A recurrent KCNK4 variant in a dominant pedigree with hypertrichosis and gingival fibromatosis syndrome: Variable phenotypic expressivity and insights on patients' dental management.

Abnormal hyperpolarization of the KCNK4 gene, expressed in the nervous system, brain, and periodontal ligament fibroblasts, leads to impaired neurotransmitter sensitivity, cardiac arrhythmias, and endocrine dysfunction, as well as, progressive cell proliferation. De novo gain of function variants in the KCNK4 gene were reported to cause a recognizable syndrome characterized by facial dysmorphism, hypertrichosis, epilepsy, intellectual/developmental delay, and gingival overgrowth (FHEIG, OMIM# 618381). FHEIG is extremely rare with only three reported cases in the literature. Herein, we describe the first inherited KCNK4 variant (c.730G>C, p.Ala244Pro) in an Egyptian boy and his mother. Variable phenotypic expressivity was noted as the patient presented with the full-blown picture of the syndrome while the mother presented only with hypertrichosis and gingival overgrowth without any neurological manifestations. The c.730G>C (p.Ala244Pro) variant was described before in a single patient and when comparing the phenotype with our patient, a phenotype-genotype correlation seems likely. Atrial fibrillation and joint laxity are new associated findings noted in our patient extending the clinical phenotype of the syndrome. Dental management was offered to the affected boy and a dramatic improvement was noted as the patient regained his smile, restored the mastication function, and resumed his psychological stability.

Antiproliferative effect of low-level laser/ photobiomodulation on gingival fibroblasts derived from calcium channel blocker-induced gingival overgrowth.

The aim of this study was to evaluate the antiproliferative properties of low-level laser therapy (LLLT) on gingival fibroblasts obtained from calcium channel blocker-induced gingival overgrowth (GO). Gingival fibroblasts of patients with GO were compared to healthy gingival fibroblasts (H). Both cells were exposed to LLLT (685 nm wavelength, 25mW power, diode laser) and compared to those not treated with LLLT. Cell proliferation and viability were measured with MTT assay at baseline and after 24 and 72 h. TGF-ß1, CTGF, and collagen Type 1 levels were evaluated with Enzyme-Linked Immunosorbent Assay (ELISA). LLLT significantly decreased the proliferation of GO fibroblasts (p < 0.05) while leading to a significantly higher proliferation in H fibroblasts compared to the untreated cells (p < 0.05). GO cells showed significantly higher CTGF, TGF-ß, and collagen Type 1 expression than the H cells (p < 0.05). LLLT significantly reduced CTGF levels in GO cells compared to the control group (p < 0.05). In H cells, CTGF and TGF-ß levels were also significantly decreased in response to LLLT compared to the control group (p < 0.05). While LLLT significantly reduced collagen expression in the H group (p < 0.05), it did not significantly impact the GO cells. LLLT significantly reduced the synthesis of the growth factors and collagen in both groups with an antiproliferative effect on the gingival fibroblasts from calcium channel blocker-induced GO, suggesting that it can offer a therapeutic approach in the clinical management of drug-induced GO, reversing the fibrotic changes.

Individualized digitally designed surgical template for guided soft tissue surgery in cases with severe gingival enlargement: A clinical application in hereditary gingival fibromatosis.

AIM: The purpose of this study was to present the use of computer-assisted periodontal surgery utilizing a novel surgical guide for cases with severe gingival enlargement through a clinical application in a patient with hereditary gingival fibromatosis. MATERIALS AND METHODS: The treatment plan included nonsurgical periodontal therapy, surgical periodontal treatment, and regular periodontal maintenance before the initiation of orthodontic treatment. Due to the increased soft tissue thickness, a surgical guide with a novel design was fabricated to facilitate the periodontal surgery since most of the patient's teeth were malpositioned and underexposed due to fibromatosis. For this purpose, the patient's intraoral scan was merged with a CBCT image in order to plan surgical excisions based on the anatomy of the teeth and the bone contour. RESULTS: The customized surgical guide facilitated the gingivectomy by controlling not only the shape of the initial incisions but also their orientation toward the level of the cementoenamel junction, improving the efficiency of the clinical time compared with freehand surgery and assisting in the verification of the final soft tissue shape, based on the treatment plan. CONCLUSION: Digital technology through the superimposition of multiple data sets can assist in the diagnosis and multidisciplinary management of cases with gingival fibromatosis. The proposed design of the surgical guide can facilitate soft tissue surgery based on the digital treatment plan, leading to more predictable management of the soft tissue, especially in patients with severe gingival enlargement, as in cases with hereditary gingival fibromatosis or drug-induced gingival overgrowth.

Cyclosporine A-induced gingival overgrowth in renal transplant patients accompanied by epithelial-to-mesenchymal transition.

OBJECTIVE: To investigate the association between the prevalence of cyclosporin A-induced gingival overgrowth and the expression of the epithelial-to-mesenchymal transition factors in the gingival tissues of renal transplant patients. BACKGROUND: Gingival overgrowth (GO) is a frequent complication in organ transplant patients treated with the immunosuppressant cyclosporin A (CsA). The epithelial-to-mesenchymal transition (EMT) is considered a factor contributing to CsA-induced GO. However, current knowledge on this topic is sparse. METHODS: Sixty-three renal transplant patients were divided into two groups according to the occurrence of GO: those with gingival overgrowth (GO+ group) and those without gingival overgrowth (GO- group). Data on age, sex, and use of immunosuppressant and calcium channel blocker medications, serum creatinine values, peak concentrations of blood CsA, and gingival hyperplasia scores were recorded to identify clinically pathogenic factors. Gingival tissues from five patients with CsA-induced GO and five healthy subjects were selected for histomorphological observation with hematoxylin-eosin staining, Masson staining, and immunohistochemical staining. The mRNA expression of EMT factors was detected with reverse transcription-quantitative PCR. RESULTS: The use of CsA significantly increased the prevalence of GO in renal transplant patients. The expression of α-SMA, SMAD4, and TGM2 was upregulated and that of E-cadherin was downregulated in the gingival tissues of patients with CsA-induced GO compared with those of the corresponding controls. CONCLUSION: Treatment with CsA is closely related to the occurrence of GO in renal transplant patients and EMT plays an important role in CsA-induced gingival tissue hyperplasia.

Enhancement of receptor-mediated calcium responses by phenytoin through the suppression of calcium excretion in human gingival fibroblasts.

BACKGROUND AND OBJECTIVES: Gingival overgrowth caused by phenytoin is proposed to be associated with Ca2+ signaling; however, the mechanisms that increase the intracellular Ca2+ concentration ([Ca2+ ]i ) are controversial. The current study aimed to elucidate the mechanism underlying the phenytoin-induced increase in [Ca2+ ]i in human gingival fibroblasts (HGFs). METHODS: Effects of 100 µM phenytoin on [Ca2+ ]i in HGFs were examined at the single-cell level using fluorescence images of fura-2 captured by an imaging system consisting of an EM-CCD camera coupled to an inverted fluorescence microscope at room temperature. RESULTS: Exposure of HGFs to 100 µM phenytoin induced a transient increase in [Ca2+ ]i in the absence of extracellular Ca2+ , indicating that the phenytoin-induced increase in [Ca2+ ]i does not require an influx of extracellular Ca2+ . In addition, phenytoin increased [Ca2+ ]i in HGFs depleted of intracellular Ca2+ stores by thapsigargin, indicating that neither Ca2+ release from stores nor inhibition of Ca2+ uptake is involved. Furthermore, the phenytoin-induced [Ca2+ ]i elevation was reduced to 18.8% in the absence of extracellular Na+ , and [Ca2+ ]i elevation upon removal of extracellular Na+ was reduced to 25.9% in the presence of phenytoin. These results imply that phenytoin increases [Ca2+ ]i of HGFs by suppressing the Na+ /Ca2+ exchanger. Suppression of intracellular Ca2+ excretion is thought to enhance the Ca2+ responses induced by various stimuli. Analysis at the single-cell level showed that stimulation with 1 µM ATP or 3 µM histamine increased [Ca2+ ]i in 20-50% of cells, and [Ca2+ ]i increased in many unresponsive cells in the presence of phenytoin. CONCLUSION: Our findings demonstrate that phenytoin induced increase in [Ca2+ ]i by the inhibition of Ca2+ efflux in HGFs. It was also found that phenytoin strongly enhanced small Ca2+ responses induced by stimulation with a low concentration of ATP or histamine by inhibiting Ca2+ efflux. These findings suggest a possibility that phenytoin causes drug-induced gingival overgrowth by interacting with inflammatory bioactive substances in the gingiva.

Drug-Induced Gingival Overgrowth-Molecular Aspects of Drug Actions.

Drug-induced gingival overgrowth (DIGO) is one of the side effects produced by therapeutic agents, most commonly phenytoin, nifedipine and cyclosporin A. However, the precise mechanism of DIGO is not entirely understood. A literature search of the MEDLINE/PubMed databases was conducted to identify the mechanisms involved in DIGO. The available information suggests that the pathogenesis of DIGO is multifactorial, but common pathogenic sequelae of events emerge, i.e., sodium and calcium channel antagonism or disturbed intracellular handling of calcium, which finally lead to reductions in intracellular folic acid levels. Disturbed cellular functions, mainly in keratinocytes and fibroblasts, result in increased collagen and glycosaminoglycans accumulation in the extracellular matrix. Dysregulation of collagenase activity, as well as integrins and membrane receptors, are key mechanisms of reduced degradation or excessive synthesis of connective tissue components. This manuscript describes the cellular and molecular factors involved in the epithelial-mesenchymal transition and extracellular matrix remodeling triggered by agents producing DIGO.

Biallelic frameshift variant in the TBC1D2B gene in two siblings with progressive gingival overgrowth, fibrous dysplasia of face, and mental deterioration.

Biallelic loss-of-function variants in the TBC1D2B gene were recently reported as a cause of a neurodevelopmental disorder with seizures and gingival overgrowth. Here, we report two male siblings with the similar clinical characteristics. They started with gingival overgrowth and bilateral growth of soft tissues in the malar region at 3 years of age, which evolved with significant maxillary hypertrophy and compression of the brainstem due to fibrous dysplasia of facial bones. After disease evolution, they presented with mental deterioration, limb tremors, and gait ataxia. One of them also presented with seizures. Whole exome sequencing revealed a novel biallelic frameshift variant [c.595del; p.(Val199Trpfs*22)] in the TBC1D2B gene in both patients, which was confirmed and found in heterozygous state in each of their parents. There are strong similarities in clinical characteristics, age of onset, and evolution between the patients described here and cases reported in the literature, including cherubism-like phenotype with progressive gingival overgrowth and seizures. This is the fourth family in the world in which a biallelic loss-of-function variant in the TBC1D2B gene is associated with this phenotype. These results support that loss of TBC1D2B is the cause of this rare condition.

Phenytoin induces connective tissue growth factor (CTGF/CCN2) production through NADPH oxidase 4-mediated latent TGFß1 activation in human gingiva fibroblasts: Suppression by curcumin.

OBJECTIVE AND BACKGROUND: Gingival overgrowth (GO) is a common side effect of some drugs such as anticonvulsants, immunosuppressant, and calcium channel blockers. Among them, the antiepileptic agent phenytoin is the most common agent related to this condition due to its high incidence. Transforming growth factor ß (TGFß) importantly contributes to the pathogenesis of GO. Connective tissue growth factor (CTGF or CCN2) is a key mediator of tissue fibrosis and is positively associated with the degree of fibrosis in GO. We previously showed that Src, c-jun N-terminal kinase, and Smad3 mediate TGFß1-induced CCN2 protein expression in human gingival fibroblasts (HGFs). This study investigates whether phenytoin can induce CCN2 synthesis through activated latent TGFß in HGFs and its mechanisms. METHODS: CCN2 synthesis, latent TGFß1 activation, and cellular reactive oxygen species (ROS) generation in HGFs were studied using western blot analysis, a TGFß1 Emax® ImmunoAssay System, and 2',7'-dichlorodihydrofluorescein diacetate (an oxidation-sensitive fluorescent probe), respectively. RESULTS: Phenytoin significantly stimulated CCN2 synthesis, latent TGFß1 activation, and ROS generation in HGFs. Addition of an TGFß-neutralizing antibody, TGFß receptor kinase inhibitor SB431542, and Smad3 inhibitor SIS3 completely inhibited phenytoin-induced CCN2 synthesis. General antioxidant N-acetylcysteine, NADPH oxidase (NOX) inhibitor diphenylene iodonium, and specific NOX4 inhibitor plumbagin almost completely suppressed phenytoin-induced total cellular ROS and latent TGFß1 activation. Curcumin dose-dependently decreased phenytoin-induced TGFß1 activation and CCN2 synthesis in HGFs. CONCLUSIONS: Our findings indicated that NOX4-derived ROS play pivotal roles in phenytoin-induced latent TGFß1 activation. Molecular targeting the phenytoin/NOX4/ROS/TGFß1 pathway may provide promising strategies for the prevention and treatment of GO. Curcumin-inhibited phenytoin-induced CCN2 synthesis is caused by the suppression of latent TGFß1 activation.

Cyclosporine-induced gingival overgrowth-Review.

Drug-induced gingival overgrowth (DIGO) is an undesirable effect resulting from the therapy of one of the three groups of drugs: phenytoin, calcium channel blockers, and cyclosporine A (CsA). It is caused by a fibrous overgrowth leading to gingivitis, periodontitis, and even tooth loss. Possible consequences include tooth decay worsening, pain and difficulty in eating, bleeding gums, and bad breath. The pathomechanism of the hypertrophy is unknown, but there is a correlation between insufficient oral hygiene and the severity of this phenomenon. The gender and age predilection of gingival hyperplasia as a result of CsA therapy is also noticeable. It is most common in children and adolescents of the male sex. The beneficial effect of the removal of tartar and local irritants in reducing the above symptoms has been demonstrated. One of the treatments for DIGO is conventional gingivectomy. The paper is a review article about cyclosporine-induced gingival hyperplasia.

Oral findings in kidney transplant children and adolescents.

BACKGROUND: Children and adolescents undergoing kidney transplantation may present oral conditions after the procedure, but a few studies have recently described them. AIM: To describe the oral conditions of post-renal transplant children and adolescents. DESIGN: Two calibrated dentists examined all the participants by assessing caries experience, enamel defects, periodontal condition and soft tissue lesions. RESULTS: A total of 120 participants were included in the study, in which 63 (52.5%) were male and 57 (47.5%) were female, with a mean age of 12.78 ± 3.9 years. Among the participants, 104 (86.7%) showed at least one oral change directly related to kidney disease. The most frequent oral findings were enamel defect (49/120; 40.8%) and drug-induced gingival overgrowth (DIGO) (20/120; 16.7%). Gingival bleeding was observed on probing in 115 (95.8%) participants, whereas 69 (57.5%) presented dental calculus and 51 (42.5%) had caries experience. CONCLUSION: Gingival bleeding, enamel defects and DIGO were the most frequent oral findings in kidney transplant children and adolescents. The use of amlodipine and anticonvulsants was associated with DIGO, and there was a positive correlation between oral ulcers and use of everolimus.

Antioxidants protect against gingival overgrowth induced by cyclosporine A.

OBJECTIVE: We investigated the importance of reactive oxygen species (ROS) on developing gingival overgrowth (GO) and then introduced the antioxidant strategy to prevent, or even reduce GO. BACKGROUND: Gingival overgrowth is a common side effect of the patients receiving cyclosporine A (CsA), an immune suppressant. Although it has been broadly investigated, the exact pathogenesis of the induced GO is still uncertain. METHODS: We cultured human primary gingival fibroblasts and used animal model of GO to investigate the ameliorative effects of antioxidants on CsA-induced GO. To examine the CsA-induced oxidative stress, associated genes and protein expression, and the overgrown gingiva of rats by using immunocytochemistry, confocal laser scanning microscopy, real-time PCR, ELISA, gelatin zymography, gingival morphological, and immunohistochemical analysis. RESULTS: We found for the first time that ROS was responsible for the CsA-induced oxidative stress and TGF-ß1 expression in human primary gingival fibroblasts, as well as the GO of rats. The antioxidants (oxidative scavenger of vitamin E and an antioxidative enzyme inducer of hemin) ameliorated CsA-induced pathological and morphological alterations of GO without affected the CsA-suppressed il-2 expression in rats. CsA-induced oxidative stress, HO-1, TGF-ß1, and type II EMT were also rescued by antioxidants treatment. CONCLUSIONS: We concluded that CsA repetitively stimulating the production of ROS is the cause of CsA-GO which is ameliorated by treating antioxidants, including vitamin E and sulforaphane. Furthermore, the immunosuppressive effect of CsA is not interfered by antioxidant treatments in rats. This finding may thus help the clinician devise better prevention strategies in patients susceptible to GO.

Gingival overgrowth induced by anticonvulsant drugs: A cross-sectional study on epileptic patients.

OBJECTIVE: Our aim was to estimate the prevalence of gingival overgrowth (hyperplasia) and to determine whether active molecules affect the severity of overgrowth in a group of epileptic patients. BACKGROUND: The effects of phenytoin on oral health have been explored in different studies, yet little information is available on other antiepileptic drugs. METHODS: Data were collected from 213 subjects of both sexes, from 5 to 80 years. Patients taking the same antiepileptic therapy for at least 1 year and meeting the inclusion criteria of the study (n = 162) were subjected to measurement of gingival overgrowth according to the modified Harris and Ewalt classification and O'Leary's plaque control record (OLR). Descriptive statistics were calculated. Data were analyzed using Pearson's r correlation coefficient and chi-square test. Significance level was set at 5%. RESULTS: The active drugs lamotrigine, oxcarbazepine, and phenobarbital were significantly associated with gingival overgrowth in 61%, 71%, and 53% of cases, respectively, and phenytoin, valproic acid, and carbamazepine in 50%, 44%, and 32% of cases, respectively. CONCLUSION: Different antiepileptic molecules may be related to gingival overgrowth. In addition to phenytoin, also lamotrigine, oxcarbazepine, and phenobarbital were associated with increased prevalence of gingival overgrowth. In the management of epileptic patients, dentists should take into account different drugs as possible causes for gingival overgrowth and warn for possible alternatives.

Role of IL-6 and STAT3 signaling in dihydropyridine-induced gingival overgrowth fibroblasts.

OBJECTIVES: This study analyzed the role of the interleukin (IL)-6/signal transducer and activator of transcription 3 (STAT3) pathway in dihydropyridine-induced gingival overgrowth (DIGO) fibroblasts. MATERIALS AND METHODS: Tissue samples were obtained through surgical dissection from five DIGO patients and five healthy individuals. Cell cultures were conditioned with nifedipine (Nif) (0.34 µM) and stimulated with IL-1ß (10 ng/ml) to clarify whether IL-6 upregulates extracellular matrix overproduction or has an impact on the cell proliferation rate of DIGO fibroblasts. STAT3 was knocked down using short hairpin (sh)RNA to determine its role in collagen (Col) type I alpha 1 (Colα1(I)) synthesis. RESULTS: Results showed that phosphorylated (p)STAT3 nuclear translocation was activated by a simulated autocrine concentration (50 ng/ml) of IL-6, and application of an anti-IL-6 antibody significantly decreased the pSTAT3/STAT3 ratio in DIGO fibroblasts. STAT3 knockdown significantly decreased STAT3 and Colα1(I) expressions in DIGO cells. DIGO tissues presented stronger proliferating cell nuclear antigen (PCNA) expression than did healthy individuals under the effect of IL-1ß/Nif treatment. CONCLUSIONS: Gingival inflammation (e.g., IL-1ß) and taking dihydropyridine (e.g., Nif) may additively stimulate Col overproduction through the IL-6-STAT3-Colα1(I) cascade in DIGO cells. IL-6-STAT3 signaling may be considered a target for the control of DIGO.

Influence of different anti-hypertensive drugs on gingival overgrowth: A cross-sectional study in a Turkish population.

OBJECTIVE: The purpose of this study was to evaluate the occurrence rate of drug-induced gingival overgrowth (DIGO) in patients treated with angiotensin-converting enzyme (ACE) inhibitors, angiotensin receptor blockers (ARBs), and calcium channel blockers (CCBs) such as amlodipine, lercanidipine, and benidipine, as well as to assess the relationship of those mentioned above with medication variables and oral hygiene. METHODS: Sociodemographic details, DIGO, and clinical periodontal parameters were obtained from one hundred and thirty-one patients receiving ACE inhibitors, ARBs, and CCBs for a period of at least 2 years. RESULTS: The occurrence rate of DIGO was 19.6% in patients using CCB, 12.5% in the ARB group, and 7.5% in the ACE inhibitor group. In a subgroup analysis of CCBs, DIGO was found to be 31.8% in the amlodipine group, 13.3% in the lercanidipine group, and 7.1% in the benidipine group. While there was a significant relationship between amlodipine drug dosage and DIGO, no association was found between the duration of therapy and DIGO in all CCB subgroups. CONCLUSION: There was no difference between the groups in terms of DIGO. Duration of therapy and drug dosage did not affect the severity of DIGO in both ACE inhibitors and ARB groups.

Hereditary gingival fibromatosis in children: a systematic review of the literature.

OBJECTIVES: Hereditary gingival fibromatosis (HGF) is an uncommon, inherited condition with slow and progressive fibrous hyperplasia of the gingiva. Due to its association with mastication, speech, and occlusion problems, early diagnosis is important. We sought to summarize the available data regarding the epidemiology, clinical characteristics, and outcomes of children with HGF (< 18 years). METHODS: A systematic literature review of the MEDLINE and Cochrane Library databases was conducted with respect to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement (end-of-search date: March 1, 2019). RESULTS: A total of 99 articles reporting on 146 patients were included. The mean age was 10.82 ± 3.93 years, and generalized gingival enlargement was seen in 97.16% (95% CI 92.69 to 99.14). Jaw, gingival, and teeth abnormalities; poor oral hygiene; eating; or speech difficulties were typical HGF-induced, while 60.90% had extraoral manifestations (95% CI 52.41 to 68.78). The disease was most commonly inherited in an autosomal dominant manner (88.41%, 95% CI 78.5 to 94.26), and about one-third of the patients had syndromic HGF (33.85%, 95% CI 23.50 to 46.00). Gingivectomy was performed in the majority of cases (91.15%, 95% CI 84.31 to 95.29), and recurrence was seen in 33.85% (95% CI 23.50 to 46.00). CONCLUSION: HGF should be suspected in children with nodularity and gingival fibrosis, teeth abnormalities, or jaw distortion. Family history can help to establish the diagnosis. CLINICAL RELEVANCE: More cases should focus on longer-term follow-up after gingivectomy as disease recurrence is not uncommon.

Comprehensive evaluation of factors that induce gingival enlargement during orthodontic treatment: A cross-sectional comparative study.

BACKGROUND: Gingival enlargement (GE) is one of the most common soft tissue problems encountered during fixed orthodontic treatment. Aims: This study aimed to evaluate the factors affecting GE in adolescents and young adults, compared with their normal peers. SUBJECTS AND METHODS: This is a cross-sectional comparative study. The sample consisted of 329 subjects (ages 10-30 years) of both genders, which was divided into four main groups: The control group (G0) with no orthodontic treatment; subjects who underwent orthodontic treatment were divided according to treatment duration into G1 (4-12 months), G2 (13-24 months), and G3 (>24 months). The clinical examinations included the level of debris, calculus (simplified oral hygiene), and GE indices. Regression analyses were used to assess the GE association in all the studied groups. RESULTS: The mean GE score increased significantly with increased treatment duration (0.42 ± 0.29 for G0 and 1.03 ± 0.52 for G3). GE scores of the lower arch were significantly higher in the anterior segment than in the posterior segment among all treatment groups. Regression analysis revealed that gender, age, oral hygiene, and treatment duration had a significant effect on GE (P < 0.05), while angle classification, overjet, overbite, treatment stage, bracket type, and therapeutic extraction did not show significant associations (P > 0.05). CONCLUSION: Gender, age, oral hygiene, and treatment duration were the most important risk factors for GE during fixed orthodontic treatment.

Biallelic loss-of-function variants in TBC1D2B cause a neurodevelopmental disorder with seizures and gingival overgrowth.

The family of Tre2-Bub2-Cdc16 (TBC)-domain containing GTPase activating proteins (RABGAPs) is not only known as key regulatorof RAB GTPase activity but also has GAP-independent functions. Rab GTPases are implicated in membrane trafficking pathways, such as vesicular trafficking. We report biallelic loss-of-function variants in TBC1D2B, encoding a member of the TBC/RABGAP family with yet unknown function, as the underlying cause of cognitive impairment, seizures, and/or gingival overgrowth in three individuals from unrelated families. TBC1D2B messenger RNA amount was drastically reduced, and the protein was absent in fibroblasts of two patients. In immunofluorescence analysis, ectopically expressed TBC1D2B colocalized with vesicles positive for RAB5, a small GTPase orchestrating early endocytic vesicle trafficking. In two independent TBC1D2B CRISPR/Cas9 knockout HeLa cell lines that serve as cellular model of TBC1D2B deficiency, epidermal growth factor internalization was significantly reduced compared with the parental HeLa cell line suggesting a role of TBC1D2B in early endocytosis. Serum deprivation of TBC1D2B-deficient HeLa cell lines caused a decrease in cell viability and an increase in apoptosis. Our data reveal that loss of TBC1D2B causes a neurodevelopmental disorder with gingival overgrowth, possibly by deficits in vesicle trafficking and/or cell survival.

Galectin-8 mediates fibrogenesis induced by cyclosporine in human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: During cyclosporine-induced gingival overgrowth, the homeostatic balance of gingival connective tissue is disrupted leading to fibrosis. Galectins are glycan-binding proteins that can modulate a variety of cellular processes including fibrosis in several organs. Here, we study the role of galectin-8 (Gal-8) in the response of gingival connective tissue cells to cyclosporine. METHODS: We used human gingival fibroblasts and mouse NIH3T3 cells treated with recombinant Gal-8 and/or cyclosporine for analyzing specific mRNA and protein levels through immunoblot, real-time polymerase chain reaction, ELISA and immunofluorescence, pull-down with Gal-8-Sepharose for Gal-8-to-cell surface glycoprotein interactions, short hairpin RNA for Gal-8 silencing and Student's t test and ANOVA for statistical analysis. RESULTS: Galectin-8 stimulated type I collagen and fibronectin protein levels and potentiated CTGF protein levels in TGF-ß1-stimulated human gingival fibroblasts. Gal-8 interacted with α5ß1-integrin and type II TGF-ß receptor. Gal-8 stimulated fibronectin protein and mRNA levels, and this response was dependent on FAK activity but not Smad2/3 signaling. Cyclosporine and tumor necrosis factor alpha (TNF-α) increased Gal-8 protein levels. Finally, silencing of galectin-8 in NIH3T3 cells abolished cyclosporine-induced fibronectin protein levels. CONCLUSION: Taken together, these results reveal for the first time Gal-8 as a fibrogenic stimulus exerted through ß1-integrin/FAK pathways in human gingival fibroblasts, which can be triggered by cyclosporine. Further studies should explore the involvement of Gal-8 in human gingival tissues and its role in drug-induced gingival overgrowth.

Elevated Interleukin-17A expression in amlodipine-induced gingival overgrowth.

BACKGROUND AND OBJECTIVES: Amlodipine, a calcium channel blocker derivative, is frequently used by patients with high blood pressure. Studies reported that it can induce gingival overgrowth. However, the underlying mechanism is not fully described yet. Interleukin-17A (IL-17A) is known as a proinflammatory cytokine, but current studies indicate that it has a role in fibrotic disorders and epithelial-mesenchymal transition (EMT). The aim of this study was to figure out the possible role of IL-17A in amlodipine-induced gingival overgrowth. MATERIALS AND METHODS: Twenty-nine (29) individuals participated in the study, and they were assigned into 3 groups based on medical status and clinical periodontal examination; 9 patients with amlodipine-induced gingival overgrowth, 11 patients with inflammatory gingival overgrowth, and 9 healthy individuals as a control group. Clinical periodontal parameters including plaque index (PI), gingival index (GI), and gingival overgrowth index (GOI) were recorded. Blood and gingival crevicular fluid (GCF) samples were obtained. Gingival tissues were taken by appropriate periodontal surgery following initial periodontal therapy. To detect IL-17A on tissue samples, immunohistochemistry (IHC) was performed. Quantitative analysis was done, and the expression level of IL-17A was given as the percent positively stained cells. Enzyme-linked immunosorbent assay (ELISA) kits were used to analyze IL-17A in serum and GCF samples. RESULTS: All recorded clinical parameters were significantly higher in gingival overgrowth groups compared with control. Evaluation of inflammation on tissue sections did not show any significant change within the groups. Immunohistochemistry findings showed that IL-17A expression was increased in amlodipine samples (81.90%) compared with control samples (42.35%) (P < .001). There was an increase in the inflammatory group (66.08%) which is significantly less than the amlodipine group (P < .05). IL-17A levels in serum and GCF samples were not different within the study groups. CONCLUSION: In this study, elevated IL-17A expression regardless of inflammation shows that amlodipine might cause an increase of IL-17A in gingival tissues. This increase might induce fibrotic changes and EMT in gingival overgrowth tissues. The association of IL-17A with fibrosis and EMT in gingival tissues requires further investigation.

Therapeutic potential of proteasome inhibitors for dihydropyridine-induced gingival overgrowth.

OBJECTIVES: NF-κB plays a crucial role in collagen overproduction in dihydropyridine-induced gingival overgrowth (DIGO) fibroblasts. We aim to investigate the role of the kappa B (IκB) kinase (IKK)-NF-κB pathway and downstream collagen type I (Col I) synthesis in DIGO cells and to demonstrate the therapeutic strategy of interference of this pathway with proteasome inhibitors. METHODS: Gingival fibroblasts from DIGO (n = 5) and healthy (n = 5) patients were selected and stimulated with IL-1ß, nifedipine, or both. All experiments were run in triplicate and independently for each primary cell sample. RESULTS: The results demonstrated that both drugs additively mediated NF-κB activity by activating IKKα/ß phosphorylation. They also triggered nuclear translocation of NF-κB, Rela, and p50 (*p < .05) and increased Col I production in both healthy and DIGO cells. The addition of proteasome inhibitors, including bortezomib and MG132, promoted the accumulation of phosphorylated p-IκBα, prevented the subsequent cytosol-to-nuclear translocation of p50 and Rela (*p < .05), and abbreviated the biosynthesis of Col I in DIGO cells. CONCLUSIONS: We suggested that IKK-IκBα activation is mediated by proinflammatory cytokines and CCBs in DIGO cells and triggers downstream NF-κB-Col I synthesis. Proteasome inhibitors may strategically interfere with the IKK-IκBα-NF-κB-Col I pathway and inhibit the etiopathogenesis of DIGO.