Oral Wound Healing Potential of Polygoni Cuspidati Rhizoma et Radix Decoction-In Vitro Study.

Polygoni Cuspidati Rhizoma et Radix (syn. rhizomes of Reynoutria japonica Houtt.) is a pharmacopoeial raw material in Europe and China. In traditional medicine, one of the applications for Reynoutria japonica rhizomes is wound healing. In a recent in vitro study, we demonstrated that ethanol and acetone extracts from this herbal drug have the potential to heal oral gum wounds. However, considering that a majority of herbal medicines have been traditionally administered as water decoctions, in the present study, a decoction of Reynoutria japonica rhizomes was prepared and detailed tests to determine its in vitro gingival wound healing activity were conducted. We used the primary human gingival fibroblasts (HGF) incubated with a decoction to determine cell viability (MTT assay), cell proliferation (the confocal laser scanning microscope-CLSM), and cell migration (wound healing assay). Moreover, the collagen type III expression was examined using immunocytochemical staining. The studied decoction was qualitatively and quantitatively characterized using the validated HPLC/DAD/ESI-HR-QTOF-MS method. The Folin-Ciocalteu assay was used to determine the total phenols and tannins content. Additionally, HPLC-RI analysis of decoction and the previously obtained ethanol and acetone extracts was used to determine the composition of saccharides. Low concentration (from 50 to 1000 µg/mL) of decoction after 24 h incubation caused a significant increase in HGF cell viability. No cytotoxic effect was observed at any tested concentration (up to 2000 µg/mL). The lowest active concentration of decoction (50 µg/mL) was selected for further experiments. It significantly stimulated human gingival fibroblasts to proliferate, migrate, and increase the synthesis of collagen III. Phytochemical analysis showed significantly fewer polyphenols in the decoction than in the ethanol and acetone extracts tested earlier. In contrast, high levels of polysaccharides were observed. In our opinion, they may have a significant effect on the oral wound healing parameters analyzed in vitro. The results obtained encourage the use of this raw material in its traditional, safe form-decoction.

Proanthocyanidins and Flavan-3-ols in the Prevention and Treatment of Periodontitis-Immunomodulatory Effects, Animal and Clinical Studies.

This paper continues the systematic review on proanthocyanidins and flavan-3-ols in the prevention and treatment of periodontal disease and covers the immunomodulatory effects, and animal- and clinical studies, while the other part discussed the direct antibacterial properties. Inflammation as a major response of the periodontal tissues attacked by pathogenic microbes can significantly exacerbate the condition. However, the bidirectional activity of phytochemicals that simultaneously inhibit bacterial proliferation and proinflammatory signaling can provide a substantial alleviation of both cause and symptoms. The modulatory effects on various aspects of inflammatory and overall immune response are covered, including confirmed and postulated mechanisms of action, structure activity relationships and molecular targets. Further, the clinical relevance of flavan-3-ols and available outcomes from clinical studies is analyzed and discussed. Among the numerous natural sources of flavan-3-ols and proanthocyanidins the most promising are, similarly to antibacterial properties, constituents of various foods, such as fruits of Vaccinium species, tea leaves, grape seeds, and tannin-rich medicinal herbs. Despite a vast amount of in vitro and cell-based evidence of immunomodulatory there are still only a few animal and clinical studies. Most of the reports, regardless of the used model, indicated the efficiency of these phytochemicals from cranberries and other Vaccinium species and tea extracts (green or black). Other sources such as grape seeds and traditional medicinal plants, were seldom. In conclusion, the potential of flavan-3-ols and their derivatives in prevention and alleviation of periodontal disease is remarkable but clinical evidence is urgently needed for issuing credible dietary recommendation and complementary treatments.

Proanthocyanidins and Flavan-3-Ols in the Prevention and Treatment of Periodontitis-Antibacterial Effects.

Flavan-3-ols and their oligomeric forms called proanthocyanidins are polyphenolic compounds occurring in several foodstuffs and in many medicinal herbs. Their consumption is associated with numerous health benefits. They exhibit antioxidant, anti-inflammatory, cytoprotective, as well as antimicrobial activity. The latter property is important in the prevention and treatment of periodontal diseases. Periodontitis is a multifactorial polymicrobial infection characterized by a destructive inflammatory process affecting the periodontium. Using non-toxic and efficient natural products such as flavanol derivatives can significantly contribute to alleviating periodontitis symptoms and preventing the disease's progress. Therefore, a comprehensive systematic review of proanthocyanidins and flavan-3-ols in the prevention and treatment of periodontitis was performed. The present paper reviews the direct antibacterial effects of these compounds against periodontic pathogens. The immunomodulatory effects, including animal and clinical studies, are included in a separate, parallel article. There is significant evidence supporting the importance of the antibacterial action exerted by proanthocyanidins from edible fruits, tea, and medicinal herbs in the inhibition of periodontitis-causing pathogens.

Cytotoxic Effect of Vanicosides A and B from Reynoutria sachalinensis Against Melanotic and Amelanotic Melanoma Cell Lines and in silico Evaluation for Inhibition of BRAFV600E and MEK1.

Vanicosides A and B are the esters of hydroxycinnamic acids with sucrose, occurring in a few plant species from the Polygonaceae family. So far, vanicosides A and B have not been evaluated for anticancer activity against human malignant melanoma. In this study, we tested these two natural products, isolated from Reynoutria sachalinensis rhizomes, against two human melanoma cell lines (amelanotic C32 cell line and melanotic A375 cell line, both bearing endogenous BRAFV600E mutation) and two normal human cell lines-keratinocytes (HaCaT) and the primary fibroblast line. Additionally, a molecular docking of vanicoside A and vanicoside B with selected targets involved in melanoma progression was performed. Cell viability was studied using an MTT assay. A RealTime-Glo™ Annexin V Apoptosis and Necrosis assay was used for monitoring programmed cell death (PCD). Vanicoside A demonstrated strong cytotoxicity against the amelanotic C32 cell line (viability of the C32 cell line was decreased to 55% after 72 h incubation with 5.0 µM of vanicoside A), significantly stronger than vanicoside B. This stronger cytotoxic activity can be attributed to an additional acetyl group in vanicoside A. No significant differences in the cytotoxicity of vanicosides were observed against the less sensitive A375 cell line. Moreover, vanicosides caused the death of melanoma cells at concentrations from 2.5 to 50 µM, without harming the primary fibroblast line. The keratinocyte cell line (HaCaT) was more sensitive to vanicosides than fibroblasts, showing a clear decrease in viability after incubation with 25 µM of vanicoside A as well as a significant phosphatidylserine (PS) exposure, but without a measurable cell death-associated fluorescence. Vanicosides induced an apoptotic death pathway in melanoma cell lines, but because of the initial loss of cell membrane integrity, an additional cell death mechanism might be involved like permeability transition pore (PTP)-mediated necrosis that needs to be explored in the future. Molecular docking indicated that both compounds bind to the active site of the BRAFV600E kinase and MEK-1 kinase; further experiments on their specific inhibitory activity of these targets should be considered.

Chemical Composition of East Asian Invasive Knotweeds, their Cytotoxicity and Antimicrobial Efficacy Against Cariogenic Pathogens: An In-Vitro Study.

BACKGROUND Giant knotweeds originating from East Asia, such as Reynoutria japonica, and Reynoutria sachalinensis, and their hybrid such as Reynoutria x bohemica, are invasive plants in Europe and North America. However, R. japonica is also a traditional East Asian drug (Polygoni cuspidati rhizoma) used in Korean folk medicine to improve oral hygiene. The aim of this study was to evaluate the antibacterial activity of acetone extracts of Reynoutria species against dominant caries pathogen such as Streptococcus mutans and alternative pathogens, as well as characterize the phytochemical composition of extracts and examine their cytotoxicity. MATERIAL AND METHODS Ultrasonic extraction was used to obtain polyphenol-rich extracts. The extracts were characterized by HPLC-DAD-ESI-MS. To test bacterial viability, the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against S. mutans, S. salivarius, S. sanguinis, and S. pyogenes were determined. The cytotoxicity of the extracts to human fibroblasts derived from gingiva was evaluated using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. RESULTS The R. japonica extract had the highest bacteriostatic and bactericidal activity against pathogens causing caries, mainly dominant caries pathogen S. mutans (mean MIC 1000 µg/mL and MBC 2000 µg/mL), which was most likely associated with a higher content of stilbene aglycons and anthraquinone aglycons in the extract. Moreover, the R. japonica extract demonstrated the lowest cytotoxic effect on human fibroblasts and exhibited cytotoxic activity only at the concentration causing the death of all S. mutans. CONCLUSIONS The results indicate that the R. japonica acetone extract can be considered as a natural, antimicrobial agent for caries control.

Phytochemical Diversity in Rhizomes of Three Reynoutria Species and their Antioxidant Activity Correlations Elucidated by LC-ESI-MS/MS Analysis.

The rhizome of Reynoutria japonica is a well-known traditional herb (Hu zhang) used in East Asia to treat various inflammatory diseases, infections, skin diseases, scald, and hyperlipidemia. It is also one of the richest natural sources of resveratrol. Although, it has been recently included in the European Pharmacopoeia, in Europe it is still an untapped resource. Some of the therapeutic effects are likely to be influenced by its antioxidant properties and this in turn is frequently associated with a high stilbene content. However, compounds other than stilbenes may add to the total antioxidant capacity. Hence, the aim of this research was to examine rhizomes of R. japonica and the less studied but morphologically similar species, R. sachalinensis and R. x bohemica for their phytochemical composition and antioxidant activity and to clarify the relationship between the antioxidant activity and the components by statistical methods. HPLC/UV/ESI-MS studies of three Reynoutria species revealed 171 compounds, comprising stilbenes, carbohydrates, procyanidins, flavan-3-ols, anthraquinones, phenylpropanoids, lignin oligomers, hydroxycinnamic acids, naphthalenes and their derivatives. Our studies confirmed the presence of procyanidins with high degree of polymerization, up to decamers, in the rhizomes of R. japonica and provides new data on the presence of these compounds in other Reynoutria species. A procyanidin trimer digallate was described for the first time in, the studied plants. Moreover, we tentatively identified dianthrone glycosides new for these species and previously unrecorded phenylpropanoid disaccharide esters and hydroxycinnamic acid derivatives. Furthermore, compounds tentatively annotated as lignin oligomers were observed for the first time in the studied species. The rhizomes of all Reynoutria species exhibited strong antioxidant activity. Statistical analysis demonstrated that proanthocyanidins should be considered as important contributors to the total antioxidant capacity.

Fractal dimension analysis a supplementary mathematical method for bone defect regeneration measurement.

AIM OR PURPOSE: The geometry of a bone defect is very complex. Its shape is too complicated to measure or compare with other bone defects using only traditional measuring methods Traditional measuring techniques based on the histomorphometric analysis of a bone specimen require supplementary measuring. For the fractal dimension analysis (FDA) mathematic formulas are used to describe complicated and chaotic shapes. The FDA offers a possibility of a comparison between complicated and complex shapes such as a histological image of a bone defect. The aim of this study was to evaluate the FDA of bone defects as a supplementary method for a defect regeneration assessment. MATERIALS AND METHODS: For the purpose of this study, microscopic photographs of bone specimens stained with hematoxylin and eosin obtained during a block biopsy were used. The bone blocks used in this study were obtained during a rat animal model study. Specimens were collected from 36 Wistar rats where a cranial defect was created and augmented with five different novel biomaterials and compared to the unfilled defect in the control group. New bone formation in every specimen was histomorphometrically measured by two independent operators and compared to FDA measurements. RESULTS: Both traditional and FDA techniques have shown statistically significant differences between bone formation in test groups compared to the control one; on the other hand, no statistically significant difference was found between other groups. The Pearson's r-test was conducted to measure the linear dependence (correlation) between standard measurements and the FDA, and a positive linear correlation was found -r=0.94. CONCLUSIONS: The FDA can be used as a supplementary method for bone regeneration measurements.

Ultrasound assessment of soft tissue augmentation around implants in the aesthetic zone using a connective tissue graft and xenogeneic collagen matrix - 1-year randomised follow-up.

PURPOSE: A comparative, ultrasound evaluation of the thickness of keratinized mucosa (TKT) around implants one year after gingival augmentation (GA) by means of a connective tissue graft (CTG) and the xenogeneic collagen matrix (CMX). MATERIALS AND METHODS: A total of 75 bone level tapered implants (Conelog® Camlog) were inserted in 57 patients in the aesthetic area of both jaws. The patients were divided into 3 groups: control group I- without GA; group II- GA 3 months before implantation, and group III- GA 3 months after implantation. Groups II and III were divided into two subgroups depends on type of material used for GA: (a) CMX (Mucograft®, Geistlich Pharma AG) and (b) CTG. The patients underwent a clinical and ultrasound examination before, then after 3 and 12 months following GA respectively to evaluate TKT at two points using ultrasound equipment (Pirop®, Echoson). Point 1 was considered to be in the middle of the line connecting the cemento-enamel junction (CEJ) to the adjacent teeth, and point 2 on the mucogingival junction (MGJ). RESULTS: Three months after GA, the highest increase in gingival thickness was noted in group IIIb (point 1 - 0.95mm, 2 - 1.01mm). However, 12 months after GA the highest gingival thickness was observed in group IIb (point 1 - 1.76mm, 2 - 1.36m) and next IIIb (point 1 - 1.52mm, 2 - 1.15mm). CONCLUSIONS: Both CTG and Geistlich Mucograft® increased TKT, but higher values were noted using CTG augmentation before implantation. An ultrasonic device can be used as a non-invasive, reliable, and reproducible method for evaluating TKT.