2003 Scientific Abstracts

1. Enamel Fluoride Penetration by a Neutral Fluoride Rinse: EPMA Study
P. VILLALTA, pvillalt@nova.edu, Nova Southeastern University, Ft. Lauderdale, FL, USA, F. GARCIA-GODOY, Nova Southeastern University, Ft. Lauderdale, FL, USA, and N. CHATTERJEE, Massachusetts Institute of Technology, Cambridge, USA
Objectives: To compare fluoride (F) penetration in sound enamel exposed in vitro to a 2% neutral sodium fluoride solution rinse.
Methods: Eight teeth were treated with one of the following F-releasing agents: Group 1 (4 teeth): Dentsply All Solutions 2 %neutral sodium fluoride solution applied for 1 minute, Group 2 (4 teeth): Dentsply All Solutions neutral sodium fluoride rinse applied for 4 minutes. The teeth were submerged and agitated in the rinse for 1 or 4 minutes. After topical application, the teeth were dried in highvacuum chamber for 48 hours, then sectioned into two halves and dry-polished with 240, 320, 400 and 600 grit SiC papers and 10,1 and 0.1 µm grit alumina-coated films. F penetration was evaluated continuously at 5 µm intervals (up to 25 µm) starting from the top of the cross-section of tooth-enamel using an electron probe x-ray micro-analyzer (EPMA) (JEOL JXA-733 Superprobe) by wavelength dispersive spectrometry. The statistical analysis used was a t-test at (P<0.05 significance level).
Results: The following values in F wt% (± SD)) were observed: 1-minute exposure 0.21 (0.04) at 5 µm, 0.20 (0.02) at 10 µm, 0.15 (0.02) at 15 µm, 0.13 (0.01) at 20 µm, and 0.12 (0.004) at 25 µm. The 4-minute exposure showed 0.38 (0.19) at 5 µm, 0.41 (0.18) at 10 µm, 0.39 (0.14) at 15 µm, 0.33 (0.07) at 20 µm, and 0.24 (0.06) at 25 µm. The results were not statistically significantly different between the enamel fluoride incorporation after 1 or 4 minute application of the 2% neutral fluoride solution at any of the depth measurements made.
Conclusions: The use of the neutral fluoride solution for 1 minute or 4 minutes had no significant difference in fluoride incorporation into the enamel.

2. DEVELOPMENT OF A MODEL TO STUDY DENTIN DEMINERALIZATION
M. TORO* and C. GONZÁLEZ-CABEZAS (Indiana University School of Dentistry, Departments of Oral Biology and Preventive Dentistry)
Root caries is a common dental problem. Evaluation of therapeutic approaches requires models for de- and remineralization of root dentin. Objectives: to develop a model to produce dentin lesions in vitro within an expected range of depth according to a specific demineralization time, and to develop a technique to quantify these lesions. Methods: Thirty-six, 4´4 mm blocks of root dentin from extracted human teeth, pre-screened by Confocal Laser Scanning Microscopy (CLSM) in order to reduce dentinal tubule variability, were selected and divided into four groups (A, B, C, and D). Group A (N=6), served as baseline. Groups B, C, and D (N=10/group) were incubated in a chemical demineralizing solution (0.1 mol/L lactic acid and 0.2 % carbopol, 50 % saturated with hydroxyapatite and 0.25 ppm NaF and adjusted to pH 5.0) for 48, 96, and 144 h, respectively. After demineralization, three sections (180 ± 20 mm thick) per specimen were obtained. Transverse microradiography (TMR) was conducted under wet conditions and mineral loss (DZ), reconstructed depth (using surrounding sound tissue to compensate for dentin shrinkage), and depth were determined. Two measurements per section were made approximately 150 mm from each edge of the lesion. For each parameter, average per specimen, maximum value per specimen, and average of the section showing the highest value were calculated. Sections were then analyzed by CLSM using a modified wet technique. Results: One-way ANOVA showed significant differences among the groups (p<0.05) for all parameters, except between groups C and D. Pearson correlation showed that CLSM (fluorescent area; FA and total fluorescence;TF) had strong correlation to TMR (range: 0.91-0.73). A significant relationship between FA and the pre-screened tubule area/count was found in specimens demineralized for 144 h. Conclusions: CLSM is an alternative method to study dentin demineralization, and this model is adequate to quantify root dentin demineralization.

3. ORAL HEALTH KNOWLEDGE AND ATTITUDE TOWARDS PREVENTION OF ORAL DISEASES AMONG PROMOTORA
S. NARENDRAN1, N.GARZA1, P. GOMEZ2. Dental Branch, University of Texas Health Science Center1 and Brownsville Community Health Science Center2.
Community Health Workers (CHW) or Promotoras have been involved in health promotion activities in developing countries as well as underserved communities in the United States, including the Texas-Mexico border communities. Because promotoras themselves are lay volunteers from the target communities, they are readily accepted as health workers by the audience. Physicians and other health care personnel have trained the CHW or promotoras in different aspects of health care such as pre- and post-natal care, diabetes, cancer, etc. Promotoras working in Brownsville along the Texas-Mexico border have had no training in oral health. We recently initiated a project to provide training on oral health promotion to Community Health Workers (CHW) or Promotoras. Before the training, we collected baseline data on the oral health knowledge, attitude and preventive practices of the participating promotoras. The study population consisted of 27 female promotoras and data were collected by a pre-tested, self-administered Spanish questionnaire. While 33.3% correctly identified the number of permanent teeth in an adult, only 11.1% knew the number of primary teeth in a child. An overwhelming majority (96.3%) was aware of the link between diabetes and oral diseases; only 33.3% was knowledgeable about the role of pit-and-fissure sealants in caries prevention. Also, 96.3% correctly identified the color of healthy gingiva, but none knew about the presence of bacteria in dental plaque. While almost all the promotoras demonstrated a positive attitude toward prevention of oral diseases, a majority of them had a positive perception about their health status. Even though participants had certain deficiencies in their dental knowledge, their positive attitude towards oral disease prevention augurs well for the success of the project.

4. DENTISTRY FOR THE HISPANIC COMMUNITY: A LOCAL SURVEY
Liliana L.C. Calkins, D.D.S.^¨
The Hispanic population has become estranged from the access and use of dental health care over the past 20 years. Searching for the cause of that estrangement, this study tested support for Spanish-speaking patients by surveying every orthodontic practice in Northern Virginia along with some General Dentistry offices in the states of Maryland, Washington D.C., and Virginia. Forty-seven percent (47%) of the surveys were returned completed. The survey measured 1) employment of Spanish speaking personnel, 2) efforts to attract and educate the Hispanic community, and 3) communication behavior of patients with limited English proficiency.¹ The primary office outreach method is hiring bilingual personnel (58%). The personnel who speak Spanish, however, are generally the staff (70%). Too few dental professionals and orthodontic specialists are capable of speaking Spanish themselves. An overwhelming 90% of the participants reported bilingual children or family members translating treatment information to the non-English speaking patient. The Hispanic community under-employs dental and orthodontic care relative to its population, its wealth, and its degree of insurance coverage. This is partly because language needs of Spanish-speaking patients are not adequately addressed by orthodontic and general dentistry offices.

5. SATISFACTION AMONG COMPREHENSIVE CARE DENTAL SCHOOL PATIENTS
Oscar Arevalo, Richard C. Weiss, Lisa P. Deem, and Thomas E. Rams, Temple University School of Dentistry, Philadelphia, PA, USA
Objectives: To assess the level and determinants of satisfaction with comprehensive dental care received at an inner-city dental school located in the mid-Atlantic region of the United States. Methods: A total of 128 dentate subjects (including 21 persons of Hispanic ethnic identification) enrolled for comprehensive care at Temple University School of Dentistry in Philadelphia completed and returned a mail questionnaire surveying their demographic characteristics and responses on a 5-point Likert scale to 23 items potentially related to patient satisfaction. Significant determinants of dental care satisfaction were identified with Fishers 2-tailed exact chi-square test and multivariate logistic regression analysis. Results: A high level of satisfaction was found, with 108 (84.4%) of the comprehensive care patients indicating an overall satisfaction with dental treatment received at the dental school. In bivariate analysis, patient gender, race/ethnicity, age, payment method (Medicaid vs. private insurance/self-pay), household income, and education were not found to be significantly related to patient satisfaction. In logistic regression modeling, three statistically significant determinants of patient satisfaction were identified: student dentist consideration of patient pain, a friendly and courteous clerical staff, and getting an appointment or being seen within a reasonable period of time. Interestingly, dental school fees, clinic hours, location, and physical plant attributes, such as cleanliness and neatness of the school, the reception area environment and seating capacity, and patient’s perceptions of the modern, up-to-date status of the school’s clinic equipment, were not significantly associated with favorable patient satisfaction responses when considered together with other dimensions of patient satisfaction. Conclusions: A high level of patient satisfaction was found at an urban-based dental school in the mid-Atlantic United States, irrespective of differences in patient demographic characteristics and methods of payment, with approximately four out of five patients expressing satisfaction with comprehensive oral health care services. Favorable patient satisfaction was significantly determined by interpersonal experiences where dental students took a patient’s pain into consideration, a friendly and courteous clerical staff was encountered, and the patient was able to get an appointment or be seen at the school within a reasonable time. These findings offer guidance for attaining patient satisfaction among persons undergoing comprehensive dental treatment at an inner-city dental school educational setting.

6. CULTURAL PERSPECTIVES, BELIEFS AND MYTHS ASSOCIATED WITH THE bIRTH OF A CLEFT LIP AND PALATE CHILD  
Gonzalez S. DDS; Da Silveira A.DDS, MS, PhD; Koerber A. DDS; Shahani M.MD; Daw J. MD; Cohen M. MD, FACS.
Culture is defined as a set of shared attitudes, values, traditions and practices that characterize a social group. These traditions and beliefs have a significant influence in the behavior of some families. The objective of this study was to investigate whether there are customs or beliefs held by individuals from different cultures, and how these beliefs affect the treatment of patients with cleft lip and palate.
The sample consisted of parents (either mother or father) or guardians of cleft lip, cleft palate and cleft lip and palate patients of all ages, who are regular patients of The Craniofacial Center. Anonymous surveys were handed out at the clinic at the time of the appointment; and parents were asked to mail back the questionnaire to the center or leave it at the reception of the clinic. 
The majority of our sample consisted of Hispanics (32%), followed by African Americans (29%) and then Caucasians (29%). A 61% of this population reported a low economic status. 59% of the people surveyed were born in the United States and 38% reported were foreigners. 79% of the parents did not know about the anomaly until they had their baby. 68% of the population stated that they did not consider other possible causes besides the ones mentioned by the physician; and 32% consider other possible causes. The cause most mentioned by this group was solar/lunar eclipse (32%), followed by God’s will (38%) and tension and emotional stress during pregnancy (15%).
Analysis of the survey results revealed the presence of certain beliefs and traditions within several populations. Knowledge of cultural beliefs could play an important role for health care providers improving communication with patients, diagnosis and treatment. 

7. INTERRELATIONSHIPS BETWEEN DIABETES MELLITUS AND PERIODONTAL DISEASES IN MEXICAN AMERICANS 
Reynaldo D. Rivera, George W. Taylor, DMD, DrPH The University of Michigan School of Dentistry
Objective: We investigated the association between glycemic control and severe periodontitis in the Mexican American and non-Hispanic white American population ages 17 years and older. Our hypothesis was that Mexican Americans with poorly controlled diabetes are more likely to have severe periodontal disease than Mexican Americans with well-controlled diabetes, non-Hispanic white Americans with poorly controlled or well controlled diabetes, and Mexican Americans and non-Hispanic white Americans without diabetes. Methods: Data on 8632 Mexican Americans and non-Hispanic white Americans ages 17 and older from the National Health and Nutrition Examination Study III were analyzed using weighted multivariable logistic regression. Severe periodontitis was defined as having at least 1 tooth with both 6 mm or more attachment loss and gingival bleeding. Individuals with fasting plasma glucose ³ 126 mg/dL were classified as having diabetes, those with fasting plasma glucose < 126 mg/dL and ³ 110 mg/dL were classified as having impaired glucose tolerance. Those with poorly controlled diabetes had glycosylated hemoglobin > 8%, and those with better-controlled diabetes had glycosylated hemoglobin = 8%. Additional variables evaluated in multivariable modeling included age, education, smoking status, poverty level, sex, and other factors derived from the interview, medical and dental examinations, and laboratory assays. Results: Mexican Americans with poorly controlled diabetes mellitus had a significantly higher prevalence of severe periodontitis than non-Hispanic whites without diabetes (odds ratio = 2.63, 95% CI: 1.14, 6.25), Mexican Americans without diabetes (odds ratio = 2.86, 95% CI: 1.32, 6.25), and Mexican Americans with impaired glucose tolerance (odds ratio = 2.94, 95% CI: 1.21, 7.14), after controlling for age, education, smoking status, and poverty level. Conclusion: These results indicate an increased probability for periodontal disease in Mexican Americans with poorly controlled diabetes and are consistent with the current body of evidence supporting an association between diabetes and periodontal disease. This study was supported by the University of Michigan School of Dentistry Student Research Program, NIDCR grant #DE007101.

8. THE DYNAMIC MECHANICAL ANALYSIS OF PORCINE TMJ DISC TISSUE AND A PROTOTYPE TMJ BIOMATERIAL
R.C. AGUIRRE* and R. BAGLEY (University of Texas at San Antonio, USA) and H.R. RAWLS (University of Texas Health Science Center at San Antonio, USA) 
If one function of the temporomandibular joint (TMJ) disc is to absorb and dissipate high frequency impulse loading during chewing, then development of an implant or tissue engineered replacement for diseased or injured discs requires an understanding of the disc's viscoelastic response to dynamic loading. Objective: To investigate the viscoelastic properties of porcine TMJ disc tissue in comparison to a vibration-damping adhesive, this has properties that may be useful for designing a TMJ-disc replacement. Methods: 2 Yorkshire hog head’s were acquired from slaughter house (Animal Tech. Co., Houston, Tx). The specimens were shipped immediately after slaughter and arrived the same day to the UTHSCSA. The TMJ discs were then extracted and immediately transferred to a protease inhibiting solution, marked, labeled, and then stored at -20ºC. The intermediate zone at the center of the articulating surface, between the temporal bone and the mandibular condyle, was the area of interest for this experiment. Disc specimens were tested on the inferior side using the dynamic mechanical analyzer (DMA 7, Perkin Elmer Co.). The DMA 7 applies an oscillatory force and records the sample’s stress-strain response. Specimens were tested in compression under a frequency scan ranging from 0.01 to 51 Hertz at 37ºC. An engineering adhesive with strong vibration-damping properties (Scotch® 467MP, 3M Corp.) was also tested using the same protocol. Results: The intermediate zone of the TMJ articular disc and the synthetic adhesive were found to have strikingly similar viscoelastic properties, with the disc being more flexible and less viscous. At chewing frequency (8 Hz), storage modulus (E') is 1.1 ± 0.5 and 2.0± 0.7 MPa for disc and adhesive, respectively; while complex viscosity (η*) is 15 ± 5 and 50 ± 15 MPa-sec, respectively. Modulus increases and viscosity decreases with frequency over the 0.01 - 50 Hz range for both disc and adhesive. Conclusion: Vibrational damping and other viscoelastic properties may be important for designing an implant or tissue engineered replacement for the temporomandibular disc.
Supported by DHHS/HRSA grant 1D34 HP04079-01

9.CELL SPREADING & INVASIVENESS OF Nf1 SARCOMA PHENOTYPES 
Vogel, K.T, Cantu, R.†(T Univ. of Texas Health Science Center at San Antonio, USA, Department of Molecular and Structural Biology, †Univ. of Texas Health Science Center at San Antonio, USA, Dental HCOE)
Neurofibromatosis Type 1 (Nf1) is a disorder characterized by neurofibromas, which are benign tumors of the skin, and by malignant peripheral nerve sheath tumors (MPNSTs). The adhesion of mouse sarcoma cells associated with Nf1 on extra-cellular matrix (ECM) molecules (Laminin (LAM), Collagen IV (COL IV), and Fibronectin (FN)) was studied, as well as the invasive phenotypes of the MPNSTs in the presence of potential chemoattractants, Fibroblast Growth Factors (FGF), or Platelet-Derived Growth Factors (PDGF). We performed Cell Spreading Assays (CSA) on the ECM molecules for each tumor-derived cell line with both 1 and 2 hr tests. Random cell counts and averages were calculated for “spreading” vs. “rounded” cells that adhered to the ECM molecules. Cells were tested on Transwell® Invasiveness Assays (TIA); FGF and PDGF were placed below the well. LAM was placed on the membrane. Cells were placed on LAM and incubated for 20 hrs. Cells that penetrated through the membrane were counted in random fields; averages were determined to indicate relative invasiveness. These results show for the first time that FGF is a chemoattractant to tumor-derived Neuron/Schwann Cell Type Tu 19-1 and Neural Crest Type Tu 8-8, and PDGF is a chemoattractant to Neuron/Schwann Cell Type Tu 19-1 only. CSAs with low averages of “rounded” cells correlated to TIAs with high levels of invasiveness, suggesting that “rounded” unbound Nf1 sarcoma cells have a greater tendency for invasiveness. CSAs with the highest averages of “rounded” cells had lower averages of invasiveness suggesting that once adhered to the ECM, the cell’s potential tendency to become “spread” increases. In TIAs, MPNSTs shown to have higher levels of invasiveness were those of neuronal or neural crest derivatives, suggesting that sarcoma phenotype may correlate with invasiveness.

10. PILOCARPINE-INDUCED ACTIVATION OF MPAKS AND SALIVARY CELL GROWTH  
Authors: Edessa Castillo*, Alan L. Lin and Chih-Ko Yeh, UTHSCSA and Geriatric Research, Education and Clinical Center, STVHCS
It has been known for more than 40 years that chronic treatment of pilocarpine can induce salivary gland enlargement, yet the mechanism by which pilocarpine induced gland enlargement has not been well understood. The purposes of this study is to examine whether activation of Beta- adernergic receptor by pilocarpine can activate MAPKs p44MAPK/p42MAPK in salivary gland cells. Another purpose is whether the activation of ERK by pilocarpine is mediated through EGF receptor. Finally, whether pilocarpine induced ERK activation is associated with salivary gland cell growth. In this experiment, cells were cultured to near confluence. The HSY cells were grown in 10% FBS DMEM medium containing certain anti-bodies. It was use for the time course of 8-chlor-cyclic AMP. HSY cells were also seeded at 10,000 cells per well using DMEM which contained 2ml of 10% FBS. These cells were used for incorporation of thymidine. The results showed that thymidine incorporation was slightly increased in the pilocarpine was slightly increased in the pilocaripine treated cells at four hours and at twenty-four hours compared to the untreated counterparts. According to the data, the peak activation by pilocarpine was observed between five and ten minutes in the time course experiment. Then the level of activation returned to basal level after 1 hour of stimulation. The Total-ERK ½ shows that the same amount of protein was added at the different time intervals. Therefore, pilocarpine can activate the MAPKs ERK ½ and has a limited effect on HSY cell proliferation.

11. EFFECT OF LASER PHOTOSENSITIZATION THERAPY ON INFECTED DENTIN TISSUE 
B. I. CORTEZ¹, B. T. AMAECHI¹ and V. C. FROHLICH² (Departments of Community Dentistry¹ and Cellular/Structural Biology², Univ. of Texas Health Science Center at San Antonio, TX, USA).
There is not yet a recognized method of confirming the presence or absence of bacteria in dentin tissue retained to avoid pulpal exposure during dental restoration. It is therefore appropriate to develop a method to ensure the sterility of the tissue prior to restoration. We utilized laser irradiation (543 nm) applied for 60 seconds as a photosensitizer to Tolonium Chloride (TC) dye (0.191 and 5 mg/ml) to explore the possibility of inhibiting the growth of cariogenic bacteria, Lactobacilli acidophilus (LA). Caries-like lesions were produced on dentin tissue of 35 human teeth using acidified gel system. Following autoclave sterilization, the teeth were assigned to seven experimental groups of 5 teeth as follows. (A) Sterile (non-infected), (B) Infected with LA only (C) Infected/TC treatment (0.191mg/ml), (D) Infected/TC treatment (5 mg/ml), (E) Infected/laser treatment, (F) Infected/laser/ TC (0.191mg/ml) treatment, and (G) Infected/laser/ TC (5 mg/ml) treatment. Following treatment, a biopsy of dentin tissue was taken from each lesion and incubated anaerobically in MRS broth at 37^O C and 5% CO₂ for 48 hours. Bacteria growth was measured by a count of colony forming unit per milliliter (cfu/ml) following plating and incubation in B12 culture agar under above condition. No growth was observed in sterile group (A) and groups (D and G) treated with 5 mg/ml TC. Duncan’s multiple comparisons (n=5, α=0.05) indicated significant (p<0.001) growth inhibition in groups C (90%) and F (97%) when compared with B (positive control). No significant effect was observed with only laser treatment. We conclude that Tolonium Chloride alone at concentration currently used for cancer detection (5 mg/ml) or at low concentration in combination with low power laser beam, applied to infected dentin inhibited the growth of cariogenic bacteria, thus highlighting its potential usefulness for treatment of this tissue prior to dental restoration. 

12. GENE EXPRESSION DURING DENTINOGENESIS 
Lizeth Dehghan,* Mary MacDougall, Shou Chen University of Texas Health Science Center at San Antonio, San Antonio, TX 78229
During tooth organogenesis reciprocal epithelial-mesenchymal interactions result in the cytodifferentiation of ectomesenchymal cells into odontoblasts which synthesize and secrete non-collagenous proteins. The conversion of predentin to dentin during dentin mineralization is involved in the complex process and implicated the non-collagenous proteins as important regulators. Objective: The aim of this study was to characterize gene expression of non-collagenous proteins during dentinogenesis. Methods: An immortalized mouse odontoblastic cell line established using retrovirus transfection was used for this study. To study gene expression during dentin mineralization, RNA and protein were isolated from cell confluency at days 1, 20 and 45. Non-collagenous genes including bone sialoprotein (BSP), dentin matrix protein1 (DMP1), dentin sialophosphoprotein (DSPP) and tuftelin were examined at transcriptional and translational levels at different time courses of cell confluence. Alkaline phosphatase (ALP) expression and von Kossa analysis were also performed. Results: ALP expression and von Kossa staining exhibited the highest levels at day 45 of odontoblast confluence. Semi-quantitative RT-PCR studies revealed that expression of DSPP, BSP, and tuftelin genes was increased with cell mineralization while DMP 1 expression was the highest level at day 20 of cell confluence, then decreased at day 45 during odontoblast mineralization. The protein expression levels were coincided with RNA expression. Conclusions: expression of non-collagenous genes tested was increased during odontoblast mineralization. This study indicates that non-collagenous genes play an important role during dentin mineralization. 
(This study was supported by NIDCR Research Grants P01DE13221 and by UTHSCSA Dental School Hispanic Center of Excellence)

13. Ectopic Exposure of Ameloblast Cells to BMP4 Induces Apoptosis
M. GARCIA*, B. HERMAN and M. MACDOUGALL (Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX, USA).
Ameloblasts undergo distinct developmental phases of cytodifferentiation, matrix secretion, and maturation and then they are removed by physiological cell death (apoptosis) before tooth eruption. Earlier studies have shown that during the maturation stage of amelogenesis (enamel growth) apoptosis causes a 50% reduction in the size of the ameloblast population. Bmp4 is expressed in the oral epithelium and has been identified as a signaling molecule involved in tooth development. In a transgenic mouse model, over-expression of Bmp4 driven by a specific portion of the Bmp4 promoter in epithelium-derived ameloblasts resulted in profound tooth abnormalities. The mechanisms by which programmed cell death is spatially regulated are not well characterized. To investigate the role of Bmp4 in regulating apoptosis in ameloblasts; we first established that Bmp4 does induce cell death in a dose-dependent manner. We then sought to determine that apoptosis was the mechanism of cell death and not necrosis. We used Fluorometric Assays to detect increases in the enzymatic activity of caspases, which are proteases known to be involved in both the initiation and execution of apoptosis. These caspases fall into two classes, initiators and executioners and are differentially activated in the intrinsic vs. extrinsic apoptotic pathways. Compared to untreated cells, there was a significant increase in the enzymatic activity of caspases 3, 6, 8, and 10 in the cells treated with Bmp4. These results confirm that Bmp4 induces apoptosis in ameloblast cells and provides evidence that Bmp4 induces apoptosis through the extrinsic pathway. This study was supported by T32 DE14318 (D-Star) and NIH/NIGMS 5R25GM60655 (MBRS-RISE program).

16.  DETERMINING WHETHER A MODIFIED WORKING TIME TEST WITH NON-PUTTY MATERIALS CAN BE ACCOMODATED BY ISO STANDARD 4823  
M. Jusino, R. Wagner, Mentor – Dr. R. Baez, (The University of Texas Health Science Center at San Antonio, San Antonio, TX)
It is believed that the manufacturer’s stated working time for dental impression materials are not accurate. The aims of the study are: (1) determine whether the working time test procedure described in ISO 4823 can be modified such that the non-putty materials can be accommodated by the ISO standard 4823; (2) to verify a simpler test for determining the working time is reliable worldwide and clinically relevant. Six non-putty materials were evaluated using three different tests: the Rheometer 0.25mm (Standard test) and 1mm displacement test and the Penetrometer test. For the displacement tests, two plates were used: the standard’s plate and one with larger perforations, since it’s believed that by having a wider diameter perforation in the plate, the material is less likely to brake upon displacement. The 6 materials involved were: Reprosil Medium, Low and Very High Viscosity, Impregum Penta and Kerr Permlastic Regular and Light Body. For the Penetrometer test the mean, standard deviation and coefficient variation were calculated for the 6 materials. A t-test was performed for the 0.25mm displacement test between both plates; only three of the materials studied had significant difference. From the three tests, the 0.25mm displacement test was the only test that provided a more accurate working time reading. The 1mm displacement and the penetrometer do not seem to be clinically relevant working time tests for elastomeric materials because the results do not provide a reading for its working time. But both can show that the material is within the stated working time.

17. Reduced Leakage in Low Cure-Shrinkage Liquid Crystal Dental Restorations
C.J. MARTINEZ*, K. SPARKMAN, B.R. FURMAN, N. SATSANGI, H.L.CARDENAS, & H.R. RAWLS (University of Texas Health Science Center at San Antonio, San Antonio, TX )Objective: To overcome the problem of leakage at cavity margins caused by excessive cure shrinkage, this study investigated microleakage adjacent to experimental low-shrinkage liquid crystal monomer (LCM) composites (IADR Abst.119, 2001) compared to that of conventional composites. Methods: Diacrylate LCM with either 60% w/w (flowable consistency) or 70% (stiff paste) 0.7mm silanated barium glass (Schott Glass USA) were compared to commercial composites with comparable filler loading and consistencies (Filtek^® Flow and Z100^®, 3M-ESPE). Degree of monomer-to-polymer conversion (DC) was determined to insure that all were adequately cured to about the same level. Specimens were placed between NaCl plates and cured at intervals up to 120s with a quartz-tungsten lamp (Optilux 401, Demetron) set at 400 mW/cm². An IR spectrum was recorded (Midac FTIR spectrometer), and DC determined from the ratio of the 1638 cm^-1 band (alkene double bonds, which decrease with DC) to the1608 cm^-1 band (aromatic double bonds, which do not change). For the leakage test, 23 bovine incisors were divided into four groups. Either one or two cavities, 2mm diameter x 2mm deep, were cut into each tooth, giving 10 cavities per group. The commercial resins served as comparative controls. All groups were placed, cured 120s, thermocycled 1000 times between 5°C and 55°C in water to simulate about one year in the mouth, and soaked in silver nitrate in the dark, followed by photo developer with room illumination, both overnight. Teeth were cut mesial-distal through the restorations using a dental lathe and a serrated cutting disk, and then rated for leakage under 10x magnification (Nikon HFX) using a 0-4, 5-point rating scale. Results: Mean leakage ratings (± std dev) were Z100 (2.0±1.4) > Filtek (1.4±1.4) = LCM70% (1.2±0.7) = LCM60% (1.0±0.9). DC at 120s was: Z100 = 58%, Filtek = 56%, LCM70% = 74%, LCM60% = 88%. ANOVA indicated that Z100 leakage was greater than for the other materials, which did not differ among themselves. All samples cured adequately and the LCM composites reached a higher DC than the two 3M composites. Conclusions: A trend was seen in which composites with higher filler have greater leakage. This has been attributed to higher modulus (rigidity) leading to greater stress build up as cure shrinkage progresses (IADR Abst. 3969, 2002). This effect is reduced in the LCM composites. Thus, low shrinkage liquid-crystal resins reduce marginal leakage. This advantage is greatest in highly filled composites such as posterior resins. Supported by the Dental Hispanic Center of Excellence, UTHSCSA, and NIH P01 DE11688.

18. CREATION OF A FOOD FREQUENCY LIST
E. Reifsnider, Phd., RN and R. E. Mora (University of Texas at San Antonio, USA)
The purpose of this project was to determine what foods are eaten most frequently by low- income Hispanic children who are enrolled in a Special Supplemental Nutrition Program for Women, Infants and Children (WIC) clinic in San Antonio. The long-term outcome will be the creation of a culturally sensitive food frequency questionnaire appropriate for this population. A secondary analysis of data collected during two studies of child growth patterns, both of stunted growth and overweight, comprised the data available for analysis. The children were all of Hispanic ethnicity, between the ages of 1 and 2. The data that was analyzed comprised 375 copies of 24-hour diet recalls, taken from the children's mothers during their WIC certifications. The individual foods were recorded, entered into an Excel document and further refined into a frequency list of food eaten by this sample. The results demonstrate that the foods consumed by these children are characterized by low fiber, high calorie and high carbohydrate contents.

19. Effects of Ethanol on Signal Transduction Pathways
Andrea Patterson*,Chakavak Farangi, James Roberts, Benxu Cheng (University of Texas at San Antonio, USA)
Scientists develop pharmaceuticals to manipulate signal transduction pathways, turning on or off genes to produce new strategies to tackle diseases. When testing pharmaceuticals, ethanol is often the vehicle used to deliver the treatment. Knowing this brings about the question—does ethanol itself have an effect on the signaling pathways?
GT1-7 and AtT-20 cell lines were used in these studies. GT1-7 cells are hypothalamic neurons, responsible for Gonadotropin Releasing Hormone (GnRH) secretion. AtT-20 cells are mouse pituitary tumor cells, responsible for Adenocorticotropin (ACTH) secretion. The main signaling pathways examined were the pathways which regulate GnRH and ACTH gene expression, MAPK (Erk), Jak/Stat (Stat-3), and PI3K (Akt).
A time course and concentration experiment were analyzed. The time course experiment looked at the effects of 0.1% ethanol at varying times. The concentration experiment looked at the effects of varying concentrations of ethanol over a ten-minute period.
In the AtT-20 time course experiment, 0.1% ethanol stimulated Erk phosphorylation and inhibited Stat-3 phosphorylation. The GT1-7 time course experiment found 0.1% ethanol to stimulate both Erk and Stat-3 phosphorylation. Effects of ethanol were seen very quickly in GT1-7 cells; by ten minutes of ethanol exposure a profound amount of phosphorylation was recognizable. AtT-20 cells showed effects of ethanol later, by thirty minutes of exposure. Akt was unaffected by the ethanol exposure, showing no major increase or decrease in phosphorylation. The concentration experiment was designed to look further into the results of GT1-7 time course experiment. The GT1-7 time course experiment showed significant effects of ethanol at ten minutes, so in the concentration experiment GT1-7 cells were exposed to varying concentration of ethanol over a ten-minute period. An increase in Erk and Stat3 phosphorylation was apparent with 0.05% ethanol.
These studies do provide evidence to conclude that ethanol itself does affect signal transduction pathways, and does so differently in different cell lines. The results obtained by these experiments will be helpful in designing an experiment in which ethanol is used as the solvent for the pharmaceutical being tested. 

20. Integrins May Modulate Opiod Receptor Sensitivity in Trigemial Ganglion
A. Salcido and Gustovo Zardeneta, Department of Oral and Maixllofacial Surgery, (University of Texas Health Science Center, San Antonio, TX USA)
Integrins are receptor proteins that serve as the main way cells bind to and respond to the extracellular environment.  Functional integrins are composed of two transmembrane glycoprotein subunits and. When a ligand binds, the integrins cluster in the membrane and bind to various intracellular proteins forming a focal adhesion.  The role of integrins in the development of neurons has been documented but little is known concerning adult sensory neurons. This work shows the link between integrins and the Erk/MAP kinase signaling cascade in mature sensory neurons.  Preliminary evidence suggests that integrin ligands, specifically RGD containing proteins bind to integrins and effect opioid receptors in adult trigeminal ganglion neurons. This study builds on the proposed Erk pathway cascade leading to the desensitization of the opioid receptor via Grk2  upon integrin activation. Activation of integrins on rat trigeminal ganglion nerve cells was accomplished by adding a integrin blocking antibody to the cell culture. Once the integrin was activated, a secondary species specific antibody was applied to link the primary antibodies which cause receptor clustering or focal adhesion formation. In comparison to the controls, the ELISA showed that the primary/secondary antibody treated cells showed an increase of approximately 27% in activation of Erk (i.e..Phosphorylated Erk) demonstrating successful clustering. Pre-treatment with l/ml of bradykinin, a plasma protein with inflammatory effects, followed by antibody induced clustering produced an increase in phosphorylated Erk of about 12% higher than the primary/secondary antibody treated cells. Western blots from cell lysates of these experiments were carried out to show changes in the phosphorylation Erk and Grk2 so that the link between integrin stimulation and opioid receptor sensitization could be confirmed

21. Mathematical Relation Between Clinical and Laboratory Caries Assessment Methods
V.M. SALDAÑA¹* and B.T. AMAECHI² (¹Univ. of Texas at San Antonio, TX, USA, ²Dept. of Community Dentistry, Univ. of Texas Health Science Center, San Antonio, TX 78229, USA).
Objective: Diagnodent™ is an established method for clinical assessment of caries; however, it does not give the actual value for the mineral content of tooth tissue. There is a need to establish a relationship between Diagnodent measurement and that of a device which can measure the mineral content of the caries lesion, to facilitate monitoring of the mineral status of a lesion and/or the effect of an applied therapeutic agent. Micro-Computerized Tomography (MicroCT40™) is currently used in the laboratory to non-invasively quantify the mineral density of a calcified tissue. It measures the linear attenuation coefficient of the tissue and relates it directly to its mineral density. This study aimed to establish a mathematical equation relating MicroCT and Diagnodent, for use in evaluating the mineral status of a caries lesion by a combined use of the two systems. Methods: Using MicroCT and Diagnodent, the baseline mineral status of defined spots on the enamel surface of twenty extracted human teeth were measured and the data recorded. Caries-like lesions were created on those spots by demineralization in acidified gel system. The spots (lesions) were then re-assessed with the two systems. Results: In either systems, paired t-tests (a=0.05) indicated a significant difference between the pre- and post-demineralization data (n=20, p<0.001). Using Linear Regression coefficient and squared Pearson correlation coefficient (r²), a linear correlation (regression equation: µCT = 3.63 + 0.08Dd) was observed between the measurements of MicroCT (µCT) (range 3.0 – 4.62 cm^-1) and that of Diagnodent (Dd) (range 4 – 9) for the post-demineralization (caries lesion) measurements. Conclusion: We conclude that the equation established in this study can be used to extrapolate the mineral density in a caries lesion detected and measured using Diagnodent, and may be useful for monitoring the progress of an early caries being treated with a therapeutic agent such as fluoride. 

22. A Summary of Recent Evidence for Enhanced Cavity Protection: Reaching the Underserved Hispanic Minority with RecaldentÔ Containing Chewing-Gum
Holme, Samantha: Cadbury-Adams, Morris Plains, NJ – Samantha.Holme@am.csplc.com
Cadbury-Adams, maker of Trident Sugarless Gum, is deeply committed to enhancing access to quality health care for those most in need, particularly the urban and rural poor.
In its long tradition of promoting dental health, Trident has developed a program – Trident Cares - designed to help improve the oral health of children in underserved communities throughout the country. For 2003, Trident has announced a partnership with the Hispanic Dental Association (HDA) to conduct Trident Cares clinics in Philadelphia, San Antonio and San Diego. These clinics will run throughout the month of October and will bring free preventative dental care to 1,000 needy children as well as help educate them about the importance of a proper dental routine. Participating children and their families will receive important information on dental care and how to protect their smiles, as well the “Trident Smile Care Kit,” which will include a toothbrush, floss, toothpaste and Trident gum. Trident plans to significantly increase our investment and to expand Trident Cares to additional communities throughout the U.S. in 2004.
Further in 2004, Trident will sponsor ten Cadbury-Adams Community Outreach Scholarships through the HDA to aid in the cost of dental or dental hygiene school for qualified applicants.
In addition to the outreach programs described above, Cadbury-Adams has developed sugarless chewing gums containing RecaldentÔ (casein phosphopeptide-amorphous calcium phosphate; CPP-ACP) to help encourage tooth remineralization after eating. RecaldentÔ is derived from milk proteins and, because of the peptides shape, contains calcium phosphate in an amorphous state. Three papers have been published in the Journal of Dental Research supporting the remineralizing efficacy of RecaldentÔ and another has been submitted to the Australian Dental Journal.
The first publication¹ detailed a study of solutions of CPP-ACP in preventing caries in rats. This was a standard rat caries model with specific-pathogen-free rats inoculated with Strep. sobrinus. The rats were fed cariogenic diets free of dairy products and the molars treated twice a day with solutions of CPP-ACP (0.1, 0.2, 0.5, 1.0% w/v). A 500-ppm fluoride solution was included as a control. The CPP-ACP solutions decreased both smooth surface and fissure caries in a dose response fashion, with the 0.5 and 1.0% CPP-ACP being comparable to the fluoride control. 
The second publication² detailed the efficacy of CPP-ACP in sugarless chewing gum using an intraoral in-situ model. This was a double-blind crossover design where the subjects were asked to wear a palatal appliance embedded with 6 demineralized human tooth half-slabs for 20 minutes of gum chewing plus 20 after four times per day for 14 days. The slabs were then removed, paired with their demineralized control half-slab, embedded in acrylic resin, sectioned and subjected to microradiography. The addition of CPP-ACP to a sugarless gum increased remineralization in a dose response fashion. The gums containing 0.19, 10.0 18.8 and 56.4 mg of CPP-ACP produced an increase in enamel remineralization of 9, 63, 102, and 152%, respectively, relative to a sugarless control gum not containing CPP-ACP.
The third publication³ detailed clinical studies showing the retention of the CPP-ACP in the plaque layer for up 3 hours post gum chewing. These studies compare the ability of CPP-ACP, with that of other sources of calcium, to be retained in the supragingival plaque and remineralize enamel subsurface lesions in-situ when delivered via a mouthrinse or a chewing gum in randomized double-blind trials. In the mouthrinse study, only the CPP-ACP-containing mouthrinse significantly increased plaque calcium and inorganic phosphate levels. In the chewing gum studies, the gum containing CPP-ACP, although not the gum containing the most calcium per piece, produced the greatest amount of enamel remineralization of the enamel half-slabs in-situ. The CPP-ACP could also be detected in plaque extracts for up to 3 hours after chewing. This is an important finding because it indicates that a “reservoir” of bioavailable calcium remains available at the tooth surface should the demineralization process begin.
An additional paper has been submitted for publication in the Australian Dental Journal detailing the results of a randomized, double blind in-situ study of CPP-ACP containing sugarless pressed mint tablets to remineralize enamel half-slabs with subsurface lesions. The subjects allowed the tablets to dissolve and then continued to wear the appliance for an additional 20 minutes, four times per day for 14 days. The tablets containing 18.8 and 56.4 mg of CPP-ACP were significantly better (p < 0.01) at remineralizing subsurface lesions compared to no treatment and a sugarless tablet without CPP-ACP.
References:
1. Reynolds E.: Anticariogenicity of calcium phosphate complexes of tryptic casein phosphopeptides in the rat. J Dent Res 74(6): 1272-1279; June 1995.
2. Shen P.; Cai F.; Nowicki A.; Vincent J.; Reynolds E.: Remineralization of enamel subsurface lesions by sugar-free chewing gum containing casein phosphopeptide-amorphous calcium phosphate. J Dent Res 80(12): 2066-2070, 2001.
3. Reynolds E.; Cai F.; Shen P.; Walker G.D.: Retention in plaque and remineralization of enamel lesions by various forms of calcium in a mouthrinse or sugar-free chewing gum. J Dent Res 82(3): 206-211, 2003.