Evaluation of Anticancer Potential of Flavones from Rhamnus alaternus against B16F10 Melanoma Cells.

Melanoma has become an important health problem and new treatment have become an imperative medical need. Therefore, the finding and identification of natural product with less toxic effects, capable of promoting melanoma cell death have become an important goal of research in oncotherapy. In this study, we want to investigate the anticancer activity of an enriched total oligomers flavonoids (TOF) extract of R. alaternus in melanoma cells. First, TOF was exhibited to be rich in flavones. We revealed that this extract reduced proliferation and increased of sub-G1 and S phase cells built-up in B16-F10 cells in a dose-related manner. Moreover, In Vivo, TOF reduced tumor volume and weight with percentages of inhibition of 92.4% and 92.9%, respectively. R. alaternus was also found to be effective in reducing the level of pro-inflammatory cytokine IL-6 during metastasis. Level of TH1 cytokine, such as IL-2, was significantly enhanced by TOF treatment. Indeed, the histological examination of the tumor revealed the absence of mitoses and the presence of numerous melanin pigmented macrophage cells in the R. alaternus extract-treated group that could be explained by the induction of macrophage activation and by the arrest of the cell cycle in the Sub-G1 and S phases.

Amelogenesis imperfecta with Class III malocclusion, reduced crown size and decreased OVD: A multi-disciplinary management and a 5-year follow-up.

This clinical report describes the oral rehabilitation of a 22-year-old-man diagnosed with a variant of hypoplastic amelogenesis imperfecta. The treatment approach was multi-disciplinary, and it included the surgical procedure of Lefort I osteotomy, surgical crown lengthening, and metal-ceramic-fixed dental prostheses. The patient was satisfied with the esthetic and functional outcome.

Effect of xylitol chewing gum enriched with propolis on dentin remineralization in vitro.

OBJECTIVES: The aim of this study was to evaluate the efficiency of xylitol chewing gums enriched with propolis, remineralizing softly demineralized dentin in vitro. DESIGN: Four groups of chewing gum were developed; Group1: xylitol (1.8 %), Group2: xylitol + casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) (3%), Group3: xylitol+Hydroxyapatite (3%) and Group4: xylitol + propolis (5%). A control group was designed without chewing gum, but with artificial saliva. Sections of embedded crowns and cleaned roots of twenty five bovine incisors were demineralized in carbonated drink. Crown specimens were half-varnished. Remineralization process was run for all the dental specimens in the 4 groups with gum extracts and in the control group with artificial saliva for 20 min at 37 °C three times a day during 7 days. Mineral contents were evaluated by scanning electron microscopy with energy dispersive X-ray spectroscopy (EDX-SEM). Surface morphology and roughness were analyzed using atomic force microscopy (AFM). Micro-hardness was measured using Vickers micro-hardness tester among varnished and unvarnished sides. RESULTS: Calcium/Phosphate mean ratio showed a significant decrease between the control group, group1, group2 and group4. Control group and group3 were not significantly different. Micro-hardness increased significantly for all treated groups. AFM showed obstruction of dentinal tubules in all the groups and roughness decreased in the treated side of the dentin compared to the untreated side for tested groups. CONCLUSION: Xylitol chewing gum enriched with propolis showed dentinal tubules occlusion, significant improvement of micro-hardness and slight decrease in roughness. Ca/P ratio analysis suggests that a mineral compound other than hydroxyapatite is responsible of tubules occlusion.

Dental caries and hypoplastic amelogenesis imperfecta: Clinical, structural, biochemical and molecular approaches.

AIM: The aim of this study was to explore the caries features in hypoplastic Amelogenesis Imperfecta (AI) patients. MATERIALS AND METHODS: A cross-sectional study was performed including 28 patients, 14 with hypoplastic AI and 14 controls for whom Decayed (D), Missed (M) and Filled (F) Teeth (T) were checked for a DMFT index evaluation. Twenty-eight saliva samples, 4 bacterial plaques and 19 teeth were used. Decayed teeth were observed under polarized light and scanning electron microscopy. Salivary pH was measured and saliva bacterial strains were biochemically identified and confirmed by PCR. Bacterial adhesions to tooth surfaces were observed by Scanning Electron Microscopy (SEM) and evaluated by colony enumeration after in vitro culture of Streptococcus mutans and Lactobacillus casei with dental fragments. RESULTS: DMFT indexes were significantly lower in AI patients (mean DMFT = 0.8) compared to controls (mean DMFT = 2.9). Decayed teeth revealed sclerotic, demineralized, invaded and disintegrated zones in dentine. Dental plaques were rich with filamentous bacteria in AI patients. Oral microbiotome of the saliva showed a low rate of Streptococci and a significant high level of Bacillus spp, Enterococcus faecalis and Enterococcus faecium in AI patients. In vitro study showed a significant high adhesion of Lactobacillus casei and a weak adhesion of Streptococcus mutans on AI dental hard tissues. CONCLUSION: Our study showed that hypoplastic AI patients have (i) a low DMFT index, (ii) an alkaline pH of saliva enriched with Bacillus spp, Enterococcus faecalis and Enterococcus faecium and (iii) dental tissues more easily invaded by Lactobacilli than Streptococci. The combination of these bacteria seems to give AI patients protection against dental caries.

Implants Placement in Contact with Dental Tissue: A Potential Paradigm Shift? Systematic Literature Review.

The aim of this study was to explore the literature for clinical and histological data of an unconventional treatment with implants placement in contact with dental tissue (IPICDT) and to try to clarify its indications and surgical procedure particularities.Relevant publications published until May 2019 on the IPICDT were thoroughly reviewed. Search strategy was developed using a controlled vocabulary combination.Medline's exploration and manual research identified 397 articles; 15 of these were selected after screening. IPICDT was indicated in three clinical situations: impacted teeth, ankylosed teeth, or residual roots. Clinical and radiological follow-up were satisfied except for implants placed in contact with (and not through) roots. Histological analysis revealed different mineralized tissues formed on the implant surface: cementum on removed implants in human and osteodentin on implants placed in contact with animal teeth dentin and pulp. These findings were described as new concept of implants' "Mineral integration."According to this study, the follow-up results of implants placed in contact with roots were controversial. Some implants were stable and others were either removed or kept and disinfected after root extraction because of bacterial infection. However, implants placed through ankylosed or impacted teeth were stable. These findings suggest that the clinicians have to be cautious when applying this unconventional approach. Further studies are recommended to explore its long follow-up. It is also interesting to explore this technique in cases of syndromic dental diseases with several impacted teeth (such as cleidocranial dysplasia; or amelogenesis imperfecta).

Coupling xylitol with remineralizing agents improves tooth protection against demineralization but reduces antibiofilm effect.

To explore the efficiency of xylitol chewing gum enriched or not with remineralizing agents to protect tooth against cariogenic biofilm formation and demineralization. Six groups of chewing gums were prepared; Group 1: isomalt (1.8%), Group 2: casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) (3%) + isomalt (1.8%), Group 3: hydroxyapatite (HAP) (2.5%) + isomalt (1.8%), Group 4: xylitol (1.8%), Group 5: CPP-ACP (3%) + xylitol (1.8%) and Group 6: HAP (2.5%) + xylitol (1.8%). The antibiofilm properties of different chewing gum extracts using seven oral bacterial species including Streptococcus mutans, Streptococcus constellatus, Streptococcus Salivarius and Streptococcus oralis were explored via the crystal violet staining assay. The remineralizing effects of those products were assessed on thirty human permanent teeth, half-protected with varnish before chemical erosion and thermocycling process with chewing gum. Remineralization was evaluated using scanning electron microscopy and microscopic measurements on polarized light microscopy. The ratio R comparing the thickness between unvarnished and varnished sides was evaluated. While the minimum biofilm inhibitory concentration (MBIC50) was low for xylitol alone compared to isomalt, it was inconsistent when enriched with remineralizing agents. The minimum biofilm eradication concentration (MBEC50) was low for xylitol groups compared to isomalt, for all the studied strains. R was significantly lower in Group 1 and Group 2, while Group 6 showed the highest ratio. Xylitol chewing gums confirmed good antibiofilm properties and showed remineralized potential on eroded teeth. When xylitol is associated to CPP-ACP or HAP, antibiofilm activity decreased while remineralization of eroded teeth increased.

Antitumoral Potency by Immunomodulation of Chloroform Extract from Leaves of Nitraria retusa, Tunisian Medicinal Plant, via its Major Compounds ß-sitosterol and Palmitic Acid in BALB/c Mice Bearing Induced Tumor.

This study evaluated the antitumoral effect of Chloroform extract from Nitraria retusa leaves, via its major compounds ß-sitosterols and palmitic acid. BALB/c mice were subcutaneously inoculated with B16-F10 cells, then treated intra-peritoneally after 7 days with the chloroform extract for 21 days. They were then euthanized, and the tumors were weighed. Lung parenchyma was analyzed. Lymphocyte and macrophages proliferation, cytotoxic T lymphocyte (CTL) activities were evaluated using the MTT assay. Macrophage phagocytosis was evaluated by measuring the lysosomal activity and nitric oxide production. Antioxidant activity was studied by cellular antioxidant activity on macrophage and splenocytes and by lipid peroxidation inhibitory activity in liver cells, kidney, and serum. ß-sitosterols and palmitic acid, major compounds of chloroform extract, impeded remarkably the expansion of the transplantable tumor, protected the lung parenchyma, and increased splenocytes proliferation and both CTL activities in tumor-bearing mice. ß-sitosterols and palmitic acid were also seen to have enhanced lysosomal activity of host macrophages and antioxidant cellular activity. Also, they showed an inhibitory effect of lipid peroxidation. Our results suggest that antitumoral effect of ß-sitosterols and palmitic acid from chloroform extract is related with its immunomodulatory activity, and opens the way for a nutrition application and coprocessing phytotherapy against cancer.

Melatonin modulates endoplasmic reticulum stress and Akt/GSK3-beta signaling pathway in a rat model of renal warm ischemia reperfusion.

Melatonin (Mel) is widely used to attenuate ischemia/reperfusion (I/R) injury in several organs. Nevertheless, the underlying mechanisms remain unclear. This study was conducted to explore the effect of Mel on endoplasmic reticulum (ER) stress, Akt and MAPK cascades after renal warm I/R. Eighteen Wistar rats were randomized into three groups: Sham, I/R, and Mel + I/R. The ischemia period was 60 min followed by 120 min of reperfusion. Mel (10 mg/kg) was administrated 30 min prior to ischemia. The creatinine clearance, MDA, LDH levels, and histopathological changes were evaluated. In addition, Western blot was performed to study ER stress and its downstream apoptosis as well as phosphorylation of Akt, GSK-3ß, VDAC, ERK, and P38. Mel decreased cytolysis and lipid peroxidation and improved renal function and morphology compared to I/R group. Parallely, it significantly reduced the ER stress parameters including GRP 78, p-PERK, XBP 1, ATF 6, CHOP, and JNK. Simultaneously, p-Akt level was significantly enhanced and its target molecules GSK-3ß and VDAC were inhibited. Furthermore, the ERK and P38 phosphorylation were evidently augmented after Mel administration in comparison to I/R group. In conclusion, Mel improves the recovery of renal function by decreasing ER stress and stimulating Akt pathway after renal I/R injury.

Topography of the pulp chamber in the maxillary primary molars of a Tunisian children.

BACKGROUND: Knowledge of the variations in pulp chamber topography is very useful to the dentist while performing endodontic access cavity in primary teeth. AIM: To determine the horizontal crown dimensions and the pulp chamber topography of shedding maxillary primary molars in a sample of Tunisian children. MATERIALS AND METHODS: Forty two maxillary primary molars (24 first and 18 second molars) were used. Teeth were ultrasonically cleaned before taking morphometric measurements of the buccolingual and mesiodistal sizes. Statistical student test was used comparing the mean sizes. RESULTS: All the crown measurements were higher in the second molar. While the buccolingual dimension of the pulp chamber was higher in the second molar, the mesiodistal sizes were equivalent among the teeth. The thickest walls were in the buccal side of the first molar and in the mesial side of the second one. The ratio of several measurements indicate that the pulp cavity is more vestibularly and distally located in the second molar. CONCLUSION: Mesiodistal dimension of the pulp cavity is not proportional to that of the dental crown. Endodontic access cavity have to be shifted to the distal and vestibular sides from the first to the second upper molar.

RANK, RANKL and OPG expressions in a permanent molar with a replacement resorption.

The aim of this study was to explore the expression of RANK, RANKL and OPG during ankylosis. Structural details and immunohistochemical investigations of the expression of RANK, RANKL and OPG in an extracted secondary retained permanent molar of a 12-year-old girl are reported. Woven and lamellar bones were observed in the thickness of the remodeled dental wall and a tertiary dentin was noticed around the pulp cavity. The resorbing multinucleated cells expressed TRAP and RANK but few of them also expressed RANKL. Both odontoblasts and osteoblasts expressed TRAP and RANK, but the expression of RANKL was limited to osteoblasts. OPG remained undetected. The present case reveals unusual expression of RANKL in the resorbing cells, TRAP and RANK in both osteoblasts and odontoblasts, and a failure of detection of OPG. These proteins could be involved in the pathogenesis of tooth ankylosis.

Attenuation of endoplasmic reticulum stress and mitochondrial injury in kidney with ischemic postconditioning application and trimetazidine treatment.

BACKGROUND: Endoplasmic reticulum (ER) and mitochondria have been implicated in the pathology of renal ischemia/reperfusion (I/R). In the present study, we investigated whether the use of ischemic postconditioning (IPostC) and trimetazidine (TMZ) separately or combined could reduce ER stress and mitochondria damage after renal ischemia. METHODS: Kidneys of Wistar rats were subjected to 60-min of warm ischemia followed by 120-min of reperfusion (I/R group, n = 6), or to 6 cycles of ischemia/reperfusion (10-s each cycle) just after 60-min of warm ischemia (IPostC group, n = 6), or to i.p. injection of TMZ (3 mg/kg) 30-min before ischemia (TMZ group, n = 6), or to the combination of both treatments (IPostC+TMZ group, n = 6). The results of these experimental groups were compared to those of a sham-operated group in which rat renal pedicles were only dissected. Sodium reabsorption rate, creatinine clearance lactate deshydrogenase (LDH) activity in plasma, and concentration of malonedialdehyde (MDA) in tissue were determined. In addition, Western blot analysis was performed to identify the amounts of cytochrome c, c-JunNH2-terminal kinase (JNK), voltage-dependent anion channel (VDAC), glycogen synthase kinase 3-beta (GSK3-ß), and ER stress parameters. RESULTS: IPostC or/and TMZ significantly decreased cytolysis, oxidative stress and improved renal function in comparison to I/R group. IPostC but not TMZ significantly attenuated ER stress parameters versus I/R group. Indeed, it down-regulated the glucose-regulated protein 78 (GRP78), the activating transcription factor 4 (ATF4), the RNA activated protein kinase (PKR)-like ER kinas (PERK), the X box binding protein-1 (XBP-1) and the caspase12 protein levels. TMZ treatment significantly augmented GSK3-ß phosphorylation and reduced levels of cytochrome c and VDAC phosphorylation in comparison to IPostC application. The combination of both treatments gave a synergetic effect. It significantly improved the survival rate, attenuated cytolysis, oxidative stress and improved renal function. CONCLUSION: This study revealed that IPostC protects kidney from I/R injury by suppressing ER stress while the beneficial effects of TMZ are mediated by mitochondria protection. The combination of both treatments ameliorated functional recovery.

The effect of etidronate on the periodontium of ovariectomized rats.

BACKGROUND: Bisphosphonates are indicated for the treatment of osteoporosis. However, they could have an adverse effect on specific sites, such as the bisphosphonate-related osteonecrosis of the jaw. The aim of this study is to investigate the effect of etidronate on the resorption and apposition sides of the periodontium in ovariectomized rats. METHODS: Twenty-four female Wistar rats were randomly subjected to either ovariectomy or sham operation. After 8 weeks, six animals of each group were sacrificed. The other 12 rats received 5 mg/kg/day etidronate subcutaneously during 4 weeks. Tartrate-resistant acid phosphatase reaction and immunohistochemical staining for receptor activator of nuclear factor-κB (RANK), RANK-ligand (RANKL), osteoprotegerin (OPG), and osteocalcin was performed. Immunoreactivity was evaluated using a semiquantitative analysis. RESULTS: In ovariectomized rats, osteoclasts were noticed in the root socket of molars, including the apposition side of the periodontium, in which RANKL expression was significantly evidenced. In the etidronate-treated group, OPG expression was significantly expressed and osteoclasts that were noticed in the resorption side remained undetected in the apposition side even under ovariectomy. RANK was significantly expressed in ovariectomized rats treated with etidronate. Osteoid formation and osteocalcin expression were described on the alveolar bone surfaces in etidronate-treated rats, with or without ovariectomy. CONCLUSIONS: Etidronate has specific site and bone cell actions in the periodontium. It inhibits the osteoclast differentiation induced by ovariectomy in the apposition side of the periodontium but maintains bone formation over all the socket surfaces. Such specificity may be related to the pathogenesis of the bisphosphonate-induced osteonecrosis of the jaw.

Disruptions of the pulp-dentin complex in an investigated case of pink secondary retained molar.

A pink retained left mandibular first molar without carious lesions was diagnosed in a healthy 12-year-old girl presenting normal clinical tests. An orthopantomogram failed to detect other retained teeth. Both periapical radiography and computed tomography showed the absence of a periodontal ligament space in the bifurcation area and the presence of radiolucency or calcifications in the pulp cavity. The coronal part of the removed tooth was subjected to histological and immunohistochemical analysis using anti-PCNA (proliferation marker) and anti-p53 (apoptosis marker) antibodies. Root surfaces were observed by scanning electron microscopy. The pink color of the molar reflected the extension of resorptive tissue into the clinical crown and the underlining proliferation of pulp vessels. Ankylosis observed in the bifurcation area was also detected in the coronal part of the pulp. Whereas odontoblasts secreted tertiary dentin despite no evidence for a carious lesion, only osteocytes in the newly formed bone were apoptotic and the root surfaces were free of resorption lacunae. The etiopathology of the lesion in this case indicated a pulp origin, suggesting that new therapies targeting this tissue should be developed.

Effect of in vitro exposure to Vibrio vulnificus on hydroelectrolytic transport and structural changes of sea bream (Sparus aurata L.) intestine.

The everted gut sac technique has been used to investigate the effect of Vibrio vulnificus on water and electrolyte (Na(+), K(+), Cl(-), HCO(3)(-)) transport on the intestine of sea bream (Sparus aurata L.). Both the anterior and the posterior intestine were incubated in a medium containing 10(8) V. vulnificus cells ml(-1) at 25 degrees C for 2 h. The presence of V. vulnificus resulted in a significant reduction (P < 0.05) of water absorption in the anterior intestine, while sodium absorption in the anterior (P < 0.01) and posterior (P < 0.05) intestine was elevated. Chloride absorption was increased, but the changed was not significant, while potassium absorption decreased significantly (P < 0.05), but only in the posterior intestine. Incubation the sea bream intestine with V. vulnificus did not affect carbonate secretion in the anterior segment, whereas high secretion was stimulated in the posterior segment (P < 0.01). Histological evaluations demonstrated damage in the anterior intestine of sea bream that was characterized by the detachment of degenerative enterocytes, alterations in the microvilli, and the presence of a heterogenous cell population, indicating inflammation. Based on our results, we conclude that V. vulnificus caused cell damage to the intestine of sea bream and that the anterior intestine is more susceptible than the posterior part of the intestine. Several hypotheses are suggested to explain our observations, such as the presence of higher numbers of villosities in the anterior intestine than in the posterior one and/or the presence of endogenous bacteria in the posterior intestine which may have a protector role.

Msx and dlx homeogene expression in epithelial odontogenic tumors.

Epithelial odontogenic tumors are rare jaw pathologies that raise clinical diagnosis and prognosis dilemmas notably between ameloblastomas and clear cell odontogenic carcinomas (CCOCs). In line with previous studies, the molecular determinants of tooth development-amelogenin, Msx1, Msx2, Dlx2, Dlx3, Bmp2, and Bmp4-were analyzed by RT-PCR, ISH, and immunolabeling in 12 recurrent ameloblastomas and in one case of CCOC. Although Msx1 expression imitates normal cell differentiation in these tumors, other genes showed a distinct pattern depending on the type of tumor and the tissue involved. In benign ameloblastomas, ISH localized Dlx3 transcripts and inconstantly detected Msx2 transcripts in epithelial cells. In the CCOC, ISH established a lack of both Dlx3 and Msx2 transcripts but allowed identification of the antisense transcript of Msx1, which imitates the same scheme of distribution between mesenchyme and epithelium as in the cup stage of tooth development. Furthermore, while exploring the expression pattern of signal molecules by RT-PCR, Bmp2 was shown to be completely inactivated in the CCOC and irregularly noticeable in ameloblastomas. Bmp4 was always expressed in all the tumors. Based on the established roles of Msx and Dlx transcription factors in dental cell fates, these data suggest that their altered expression is a proposed trail to explain the genesis and/or the progression of odontogenic tumors.

Expression pattern of Dlx3 during cell differentiation in mineralized tissues.

The present study was designed to compare the expression pattern of Dlx3 in four different mineralized tissues because of: 1-its role in skeleton patterning, 2-its expression in dental epithelium and mesenchyme during morphogenesis, 3-the membranous and endochondral bone and tooth phenotype of tricho-dento-osseous syndrome related to Dlx3 gene mutation and 4-recently emerging knowledge on Dlx family members in the bone field. Ameloblasts, odontoblasts, osteoblasts and chondrocytes were analyzed in vitro and in vivo. Dlx3 transcripts were detected by RT-PCR in established model systems (microdissected dental epithelium and mesenchyme; primary cultures of rat chondrocytes), as recently performed in osteoblasts in vitro. A human 414-bp Dlx3 probe was generated. A 4.5-kb human Dlx3 sense RNA was identified in maxillo-facial samples by Northern blotting. Immunolabeling and in situ hybridization were performed in mice from Theiler stage E 14.5 until birth. In teeth, although Dlx3 was still expressed in differentiated ameloblasts, it was down regulated during odontoblast polarization. During endochondral bone formation, Dlx3 protein was detected in chondrocytes and was most strongly expressed in the prehypertrophic cartilage zone and in differentiating and differentiated osteoblasts of metaphyseal periosteum. In vitro, real-time PCR studies supported this upregulation in prehypertrophic chondrocytes, closely correlated with Ihh variations. In membranous bone, Dlx3 was present in preosteoblasts, osteoblasts and osteoid-osteocytes. The present data on Dlx3 and recently published functional studies show that this transcription factor may be instrumental during growth in the control of matrix deposition and biomineralization in the entire skeleton.