Reversal of drug-induced gingival overgrowth by UV-mediated apoptosis of gingival fibroblasts - an in vitro study.

Gingival overgrowth (GO) is an undesirable result of certain drugs like Cyclosporine A (CsA). Histopathology of GO shows hyperplasia of gingival epithelium, expansion of connective tissue with increased collagen, or a combination. Factors such as age, gender, oral hygiene, duration, and dosage also influence onset and severity of GO. One of the mechanisms behind uncontrolled cell proliferation in drug-induced GO is inhibition of apoptotic pathways, with a consequent effect on normal cell turnover. Our objective was to determine if UV photo-treatment would activate apoptosis in the gingival fibroblast component. Human gingival fibroblast cells (HGF-1) were exposed to 200ng/ml or 400ng/ml CsA and maintained for 3, 6, and 9 days, followed by UV radiation for 2, 5, or 10min (N=6). Naïve (no CsA or UV), negative (UV, no CsA), and positive controls (CsA, no UV) were designated. Prior to UV treatment, growth media was replaced with 1M PBS to prevent absorption of UV radiation by serum proteins, and cells were incubated in growth media for 24h post-UV before processing for TUNEL assay, cell proliferation assays, or immunofluorescence. Data showed a temporal increase in proliferation of HGF-1 cells under the influence of CsA. The 200ng/ml dose was more effective in causing over-proliferation. UV treatment for 10min resulted in significant reduction in cell numbers, as evidenced by counts and proliferation assays. Our study is a first step to further evaluate UV-mediated apoptosis as a mechanism to control certain forms of GO.

Cancer Secretome May Influence BSP and DSP Expression in Human Salivary Gland Cells.

One of the biggest challenges in managing head and neck cancers, especially salivary gland cancers, is the identification of secreted biomarkers of the disease that can be evaluated noninvasively. A relevant source of enriched tumor markers could potentially be found in the tumor secretome. Although numerous studies have evaluated secretomes from various cancers, the influence of the cancer secretome derived from salivary gland cancers on the behavior of normal cells has not yet been elucidated. Our data indicate that secretome derived from salivary gland cancer cells can influence the expression of two potential biomarkers of oral cancer-namely, bone sialoprotein (BSP) and dentin sialoprotein (DSP)-in normal salivary gland cells. Using routine immunohistochemistry, immunofluorescence, and immunoblotting techniques, we demonstrate an enrichment of BSP and DSP in human salivary gland (HSG) cancer tissue, unique localizations of BSP and DSP in HSG cancer cells, and enriched expression of BSP and DSP in normal salivary gland cells exposed to a cancer secretome. The secretome domain of the cancer microenvironment could alter signaling cascades responsible for normal cell proliferation, migration, and invasion, thus enhancing cancer cell survival and the potential for cancer progression. The cancer secretome may be critical in maintaining and stimulating "cancer-ness," thus potentially promoting specific hallmarks of metastasis.

Variant Inferior Alveolar Nerves and Implications for Local Anesthesia.

A sound knowledge of anatomical variations that could be encountered during surgical procedures is helpful in avoiding surgical complications. The current article details anomalous morphology of inferior alveolar nerves encountered during routine dissection of the craniofacial region in the Gross Anatomy laboratory. We also report variations of the lingual nerves, associated with the inferior alveolar nerves. The variations were documented and a thorough review of literature was carried out. We focus on the variations themselves, and the clinical implications that these variations present. Thorough understanding of variant anatomy of the lingual and inferior alveolar nerves may determine the success of procedural anesthesia, the etiology of pathologic processes, and the avoidance of surgical misadventure.

The gross anatomy laboratory: a novel venue for critical thinking and interdisciplinary teaching in dental education.

Reports on the status of dental education have concluded that there is a need for various types of curricular reform, making recommendations that include better integration of basic, behavioral, and clinical sciences, increased case-based teaching, emphasis on student-driven learning, and creation of lifelong learners. Dental schools faced with decreasing contact hours, increasing teaching material, and technological advancements have experimented with alternate curricular strategies. At Southern Illinois University School of Dental Medicine, curricular changes have begun with a series of integrated biomedical sciences courses. During the process of planning and implementing the integrated courses, a novel venue-the gross anatomy laboratory-was used to introduce all Year 1 students to critical thinking, self-directed learning, and the scientific method. The venture included student-driven documentation of anatomical variations encountered in the laboratory using robust scientific methods, thorough literature review, and subsequent presentation of findings in peer review settings. Students responded positively, with over 75% agreeing the experience intellectually challenged them. This article describes the process of re-envisioning the gross anatomy laboratory as an effective venue for small group-based, student-driven projects that focus on key pedagogical concepts to encourage the development of lifelong learners.

Basal membrane complex architecture is disrupted during posterior subcapsular cataract formation in Royal College of Surgeons rats.

PURPOSE: Previous studies detailing the development of posterior subcapsular cataracts (PSC) in Royal College of Surgeons (RCS) rats have shown that aberrant fiber-end migration underlies the structural compromise. This investigation was conducted to examine the distribution of select basal membrane complex (BMC) components and to assess the intravitreal levels of specific cytokines during PSC formation. METHODS: Lenses from 52 RCS dystrophic rats (RCS/Lav) and 28 genetically matched control animals (RCS-rdy(+)/Lav) from 2 to 8 weeks old were used. After enucleation, vitreous was collected for eventual cytokine level analyses; lenses were then removed and processed for immunocytochemical localization of actin, cadherin, ß integrin, vinculin, and cell nuclei. RESULTS: At 2-3 weeks postnatal, dystrophic lenses showed normal BMC distribution of actin, cadherin, and vinculin; however ß integrin distribution was altered as compared to controls. By 4-6 weeks of age, F-actin was visible as bright foci arranged in a "rosette" pattern around fiber-end profiles. Concurrently, vinculin was rearranged into a diffuse pattern within the BMC. Cadherin delineated the fiber ends in dystrophic lenses until 5 weeks postnatal, after which it displayed diffuse cytoplasmic staining with more definitive labeling at the BMC periphery. ß integrin was initially distributed as punctuate spots at 2-3 weeks postnatal; however, by 4-6 weeks it was co-localized with F-actin around the periphery of fiber ends. The distribution of F-actin, cadherin, and ß integrin components did not undergo further changes after 6 weeks of age; however, vinculin was present predominantly at the periphery of the BMC in 7-8-week-old dystrophic lenses. Intravitreal cytokine levels were assessed for interleukin (IL)-1α, IL-4, IL-6, IL-8, tumor necrosis factor (TNF), and interferon (IFN)-γ. Levels of IL-1α, IL-4, TNF, and IFN-γ demonstrated a similar pattern, with concentrations increasing from 2 to 6 weeks postnatal and then decreasing slightly up to 8 weeks of age. IL-4 and TNF had the highest average concentrations, with peaks of 148.00 pg/ml and 34.20 pg/ml, respectively. CONCLUSIONS: The data indicate that defined rearrangements of normal BMC architecture precede and characterize the structural changes that culminate in the PSC. These are consistent with modifications of adhesion mechanics involving cell-cell attachment, cell-matrix adhesion, and timely fiber-end detachment. Further, the results suggest that pro-inflammatory cytokines are potential initiating factors in aberrant fiber-end migration and subsequent PSC formation.

Abnormal fiber end migration in Royal College of Surgeons rats during posterior subcapsular cataract formation.

PURPOSE: Prior structural studies of posterior subcapsular cataract (PSC) development in Royal College of Surgeons (RCS) rats suggest that migration of basal fiber ends was disrupted, ultimately resulting in a PSC. Therefore the goal of this study was to assess the overall migration patterns as well as changes to the structure and cytoskeleton of basal fiber ends during PSC development. METHODS: Lenses from 48 RCS dystrophic rats (RCS/Lav) and 24 genetically matched control animals (RCS-rdy(+)/Lav) from 2 to 8 weeks old were examined. Equatorial diameters were measured and suture patterns were photographed immediately following enucleation/dissection. Right eye lenses were fixed and processed to visualize the actin cytoskeleton via laser scanning confocal microscopy (LSCM), left eye lenses were decapsulated, fixed and processed for scanning electron microscopy (SEM). Scaled 3D-computer assisted drawings (CADs) and animations were constructed from the data to depict the changes in suture patterns and fiber end architecture. RESULTS: At 2 weeks, dystrophic lenses displayed an inverted Y suture on the posterior, and by 3 weeks most lenses had at least one sub-branch. Additional sub-branches were observed with time, opacities being visible as early as 4 weeks and progressing into PSC plaques by 6 weeks. Control lenses displayed inverted Y sutures at all ages and were transparent. SEM of dystrophic lenses revealed fiber ends with normal size, shape, arrangement, and filopodia at 2 weeks; scattered areas of dome-shaped fiber ends and small filopodia were present at 3 weeks. At 4 weeks the irregularly arranged domed fiber ends had extremely long filopodia with 'boutons' at their tips. By 6 weeks all fiber ends within plaques displayed rounded or domed basal membranes and lacked filopodial extensions. Control lenses at all time points had comparable ultrastructure to the 2 week old dystrophic lenses. F-actin arrangement within the basal membrane complex (BMC) of control lenses showed the expected peripheral pattern of labeling at all ages. Dystrophic RCS lenses at 2 weeks were comparable to controls, however by 3-4 weeks they displayed scattered foci of F-actin within the BMC. At all time points thereafter, F-actin was rearranged into a 'rosette' pattern of prominent foci at cell vertices. CONCLUSIONS: The data are consistent with the hypothesis that migration of basal fiber ends is altered in a two stage process wherein initially, migration patterns undergo a rapid shift resulting in abnormal suture sub-branch formation. Subsequent cytological alterations are consistent with an eventual cessation of migration, precluding proper targeting of basal ends to their sutural destinations and leading to cataract plaque formation.

Aberrant basal fiber end migration underlies structural malformations in a streptozotocin-induced diabetic rat model.

This study characterized early structural changes at posterior fiber ends in the crystalline lens after diabetic induction. Wistar rats (n = 49), randomized into one naïve control group and four experimental groups, were rendered diabetic via streptozotocin injection. Animals were euthanized at 1 week intervals, blood glucose levels recorded and lenses were evaluated grossly, by SEM and by confocal microscopy. Scoring Indices were developed to assess structural alterations and for statistical correlations between the scores and the duration of hyperglycemic exposure as well as blood glucose levels. Average blood glucose levels increased progressively from 98.5 mg/dL (controls) to 331.4 mg/dL (4 weeks). Diabetic lenses displayed abnormal suture sub-branches and opacity formation beginning late in the first week post-injection and rapidly progressing in severity through four weeks. SEM analyses showed gradual elongation of fiber ends and filopodia which comprised a disorganized configuration and a loss of recognizable migration patterns. Structural alterations culminated in foci of fiber degeneration by the third to fourth weeks. The F-actin distribution at basal fiber ends was significantly altered as compared to naïve controls. Cadherin distribution was altered as compared to controls, but largely at later time points. The grading system clearly shows increased structural compromise with elevated blood glucose levels in streptozotocin-induced diabetes. Further, although the initial rise in blood glucose levels was associated with pathological changes, their progression depended to a larger extent on continued hyperglycemic exposure. The data suggests that hyperglycemia initially disrupts fiber end migration, resulting in structural alterations and eventual fiber degeneration.