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Miscellaneous: In-Vitro

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Anti-inflammatory and macrophage polarization effects of Cranberry Proanthocyanidins (PACs) for periodontal and peri-implant disease therapy

Posted: 
March 2, 2021
Authors: 
Galarraga-Vinueza ME; Dohle E; Ramanauskaite A; Al-Maawi S; Obreja K; Magini R; Sader R; Ghanaati S; Schwarz F.
Journal: 
Journal of Periodontal Research. 55(6):821-829, 2020 Dec
Abstract: 

BACKGROUND AND OBJECTIVE: Macrophages' cytokine expression and polarization play a substantial role in the host's "destructive" inflammatory response to periodontal and peri-implant pathogens. This study aimed to evaluate cell viability, anti-inflammatory activity, and macrophage polarization properties of different cranberry concentrates. METHODS: THP-1 cells (monocytic line) were treated with phorbol myristic acid to induce macrophage differentiation. Human gingival fibroblasts (HFIB-G cell line), osteosarcoma-derived osteoblasts (SAOS-2 cell line), and induced macrophages were treated with cranberry concentrates at 25, 50, and 100 microg/mL for 120 seconds, 1 hour and 24 hours. Untreated cells at the same time points served as controls. For anti-inflammatory analysis, induced macrophages exposed to cranberry concentrates (A-type PACs) were stimulated with lipopolysaccharides (LPS) derived from E coli for 24 hours. Cell viability, interleukin (IL)-8, IL-1 s, IL-6, and IL-10 expression of LPS-stimulated macrophages, and macrophage polarization markers were evaluated through determination of live-cell protease activity, enzyme-linked immunosorbent assay, and immunofluorescence staining semi-quantification. RESULTS: Cranberry concentrates (A-type PACs) did not reduce HGF, SAOS-2, and macrophage viability after 24 hours of exposure. Pro-inflammatory cytokine expression (ie IL-8 and IL-6) was downregulated in LPS-stimulated macrophages by cranberry concentrates at 50 and 100 microg/mL. Anti-inflammatory IL-10 expression was significantly upregulated in LPS-stimulated macrophages by cranberry concentrates at 100 microg/mL after 24 hours of exposure. M1 polarization significantly decreased when LPS-stimulated macrophages were exposed to cranberry concentrates. High levels of positive M1 macrophages were present in all untreated control groups. M2 polarization significantly increased at all LPS-stimulated macrophages exposed to cranberry concentrates for 1 and 24 hours. CONCLUSION: Cranberry-derived proanthocyanidins may have the potential to act as an anti-inflammatory component in the therapy of periodontal and peri-implant diseases.

Cranberry extract-based formulations for preventing bacterial biofilms

Posted: 
March 2, 2021
Authors: 
Greene AC; Acharya AP; Lee SB; Gottardi R; Zaleski E; Little SR.
Journal: 
Drug Delivery & Translational Research. 2020 Aug 11.
Abstract: 

Generating formulations for the delivery of a mixture of natural compounds extracted from natural sources is a challenge because of unknown active and inactive ingredients and possible interactions between them. As one example, natural cranberry extracts have been proposed for the prevention of biofilm formation on dental pellicle or teeth. However, such extracts may contain phenolic acids, flavonol glycosides along with other constituents like coumaroyl iridoid glycosides, flavonoids, alpha-linolenic acid, n-6 (or n-3) fatty acids, and crude fiber. Due to the presence of a variety of compounds, determining which molecules (and how many molecules) are essential for preventing biofilm growth is nontrivial to ascertain. Therefore, a formulation that could contain natural, unrefined, cranberry extract (with all its constituent compounds) at high loading would be ideal. Accordingly, we have generated several candidate formulations including poly(lactic-co-glycolic) acid (PLGA)-based microencapsulation of cranberry extract (CE15) as well as formulations including stearic acid along with polyvinylpyrrolidone (PVP) or Ethyl lauroyl arginate (LAE) complexed with cranberry extracts (CE15). We found that stearic acid in combination with PVP or LAE as excipients led to higher loading of the active and inactive compounds in CE15 as compared with a PLGA microencapsulation and also sustained release of CE15 in a tunable manner. Using this method, we have been able to generate two successful formulations (one preventative based, one treatment based) that effectively inhibit biofilm growth when incubated with saliva. In addition to cranberry extract, this technique could also be a promising candidate for other natural extracts to form controlled release systems.

Development of a cranberry standard for quantification of insoluble cranberry (Vaccinium macrocarpon Ait.) proanthocyanidins

Posted: 
March 2, 2021
Authors: 
Gullickson, E. R.; Krueger, C. G.; Birmingham, A.; Maranan, M.; Reed, J. D
Journal: 
Journal of Agricultural and Food Chemistry; 2020. 68(10):2900-2905.
Abstract: 

Cranberry proanthocyanidins (PACs) can be partitioned into soluble PACs, which are extracted with solvents, and insoluble PACs, which remain associated with fibers and proteins after extraction. Most research on cranberry products only quantifies soluble PACs because proper standards for quantifying insoluble PACs are lacking. In this study, we evaluated the ability of a cranberry PAC (c-PAC) standard, reflective of the structural heterogeneity of PACs found in cranberry fruit, to quantify insoluble PACs by the butanol-hydrochloric acid (BuOH-HCl) method. For the first time, a c-PAC standard enabled conversion of BuOH-HCl absorbance values (550 nm) to a weight (milligram) basis, allowing for quantification of insoluble PACs in cranberries. The use of the c-PAC reference standard for sequential analysis of soluble PACs by the method of 4-(dimethylamino)cinnamaldehyde and insoluble PACs by the method of BuOH-HCl provides analytical tools for the standardization of cranberry-based ingredients

Antibiofilm Properties of Triclosan with EDTA or Cranberry as Foley Catheter Lock Solutions.

Posted: 
March 20, 2020
Authors: 
Ayyash M; Shehabi AA; Mahmoud NN; Al-Bakri AG.
Journal: 
Journal of Applied Microbiology. 127(6):1876-1888
Abstract: 

AIMS: To investigate the efficiency of triclosan, ethylenediaminetetraacetic acid (EDTA) and cranberry alone or in combinations against Escherichia coli strains as urinary catheter lock solutions to reduce catheter-associated urinary tract infections.METHODS AND RESULTS: Viable counting was used to assess antibiofilm activities for triclosan, EDTA and cranberry alone or in combinations against E. coli strains embedded in biofilm onto all-silicon Foley catheter surface. The results revealed that combination of triclosan (10 mg ml-1 /EDTA 30 mg ml-1 ) when filling the catheter balloon was able to eradicate and prevent biofilm formation among all tested E. coli including the resistant strains, whereas triclosan (8.5 mg ml-1 )/ cranberry (103 mg ml-1 ) combination was a successful catheter lock solution by preventing all tested strains from adhering onto catheter surface when filled via the eye hole.CONCLUSIONS: The combinations of triclosan/EDTA and triclosan/cranberry were significantly effective in eradicating and preventing biofilm formation of the tested E. coli strains on Foley catheters.SIGNIFICANCE AND IMPACT OF THE STUDY: Combinations of triclosan/EDTA and triclosan/cranberry have a promising application as nonantibiotic catheter lock solution.

Comparative Evaluation of the Antimicrobial Effects of Different Mouthrinses against Streptococcus Mutans: An in Vitro Study.

Posted: 
March 20, 2020
Authors: 
Abu-Obaid E; Salama F; Abu-Obaid A; Alanazi F; Salem M; Auda S.
Journal: 
Journal of Clinical Pediatric Dentistry. 43(6):398-407
Abstract: 

Purpose: To assess the antimicrobial effects of different natural and semi-natural mouthrinses on isolates of S. mutans obtained from the saliva of Saudi children and reference strains of S mutans (ATCC 25175). Study design: Saliva samples were collected from 20 children. Natural and semi-natural mouthrinses included were herbal mix mouthrinse, cranberry mouthrinse, chlorhexidine digluconate mouthrinse, cranberry extract mixed with chlorhexidine digluconate mouthrinse, chlorhexidine digluconate mouthrinse with alcohol (positive control), and distilled water (negative control). The microbiological examination tests were minimal inhibitory concentration, minimal bactericidal concentration, and zone of inhibition for the saliva isolates of S. mutans while zone of inhibition test only for reference strain of S. mutans. Results: For reference strain in a comparison with the distilled water, the herbal mix, cranberry, cranberry mixed with chlorhexidine, chlorhexidine, and chlorhexidine with alcohol showed significantly increased zones of inhibition by 36.38, 36.25, 26.13, 17.75, and 12.38, respectively. For saliva isolates in a comparison with the distilled water, the herbal mix, cranberry, cranberry mixed with chlorhexidine, chlorhexidine, and chlorhexidine with alcohol showed significantly increased zones of inhibition by 38.00, 34.25, 22.94, 16.50, and 16.44, respectively. Chlorhexidine with alcohol showed significantly lower minimum inhibitory and bactericidal concentration than the other groups. Conclusions: Herbal mix and cranberry mouthrinses could be effective natural alternative to chlorhexidine mouthrinse with or without alcohol in affecting tested parameters.

Cranberry Extracts Promote Growth of Bacteroidaceae and Decrease Abundance of Enterobacteriaceae in A Guman Gut Simulator Model.

Posted: 
March 20, 2020
Authors: 
O'Connor, K. Morrissette, M. Strandwitz, P. Ghiglieri, M. Caboni, M. Liu HaiYan Khoo, C. D'Onofrio, A. Lewis, K.
Journal: 
PLoS ONE; 2019. 14(11):e0224836.
Abstract: 

The opportunistic pathogen Escherichia coli, a common member of the human gut microbiota belonging to the Enterobacteriaceae family, is the causative agent of the majority of urinary tract infections (UTIs). The gut microbiota serves as a reservoir for uropathogenic E. coli where they are shed in feces, colonize the periurethral area, and infect the urinary tract. Currently, front line treatment for UTIs consists of oral antibiotics, but the rise of antibiotic resistance is leading to higher rates of recurrence, and antibiotics cause collateral damage to other members of the gut microbiota. It is commonly believed that incorporation of the American cranberry, Vaccinium macrocarpon, into the diet is useful for reducing recurrence of UTIs. We hypothesized such a benefit might be explained by a prebiotic or antimicrobial effect on the gut microbiota. As such, we tested cranberry extracts and whole cranberry powder on a human gut microbiome-derived community in a gut simulator and found that cranberry components broadly modulate the microbiota by reducing the abundance of Enterobacteriaceae and increasing the abundance of Bacteroidaceae. To identify the specific compounds responsible for this, we tested a panel of compounds isolated from cranberries for activity against E. coli, and found that salicylate exhibited antimicrobial activity against both laboratory E. coli and human UTI E. coli isolates. In a gut simulator, salicylate reduced levels of Enterobacteriaceae and elevated Bacteroidaceae in a dose dependent manner.

Cranberry Proanthocyanidins Neutralize the Effects of Aggregatibacter actinomycetemcomitans Leukotoxin

Posted: 
March 20, 2020
Authors: 
Amel Ben Lagha, Amy Howell and Daniel Grenier
Journal: 
Toxins 2019, 11(11), 662; https://doi.org/10.3390/toxins11110662
Abstract: 

Aggregatibacter actinomycetemcomitans is a Gram-negative bacterium that has been strongly associated with localized aggressive periodontitis. The capacity of A. actinomycetemcomitans to produce a leukotoxin (LtxA) that activates pyroptosis in macrophages and induces the release of endogenous danger signals is thought to play a key role in the disease process. The aim of the present study was to investigate the effects of cranberry proanthocyanidins (PACs) on gene expression and cytotoxic activities of LtxA. We showed that cranberry PACs dose-dependently attenuate the expression of genes making up the leukotoxin operon, including ltxB and ltxC, in the two strains of A. actinomycetemcomitans tested. Cranberry PACs (≥62.5 µg/mL) protected macrophages against the cytotoxic effect of purified LtxA. Moreover, cranberry PACs reduced caspase-1 activation in LtxA-treated macrophages and consequently decreased the release of both IL-1β and IL-18, which are known as damage-associated molecular patterns (DAMPs) and contribute to the progression of periodontitis by increasing cell migration and osteoclastogenesis. In addition, cranberry PACs reduced the expression of genes encoding the P2X7 receptor and NALP3 (NACHT, LRR and PYD domains-containing protein 3), which play key roles in pore formation and cell death. Lastly, cranberry PACs blocked the binding of LtxA to macrophages and consequently reduced the LtxA-mediated cytotoxicity. In summary, the present study showed that cranberry PACs reduced LtxA gene expression in A. actinomycetemcomitans and neutralized the cytolytic and pro-inflammatory responses of human macrophages treated with LtxA. Given these properties, cranberry PACs may represent promising molecules for prevention and treatment of the aggressive form of periodontitis caused by A. actinomycetemcomitans.

The Effect of Cranberry Fruit Extract on Alpha -Synuclein Protein Expression using Immunostaining Techniques

Posted: 
March 20, 2020
Authors: 
Kumar, J. Sarvesh; Priya, V. Vishnu; Gayathri, R.
Journal: 
Drug Invention Today . Jun2019, Vol. 11 Issue 6, p1506-1508
Abstract: 

Abstract: Aim: The aim of the study was to identify the neuroprotective effect of cranberry fruit extract by reducing the release of this biomarker by immunostaining techniques. Objective: The present study is to find out the alpha-lipoic acid cranberry fruit extract on alpha-synuclein protein expression using immunostaining techniques. Materials and Methods: The neuroblastoma SH-SY5Y cells were cultured and treated with various concentrations of cranberry fruit extract and were incubated with primary antibody alpha-synuclein and the antigen-antibody activity was visualized under a light microscope, and the results were quantified using image analysis software. Results: The study infers that when cranberry extract is added to the neurotoxic cell lines, the antigen-antibody reaction is interfered and as the concentration of cranberry fruit extract increases the corresponding decrease in the expression of the alpha-synuclein protein can be seen. Conclusion: Cranberry fruit extract proves to effectively reduce the expression of alpha-synuclein protein in neurotoxic cell lines so further research must be conducted in this field to discover the useful effects of cranberry so that it can be used as a neuroprotective agent in medicines to treat neurodegenerative disorders.

Development of a Cranberry Standard for Quantification of Insoluble Cranberry (Vaccinium macrocarpon Ait.) Proanthocyanidins.

Posted: 
August 29, 2019
Authors: 
Gullickson ER; Krueger CG; Birmingham A; Maranan M; Reed JD.
Journal: 
Journal of Agricultural & Food Chemistry 10.1021/acs.jafc.9b03696 [doi]
Abstract: 

Cranberry proanthocyanidins (PACs) can be partitioned into soluble PACs, which are extracted with solvents, and insoluble PACs, which remain associated with fibers and proteins after extraction. Most research on cranberry products only quantifies soluble PACs because proper standards for quantifying insoluble PACs are lacking. In this study, we evaluated the ability of a cranberry PAC (c-PAC) standard, reflective of the structural heterogeneity of PACs found in cranberry fruit, to quantify insoluble PACs by the butanol-hydrochloric acid (BuOH-HCl) method. For the first time, a c-PAC standard enabled conversion of BuOH-HCl absorbance values (550 nm) to a weight (milligram) basis, allowing for quantification of insoluble PACs in cranberries. The use of the c-PAC reference standard for sequential analysis of soluble PACs by the method of 4-(dimethylamino)cinnamaldehyde and insoluble PACs by the method of BuOH-HCl provides analytical tools for the standardization of cranberry-based ingredients.

Evaluation of the Degree of Polymerization of the Proanthocyanidins in Cranberry by Molecular Sieving and Characterization of the Low Molecular Weight Fractions by UHPLC-Orbitrap Mass Spectrometry.

Posted: 
August 29, 2019
Authors: 
Gardana, C. Simonetti, P.
Journal: 
Molecules. 24(8):1504
Abstract: 

4-dimethylammino-cinnamaldehyde (DMAC) assays quantify total proanthocyanidins (PACs) but do not provide qualitative PAC molecular weight distribution information and cannot discriminate between A- and B-type PACs. We developed an efficient method for assessing PAC molecular weight distributions. The PACs from three commercial cranberry extracts (A1-A3) were fractionated by molecular sieves with cut-offs of 3, 10, 30, 50, and 100 kDa, and each fraction was analyzed by DMAC assays. A1, A2, and A3 contained 27%, 33%, and 15% PACs, respectively. Approximately 28 PACs, 20 flavonols, and 15 phenolic acids were identified by UHPLC-DAD-Orbitrap MS in A1 and A3, while A2 contained only flavan-3-ols. Epicatechin was the main monomer in A1 and A3, and catechin was the main in A2. Procyanidin A2 was the main dimer in A1 and A3, representing more than 85% of the total dimers, while it constituted approximately only 24% of A2. A1 and A3 contained quercetin, isorhamnetin, myricetin, and their glycosides, which were totally absent in A2. In A1 and A3 the PACs were mainly distributed in the fractions 30-3 and <3 kDa, while in A2 more than 70% were present in the fraction less than 3 kDa. Overall, obtained data strongly suggests that A2 is not cranberry-derived, or is adulterated with another source of PACs.

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