Effects of Sjogren's syndrome and high sugar diet on oral microbiome in patients with rampant caries: a clinical study.

OBJECTIVE: The purpose of this study was to assess the composition of the oral microbial flora of adults with rampant caries in China to provide guidance for treatment. PATIENTS AND METHODS: Sixty human salivary and supragingival plaque samples were collected. They were characterized into four groups: patients with rampant caries with Sjogren's syndrome (RC-SS) or high-sugar diet (RC-HD), common dental caries (DC), and healthy individuals (HP). The 16S rRNA V3-V4 region of the bacterial DNA was detected by Illumina sequencing. PCoA based on OTU with Bray-Curtis algorithm, the abundance of each level, LEfSe analysis, network analysis, and PICRUSt analysis were carried out between the four groups and two sample types. Clinical and demographic data were compared using analysis of variance (ANOVA) or the nonparametric Kruskal-Wallis rank-sum test, depending on the normality of the data, using GraphPad Prism 8 (P < 0.05). RESULTS: OTU principal component analysis revealed a significant difference between healthy individuals and those with RC-SS. In the saliva of patients with rampant caries, the relative abundance of Firmicutes increased significantly at the phylum level. Further, Streptocpccus, Veillonella, Prevotella, and Dialister increased, while Neisseria and Haemophilus decreased at the genus level. Veillonella increased in the plaque samples of patients with rampant caries. CONCLUSION: Both salivary and dental plaque composition were significantly different between healthy individuals and patients with rampant caries. This study provides a microbiological basis for exploring the etiology of rampant caries. CLINICAL RELEVANCE: This study provides basic information on the flora of the oral cavity in adults with rampant caries in China. These findings could serve as a reference for the treatment of this disease.

Effect of gal/GalNAc regioisomerism in galactosylated liposomes on asialoglycoprotein receptor-mediated hepatocyte-selective targeting in vivo.

In this study, we describe a novel synthesis of galactosylated lipids by lipase catalysis. Lactitol (Lac), galactose (Gal), or N-acetyl galactosamine (GalNAc) was coupled with cholesterol (CHS) as target head groups by enzyme-catalyzed regioselective esterification to produce three kinds of lipids: CHS-1-Gal, CHS-6-Gal, or CHS-6-GalNAc1. The biological effects of galactosylated lipids carrying different constitutional isomers of the pendent sugar species were investigated. LP-1-Gal (liposomes containing 5.0 molar% of CHS-1-Gal) showed strong binding to tetrameric lectins of Ricinus communis agglutinin (RCA120) in vitro, while LP-6-Gal (liposomes containing 5.0 molar% of CHS-6-Gal) and LP-6-GalNAc (liposomes containing 5.0 molar% of CHS-6-GalNAc) did not. After intravenous injection, LP-6-GalNAc, LP-1-Gal and LP-6-Gal rapidly disappeared from the blood and accumulated rapidly in liver (up to 74.88 ± 4.11%, 58.67 ± 5.75%, and 47.66 ± 4.56% of injected dose/g organ within 4 h, respectively). This is significantly higher than the uptake of unmodified liposomes (Unmod-LP) (18.67 ± 6.07%). Pre-injection of asialofetuin significantly inhibits liver uptake of Gal-liposomes (P < 0.01), with the degree of inhibition appearing in the following order: LP-6-GalNAc (73.29%) > LP-1-Gal (67.06%) > LP-6-Gal (53.61%). More importantly, LP-6-GalNAc was preferentially taken up by hepatocytes and the uptake ratio by parenchymal cells (PC) and nonparenchymal cells (NPC) (PC/NPC ratio) was 11.03 higher than LP-1-Gal (7.32), LP-6-Gal (5.83) and Unmod-LP (2.39). We suggest that liposomes containing the novel galactosylated lipid CHS-6-GalNAc have potential as drug delivery carriers for hepatocyte-selective targeting.

Neural epidermal growth factor-like 1 protein (Nell-1) alleviates periodontal tissue destruction in periodontitis by regulating the ratio of M2/M1 macrophage phenotypes.

BACKGROUND: Periodontitis is a common oral disease with high prevalence worldwide. Neural epidermal growth factor-like 1 protein (Nell-1) has recently been reported to have anti-inflammation effects and may be a drug candidate for osteoarthritis. However, its immunotherapeutic effects in periodontitis remain unknown. Therefore, this study aimed to investigate the effects of Nell-1 on periodontitis in terms of macrophage polarization and analyze its possible underlying mechanism. METHODS: A rat ligation-induced experimental periodontitis model was established and locally injected with Nell-1 (n = 6/group). Periodontal tissue destruction and macrophage polarization in vivo were analyzed using micro-CT, histology analysis, and western blot. Enzyme-linked immunosorbent assay was used to evaluate serum inflammatory cytokines. Then, the RAW 264.7 macrophage cells were treated with lipopolysaccharide (LPS), Nell-1, and the c-Jun N-terminal kinases (JNK) inhibitor (SP600125). RT-PCR, western blot, and flow cytometry were performed to further analyze the effect of Nell-1 on macrophage polarization and the underlying mechanism in vitro. RESULTS: Local treatment with Nell-1 significantly alleviated the destruction of alveolar bone and fibers in periodontitis, and upregulated the ratio of M2/M1 macrophages in periodontal tissues (P < 0.05). In vitro, Nell-1 at the concentrations of 200 and 500 ng/mL could significantly inhibit the expression of M1-related inflammatory factors in LPS-stimulated macrophages, and increase the expression of M2-related markers, regulating the macrophage phenotype switch into M2 (P < 0.05). The mRNA of JNK and relative protein level of phospho-JNK/JNK were also upregulated by Nell-1 (P < 0.05). Additionally, the JNK inhibitor (SP600125) could reverse the effect of Nell-1 on macrophage polarization (P < 0.05). CONCLUSIONS: Nell-1 could modulate the ratio of M2/M1 macrophages possibly through the JNK/MAPK signaling pathway, subsequently attenuating the inflammation and destruction of periodontal tissues caused by periodontitis.

Veillonella parvula acts as a pathobiont promoting the biofilm virulence and cariogenicity of Streptococcus mutans in adult severe caries.

Adult severe caries (ASC) brings severe oral dysfunction and treatment difficulties to patients, and yet no clear pathogenic mechanism for it has been found. This study is focused on the composition of dental plaque microbiome profiles in order to identify disease-relevant species and to investigate into their interactions with the S. mutans. Samples of dental plaque were collected for metagenomic analysis. The acidification, aciduricity, oxidative stress tolerance, and gtf (glucosyltransferase) gene expression of S. mutans cocultured with V. parvula which was identified as ASC-related dominant bacterium. The biofilm formation and extracellular exopolysaccharide (EPS) synthesis of dual-strain were analyzed with scanning electron microscopy (SEM), crystal violet (CV) staining, live/dead bacterial staining, and confocal laser scanning microscopy (CLSM). Furthermore, rodent model experiments were performed to validate the in vivo cariogenicity of the dual-species biofilm. The most significantly abundant taxon found associated with ASC was V. parvula. In vitro experiments found that V. parvula can effectively promote S. mutans mature biofilm formation with enhanced acid resistance, hydrogen peroxide detoxicity, and biofilm virulence. Rodent model experiments revealed that V. parvula was incapable of causing disease on its own, but it significantly heightened the biofilm virulence of S. mutans when being co-infected and augmented the progression, quantity, and severity of dental caries. Our findings demonstrated that V. parvula may act as a synergistic pathobiont to modulate the metabolic activity, spatial structure, and pathogenicity of biofilms of S. mutans in the context of ASC.IMPORTANCEAdult severe caries (ASC), as a special type of acute caries, is rarely reported and its worthiness of further study is still in dispute. Yet studies on the etiology of severe caries in adults have not found a clear pathogenic mechanism for it. Knowledge of the oral microbiota is important for the treatment of dental caries. We discovered that the interaction between V. parvula and S. mutans augments the severity of dental caries in vivo, suggesting V. parvula may act as a synergistic pathobiont exacerbating biofilm virulence of S. mutans in ASC. Our findings may improve the understanding of ASC pathogenesis and are likely to provide a basis for planning appropriate therapeutic strategies.

Targeted liposomes encapsulated iridium(III) compound greatly enhance anticancer efficacy and induce cell death via ferroptosis on HepG2 cells.

In this study, ligands 2-phenyl-1H-imidazo[4,5-f][1,10]phenanthroline (PIP), 2-(2-nitrophenyl)-1H-imidazo[4,5-f][1,10]phenanthroline (NPIP), 2-(2-nitronaphthalen-1-yl)-1H-imidazo[4,5-f][1,10]phenanthroline (NNIP) and their iridium(III) metal compounds [Ir(ppy)2(PIP)](PF6) (ppy = 2-phenylpyridine, 1a), [Ir(ppy)2(NPIP)](PF6) (1b), [Ir(ppy)2(NNIP)](PF6) (1c) were designed and synthesized. The anti-cancer activities of 1a, 1b and 1c on BEL-7402, HepG2, SK-Hep1 and non-cancer LO2 were detected using MTT method. 1a shows moderate, 1b and 1c display low or no anti-cancer activities. To elevate the anti-cancer effectiveness, encapsulating the compounds 1a, 1b and 1c into the ordinary or targeted liposomes to produce 1alip, 1blip, 1clip, or targeted 1aTlip, 1bTlip and 1cTlip. The IC50 values of 1alip, 1blip, 1clip, 1aTlip, 1bTlip and 1cTlip against HepG2 cells are 7.9 ± 0.1, 8.6 ± 0.2, 16.9 ± 0.5, 5.9 ± 0.2, 7.3 ± 0.1 and 9.7 ± 0.7 µM, respectively. Specifically, the anti-tumor activity assays in vivo found that the inhibitory rates are 23.24 % for 1a, 61.27 % for 1alip, 76.06 % for 1aTlip. It is obvious that the targeted liposomes entrapped iridium(III) compound greatly enhance anti-cancer efficacy. Additionally, 1alip, 1blip and 1clip or targeted 1aTlip, 1bTlip and 1cTlip can effectively restrain the cell colony and proliferation in the G0/G1 period. 1alip, 1blip, 1clip, 1aTlip, 1bTlip and 1cTlip can increase reactive oxygen species (ROS) concentration, arouse a decline in the mitochondrial membrane potential and promote Ca2+ release. RNA-sequence was applied to examine the signaling pathways. Taken together, the liposomes or targeted liposomes encapsulated compounds trigger cell death by way of apoptosis, autophagy, ferroptosis, disruption of mitochondrial function and PI3K/AKT/mTOR signaling pathways.

Effect of hydrophile-lipophile balance of the linker in Gal/GalNAc ligands on high-affinity binding of galactosylated liposomes by the asialoglycoprotein receptor.

In this study, we explored the effect of the hydrophile-lipophile balance (HLB) in the linker unit of Galactose (Gal)/N-acetylgalactosamine (GalNAc) ligands on their affinity toward asialoglycoprotein receptors (ASGPRs). Two Gal/GalNAc ligands with lipophilic linkers-{(5-cholesten-3b-ol)[(2-acetamido-2-deoxy-d-galactopyranose-6-o)sebacate]} (CHS-6-GalNAc) and {(5-cholesten-3b-ol)[(d-galactopyranose-6-o)sebacate]} (CHS-6-Gal)-and two with hydrophilic linkers-{(5-cholesten-yl)[(4-O-b-D-galactopyranosyl)-D-glucitol-6-yl]sebacate} (CHS-1-Gal) and {(5-cholesten-3a-ol)[(2-acetamido-2-deoxy-d-galactopyranose-6-o)3,6-dioxa-octanedioate]} (CHS-PEG2-6-GalNAc)-were synthesized by enzymatic catalysis. Compared with unmodified liposomes, all Gal/GalNAc ligand-modified liposomes showed higher efficiency toward the hepatocyte target as evaluated by weighted-average overall drug-targeting efficiency (Te*) in vivo and HepG2 cell uptake efficiency in vitro. The ligands containing linkers with high HLB values (i.e., CHS-PEG2-6-GalNAc and CHS-1-Gal) exhibited higher ASGPR affinity than those containing linkers with low HLB values (i.e., CHS-6-GalNAc and CHS-6-Gal). We used molecular-dynamics (MD) simulations to investigate the structure-activity relationship between the HLB value of the linker in a ligand and ASGPR affinity. MD simulation results indicated that a Gal/GalNAc ligand with a more hydrophilic linker (i.e., higher HLB value) unit tended to have a higher solvent-accessible surface area (SASA), leading to lower steric hindrance for effective ASGPR recognition. The results of this study will provide an improved design for Gal/GalNAc ligand-based surface-modified liposomes with high ASGPR affinity.

Removal of Cu2+ from aqueous solution by adsorption onto a novel activated nylon-based membrane.

A novel activated nylon-based membrane was prepared and applied as an adsorbent for the removal of Cu2+ from aqueous solutions. It involved three stages: (i) deposition of a chitosan layer that functionalized the nylon membrane, (ii) cross-linking with epichlorohydrin to stabilize the polymer layer and enabling grafting, and (iii) iminodiacetic acid grafting. SEM and EDX techniques were used to characterize the composition of the membranes. Dynamic adsorption experiments on membranes were carried out at various pH values, contact times, adsorption dosages and initial metal concentrations to determine optimum membrane adsorption properties. The adsorption isotherm relating to Cu2+ fitted the Langmuir equation and an adsorption equilibrium constant and adsorption capacity of 2.345x10(-3)mg/ml and 10.794mg/g were determined, respectively. The experimental data was analyzed using two adsorption kinetic models, pseudo-first-order and pseudo-second-order with the latter system providing the best fit. Finally complete regeneration of the activated nylon membrane was possible using 100mmol/l Na2EDTA.

Adsorption of papain with Cibacron Blue F3GA carrying chitosan-coated nylon affinity membranes.

Covalent coupling of chitosan (CS) to activated nylon membrane was performed after the reaction of the microporous nylon membrane with formaldehyde. Non-specific adsorption on the CS-coated nylon membrane decreased greatly, compared with plain nylon membrane. The dye Cibacron Blue F3GA (CB F3GA) as a ligand was then covalently immobilized on the CS-coated membranes. Physical properties of the composite membrane and its applications in affinity membrane chromatography were examined. The contents of CS and CB F3GA-attached membranes were 89.6 mg/g nylon membrane and 146.1 micromol/g nylon membrane, respectively. These CB F3GA-attached composite membranes were used in the papain adsorption studies. Higher papain adsorption capacity, up to 235.3mg/g affinity membrane, was obtained. The adsorption isotherm fitted the Freundlich model well. Significant amount of the adsorbed papain (about 94.3%) was eluted by 1.0M NaSCN at pH 9.0. Experiments on regeneration and dynamic adsorption were also performed. It appears that CB F3GA-CS nylon membranes can be applied for papain separation without causing any denaturation.

Pharmacokinetics and tissue distribution of docetaxel liposome mediated by a novel galactosylated cholesterol derivatives synthesized by lipase-catalyzed esterification in non-aqueous phase.

The purpose of this study is to synthesize a novel galactosylated cholesterol derivative, cholesterol-diethenyl decanedioate-lactitol (CHS-DD-LA) through lipase-catalyzed esterification in non-aqueous and to evaluate the preparation, pharmacokinetics and biodistribution of docetaxel (DOC) liposomes modified with CHS-DD-LA (G-DOC-L), which may actively gather at the liver compared with the conventional DOC liposomes (DOC-L) and commercial dosage form of DOC injection (DOC-i). A rapid and simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed for the determination of the DOC concentration in plasma and tissues with Taxol as the internal standard (IS). To measure the liver-targeting effect of the G-DOC-L, relative uptake rate (Re), peak concentration ratio (Ce), targeting efficiency (Te) and relative targeting efficiency (RTe) were reduced as the evaluation parameters. The results showed that the entrapment efficiency, particle size and Zeta potential of G-DOC-L was 76.8 ± 3.5%, 95.6 nm and 27.19 mV, respectively. After i.v. administration at the dose of 2.5 mg/kg in rats, a decrease in the AUC, MRT and an increase in CL (p < 0.05) were observed in the G-DOC-L group compared with DOC-L. All these results suggested that galactose-anchored liposomes could rapidly be removed from the circulation in vivo. The tissue distribution of G-DOC-L was widely different from that of DOC-L. The Re of G-DOC-L, DOC-L on liver was 4.011, 0.102; Ce was 3.391, 0.111; Te was 55.01, 3.08, respectively, demonstrating that G-DOC-L had an excellent effect on liver-targeting, which may help to improve the therapeutic effect of hepatic diseases.

Electrospun polyvinyl alcohol/carbon dioxide modified polyethyleneimine composite nanofiber scaffolds.

A novel biocompatible polyvinyl alcohol/carbon dioxide modified polyethyleneimine (PVA/PEI-CO2) composite nanofiber was fabricated by a green and facile protocol, which reduces the cytotoxicity of PEI through the surface modification of the PEI with CO2. The (13)C NMR spectrum, elemental analysis, and TGA show that CO2 has been incorporated in the PEI surface resulting in a relatively stable structure. The resulting PVA/PEI-CO2 composite nanofibers have been characterized by attenuated total reflection-Fourier transform infrared spectroscopy (ATR-FTIR), contact angle, and scanning electron microscopy (SEM). The results show that the average diameters of the nanofibers range from 265 ± 53 nm to 423 ± 80 nm. The cytotoxicity of PVA/PEI-CO2 composite nanofibers was assessed by cytotoxicity evaluation using the growth and cell proliferation of normal mice Schwann cells. SEM and the MTT assay demonstrated the promotion of cell growth and proliferation on the PVA/PEI-CO2 composite scaffold. It suggests that PEI-CO2 can have tremendous potential applications in biological material research.

Facile fabrication of P(OVNG-co-NVCL) thermoresponsive double-hydrophilic glycopolymer nanofibers for sustained drug release.

The thermoresponsive double-hydrophilic glycopolymer (DHG), Poly (6-O-vinyl-nonanedioyl-D-galactose-co-N-vinylcaprolactam) (P(OVNG-co-NVCL)) was synthesized via a chemo-enzymatic process and a free radical copolymerization and the resulting nanofibers were fabricated using an electrospinning process. The desired lower critical solution temperature (LCST) between 32 and 40 °C of the DHG polymers was achieved by adjusting the molar fraction of galactose monomer in the copolymers during the synthesis. The thermoresponsive DHG polymers were found to have good cytocompatibility with Hela cells as determined by the MTT assay, and special recognition of the protein peanut agglutinin (PNA). The drug release properties of these newly designed thermoresponsive DHG P(OVNG-co-NVCL) nanofibers are temperature regulated, can target specific proteins and have the potential application in the field of sustained drug release.

Fabrication and aggregation of thermoresponsive glucose-functionalized double hydrophilic copolymers.

Novel double-hydrophilic thermosensitive statistical glycopolymers, poly(N-isopropylacrylamide-co-6-O-vinyladipoyl-D-glucose), were fabricated using a chemoenzymatic process and free radical copolymerization. The structures of the glycopolymers were confirmed by (1)H and (13)C NMR, and their molar mass distributions determined by gel permeation chromatography. UV-vis spectroscopy data showed that the polymers exhibited reproducible temperature-responsive behavior. The self-assembly and critical aggregation concentration was verified by fluorescence spectroscopy with pyrene acting as a hydrophobic probe. Measurements by laser light scattering and transmission electron microscopy revealed that the glycopolymers were able to self-assemble into aggregates with varying particle sizes and morphologies in aqueous solutions.

Metal chelate affinity precipitation: purification of BSA using poly(N-vinylcaprolactam-co-methacrylic acid) copolymers.

This investigation involves the metal chelate affinity precipitation of bovine serum albumin (BSA) using a copper ion loaded thermo-sensitive copolymer. The copolymer of N-vinylcaprolactam with methacrylic acid PNVCL-co-MAA was synthesized by free radical polymerization in aqueous solution, and Cu(II) ions were attached to provide affinity properties for BSA. A maximum loading of 48.1mg Cu(2+) per gram of polymer was attained. The influence of pH, temperature, BSA and NaCl concentrations on BSA precipitation and of pH, ethylenediaminetetraacetic acid (EDTA) and NaCl concentrations on elution were systematically probed. The optimum conditions for BSA precipitation occurred when pH, temperature and BSA concentration were 6.0, 10°C and 1.0 mg/ml, respectively and the most favorable elution conditions were at pH 4.0, with 0.2M NaCl and 0.06 M EDTA. The maximum amounts of BSA precipitation and elution were 37.5 and 33.7 mg BSA/g polymer, respectively. It proved possible to perform multiple precipitation/elution cycles with a minimal loss of polymer efficacy. The results show that PNVCL-co-MAA is a suitable matrix for the purification of target proteins from unfractionated materials.

Optimization of adsorption conditions of BSA on thermosensitive magnetic composite particles using response surface methodology.

Thermosensitive core-shell magnetic composite particles with a magnetic silica core and a rich poly (N-vinylcaprolactam) (PNVCL) shell layer were developed for studying the adsorption of bovine serum albumin (BSA) in a batch system. Various analytical and spectroscopic techniques including SEM, FT-IR, VSM and DSC were used to characterize the adsorbents prepared in this study. The combined effects of operating parameters such as initial temperature, pH and initial BSA concentration on the adsorption were analyzed using response surface methodology. The optimum conditions were 40°C, pH 4.68, and initial BSA concentration 2.0 mg/mL. Desorption experiments were conducted by altering the system temperature where a high recovery rate of protein was obtained. The separation process developed here indicates that the dual-responsive smart adsorbent could be an ideal candidate for the separation of protein.

Optimization of adsorption conditions of papain on dye affinity membrane using response surface methodology.

The adsorption of papain on Reactive Blue 4 dye-ligand affinity membrane was investigated in a batch system. The combined effects of operating parameters such as initial pH, temperature, and initial papain concentration on the adsorption were analyzed using response surface methodology. The optimum adsorption conditions were determined as initial pH 7.05, temperature 39 degrees C, and initial papain concentration 11.0mg/ml. At optimum conditions, the adsorption capacity of dye-ligand affinity membrane for papain was found to be 27.85 mg/g after 120 min adsorption. The papain was purified 34.6-fold in a single step determined by fast protein liquid chromatography. More than 85% of the adsorbed papain was desorbed using 1.0M NaCl at pH 9.0 as the elution agent. The purification process showed that the dye-ligand immobilized composite membrane gave good separation of papain from aqueous solution.

Comparison: adsorption of papain using immobilized dye ligands on affinity membranes.

Novel membranes that involve the immobilization of Reactive Red 120 or Reactive Brown 10 as dye ligands were prepared. These were used in the purification of papain from papaya powder extracts. Papain adsorption capacities for the Red 120 and Brown 10 membranes were 143.6 mg/g and 107.3mg/g, respectively. The effectiveness of adsorption was demonstrated by Freundlich isotherm proficiency. The enzyme was eluted from the respective dye membranes using 1.0M NaCl at pH 6.0 and yields of over 80% were found for the Red 120-CS (chitosan)-nylon membrane whereas only a 50% recovery was possible using the Brown 10-CS-nylon membranes. It is concluded that Red 120-CS-nylon membranes could play an active role in the separation and purification of papain from crude extracts. This system has the potential to be developed for the commercial isolation of the protein.

Polyacrylonitrile fibers efficiently loaded with tamoxifen citrate using wet-spinning from co-dissolving solution.

Tamoxifen citrate (TAM)-loaded polyacrylonitrile (PAN) fibers were prepared using an improved wet-spinning technique. TAM was used as a model drug to evaluate the potential application of the loaded fiber system for drug delivery. PAN was first homogeneously dissolved in the N,N-dimethylacetamide (DMAc) solution containing TAM and then the co-dissolving solution was solidified to prepare the fibers using a wet-spinning method. Chemical, morphological and mechanical property characterizations were carried out, as well as the studies of the drug release properties. TAM was successfully encapsulated into a monofilament fiber, and this system was stable in terms of high loading capacity and effectiveness in release. The diameter of drug-loaded fiber was in the range of 40-60 microm. The best values of the tensile strength at 2.968 cN/dtex and breaking elongation at 14.9% of drug-loaded fibers were obtained when the drug loading content was 23.1 wt.%. These characteristics were suitable for the weaving process. The in vitro release experiment indicated that constant drug release from the fiber was observed for a long duration of time. Kinetic studies demonstrated that the system followed the Higuchi kinetics. These findings demonstrate that controlled release of drugs from PAN fibers could be potentially useful in drug delivery systems.