Promising valorisation method of chitin biomass by producing 5-hydroxymethylfurfural using microwave hydrothermal treatment.

Chitin biomass is the second largest biomass resource on Earth but under-utilized. In this study, pretreated shrimp shells were converted into value-added platform chemical 5-hydroxymethylfurfural (HMF) using microwave hydrothermal treatment. Under the combined pretreatment of acid decalcification at room temperature and microwave-assisted alkali deacetylation, the HMF yield could reach 1.8 wt%. The key process parameters, including the holding temperature, holding time, and pH value, were evaluated and optimised. The highest HMF yield of 6.5 wt% was obtained at 202.6°C at a holding time of 5.8 min and a pH value of 1.5. This result demonstrates the potential of synchronously treating waste and recycling it, thereby offering a highly promising valorisation strategy for chitin-biomass utilisation.

A threshold longitudinal Tobit quantile regression model for identification of treatment-sensitive subgroups based on interval-bounded longitudinal measurements and a continuous covariate.

Identification of a subgroup of patients who may be sensitive to a specific treatment is an important problem in precision medicine. This article considers the case where the treatment effect is assessed by longitudinal measurements, such as quality of life scores assessed over the duration of a clinical trial, and the subset is determined by a continuous baseline covariate, such as age and expression level of a biomarker. Recently, a linear mixed threshold regression model has been proposed but it assumes the longitudinal measurements are normally distributed. In many applications, longitudinal measurements, such as quality of life data obtained from answers to questions on a Likert scale, may be restricted in a fixed interval because of the floor and ceiling effects and, therefore, may be skewed. In this article, a threshold longitudinal Tobit quantile regression model is proposed and a computational approach based on alternating direction method of multipliers algorithm is developed for the estimation of parameters in the model. In addition, a random weighting method is employed to estimate the variances of the parameter estimators. The proposed procedures are evaluated through simulation studies and applications to the data from clinical trials.

Conventional Magnetic Resonance Features for Predicting 1p19q Codeletion Status of World Health Organization Grade II and III Diffuse Gliomas.

PURPOSE: The conventional magnetic resonance features of World Health Organization (WHO) grade II and III diffuse gliomas in relation to chromosome 1p and 19q deletions (1p19q codeletion) were analyzed. METHODS: We identified 147 cases of WHO grade II and III diffuse gliomas (1p/19q codeletion, 36 cases; no 1p/19q codeletion, 111 cases). χ Test and univariate and multivariate binary logistic regression analyses were conducted to evaluate the association between the imaging features and 1p19q codeletion status of WHO grade II and III diffuse gliomas in the discovery group, including the WHO grade II and III subgroups. RESULTS: (1) In the entire population, multivariate regression demonstrated that proportion contrast-enhanced tumor (>5% vs ≤5%; odds ratio [OR], 0.169; P = 0.009), enhancing margin (poorly vs well defined; OR, 12.435; P = 0.002), and hemorrhage (yes vs no; OR, 21.082; P < 0.001) were associated with a higher incidence of 1p19q codeletion status. The nomogram showed good discrimination (area under the curve [AUC], 0.803) and calibration. (2) For grade II tumors, subgroup analysis found that enhancing margin (poorly vs well defined; OR, 0.308; P = 0.007) and subventricular zone (presence vs absence-; OR, 0.137; P < 0.001) were associated with a higher incidence of 1p19q codeletion status (AUC, 0.779). (3) For grade III tumors, subgroup analysis found that age (≥40 years vs <40 years; OR, 5.977; P = 0.03) and hemorrhage (yes vs no; OR, 18.051; P < 0.001) were associated with a higher incidence of 1p19q codeletion status (AUC, 0.816). CONCLUSIONS: Conventional magnetic resonance features can be conveniently used to facilitate the preoperative prediction of 1p19q codeletion status of WHO grade II and III diffuse gliomas. Decision curve analysis demonstrated that the nomogram was clinically useful.

Soft and Hard Tissue Changes Following Immediate Placement or Immediate Restoration of Single-Tooth Implants in the Esthetic Zone: A Systematic Review and Meta-Analysis.

PURPOSE: This systematic review aimed to compare immediate protocols with conventional protocols of single-tooth implants in terms of changes in the surrounding hard and soft tissue in the esthetic area. MATERIALS AND METHODS: Electronic and manual searches were performed in PubMed, EMBASE, Cochrane, and other data systems for research articles published between January 2001 and December 2014. Only randomized controlled trials (RCTs) reporting on hard and or soft tissue characteristics following a single-tooth implant were included. Based on the protocol used in each study, the included studies were categorized into three groups to assess the relationships between the factors and related esthetic indexes. Variables such as marginal bone level changes (mesial, distal, and mean bone level), peri-implant soft tissue changes (papilla level, midbuccal mucosa, and probing depth), and other esthetic indices were taken into consideration. The data were analyzed using RevMan version 5.3, Stata 12, and GRADEpro 3.6.1 software. RESULTS: A total of 13 RCTs met the inclusion criteria. Four studies examined immediate implant placement, five studies examined immediate implant restoration, and four studies examined immediate loading. Comparing the bone level changes following immediate and conventional restoration, no significant differences were found in the bone level of the mesial site (standard mean difference [SMD] = -0.04 mm; 95% confidence interval [CI]: -0.25 to 0.17 mm), the distal site (SMD = -0.15 mm; 95% CI: -0.38 to 0.09 mm), and the mean bone level changes (SMD = 0.05 mm; 95% CI: -0.18 to 0.27 mm). The difference in the marginal bone level changes between immediate and conventional loading was also not statistically significant (SMD = -0.05 mm; 95% CI: -0.15 to 0.06 mm for the mesial site and SMD = -0.02 mm; 95% CI: -0.09 to 0.05 mm for the distal site). Soft tissue changes following immediate and conventional restoration reported no significant differences in the papillae level of the mesial site (SMD = 0.18 mm; 95% CI: -0.00 to 0.37 mm), the papillae level of the distal site (SMD = -0.12 mm; 95% CI: -0.34 to 0.09 mm), and the midbuccal mucosa (SMD = -0.22 mm; 95% CI: -1.29 to 0.85 mm). CONCLUSION: Within the limitations, it can be concluded that immediately placed, restored, or loaded single-tooth implants in the esthetic zone result in similar hard and soft tissue changes compared with conventional protocols.

Controlled release of simvastatin-loaded thermo-sensitive PLGA-PEG-PLGA hydrogel for bone tissue regeneration: in vitro and in vivo characteristics.

Reports on the local delivery of drug loaded injectable hydrogels for bone regeneration are currently limited. This study assessed the effect of controlled simvastatin (SIM) release from a thermo-sensitive hydrogel in vitro and in vivo. We successfully manufactured and evaluated thermo-sensitive poly(d,l-lactide-co-glycolide)-poly(ethylene glycol)-poly(d,l-lactide-co-glycolide) triblock copolymers (PLGA-PEG-PLGA) loaded with SIM. The osteogenic effect of this hydrogel was tested in vitro and in vivo. MC-3T3 E1 cells proliferation and osteoblastic differentiation was analyzed after cultivation with the hydrogel extracts. Cells co-cultured with SIM/PLGA-PEG-PLGA extracts showed an increase in mineralization and osteogenic gene expression compared to the other two groups. Additionally, the characteristics of this composite in vivo were demonstrated using a rat bone defect model. The bone defects injected with SIM/PLGA-PEG-PLGA hydrogel showed increased new bone formation compared to samples treated with PLGA-PEG-PLGA and control samples. The results of this study suggest that SIM/PLGA-PEG-PLGA might provide potential therapeutic value for bone healing.

The effects of dried root aqueous extract of Salvia miltiorrhiza and its major ingredient in acceleration of orthodontic tooth movement in rat.

OBJECTIVES: Salvia miltiorrhiza (SM) is a popular and classic herb in traditional Chineses medicines. The objective is to confirm the effects of aqueous extract of S. miltiorrhiza (ESM) and its main ingredient on the promotion of orthodontic tooth movement and healing of periodontal ligament in rat. MATERIALS AND METHODS: Male Sprague-Dawley rats (n= 150) were divided into five groups: model control group (0.5 ml/kg phosphate-buffered saline (PBS) injection), ESM group (0.75 g/kg/day of crude drugs) and Danshensu subgroups (250, 500, 750 mg/kg/day of body weight). All rats were administered intramuscularly into the buccal vestibular mucosa of first molar of left maxillary. The indicators such as the moving distance of orthodontic tooth, nuclear factor κB ligand (RANKL) and osteoprotegerin (OPG) expression and osteoclasts were tested. RESULTS: The expressions of RANKL and OPG in the treatment groups were obviously enhanced compared with control group (P<0.05). The increase rate of OPG expression was slower than that of RANKL. But, RANKL decreased conspicuously after no orthodontic pressure was applied, especially in the treatment groups (Danshengsu high dose group at day 30: 2.17 versus 3.47 of control, P<0.01). ESM groups promoted osteoclasts proliferation in the first 20 days. CONCLUSION: There is a relationship between RANKL/OPG ratio and the number of osteoclasts. ESM might accelerate periodontal alteration of rat orthodontic tooth via producing more osteoclasts.

Development and validation of an antibody-dependent cell-mediated cytotoxicity-reporter gene assay.

Humanized monoclonal antibodies (mAbs) are the fastest growing class of biological therapeutics that are being developed for various medical indications, and more than 30 mAbs are already approved and in the market place. Antibody-dependent cell-mediated cytotoxicity (ADCC) is an important biological function attributed to the mechanism of action of several therapeutic antibodies, particularly oncology targeting mAbs. The ADCC assay is a complicated and highly variable assay. Thus, the use of an ADCC assay as a lot release test or a stability test for clinical trial batches of mAbs has been a substantial challenge to install in quality control laboratories. We describe here the development and validation of an alternate approach, an ADCC-reporter gene assay that is based on the key attributes of the PBMC-based ADCC assay. We tested the biological relevance of this assay using an anti-CD20 based model and demonstrated that this ADCC-reporter assay correlated well with standard ADCC assays when induced with the drugable human isotypes [IgG1, IgG2, IgG4, IgG4S > P (S228P) and IgG4PAA (S228P, F234A, L235A)] and with IgG1 isotype variants with varying amounts of fucosylation. This data demonstrates that the ADCC-reporter gene assay has performance characteristics (accuracy, precision and robustness) to be used not only as a potency assay for lot release and stability testing for antibody therapeutics, but also as a key assay for the characterization and process development of therapeutic molecules.

The rheumatoid arthritis shared epitope increases cellular susceptibility to oxidative stress by antagonizing an adenosine-mediated anti-oxidative pathway.

We have recently demonstrated that the rheumatoid arthritis (RA) shared epitope (SE) acts as a ligand that triggers nitric oxide (NO) signaling in opposite cells. Given the known pro-oxidative effect of NO and the proposed role of oxidative stress in the pathogenesis of RA, this study explores whether SE-triggered signaling can increase cellular oxidative stress. cAMP levels, adenylyl cyclase activity, and protein kinase A activity were measured using commercial kits. Generation of reactive oxygen species (ROS) was quantified using the fluorochrome dichlorofluorescein diacetate. Oxidative DNA damage was quantified using the single-cell electrophoresis technique. Here, we report that cells exposed to cell surface SE-positive HLA-DR (human leukocyte antigen-DR) molecules, to cell-free recombinant proteins genetically engineered to express the SE motif, or to SE-positive synthetic peptide showed diminished cAMP-dependent signaling, increased ROS levels, and higher vulnerability to oxidative DNA damage. Introduction of single amino acid substitutions into SE-positive peptides revealed a consensus five-amino acid sequence motif of Q/R-K/R-X-X-A that is necessary and sufficient for SE-triggered signaling. The pro-oxidative effect of the SE could be reversed by inhibiting NO production. We conclude that the SE acts as a signaling ligand that activates an NO-mediated pro-oxidative pathway. The potential contribution of this signaling aberration to RA pathogenesis is discussed.

Aberrant Gi protein coupled receptor-mediated cell survival signaling in rheumatoid arthritis B cell lines.

Sphingosine 1-phosphate (S1P) is a pleiotropic bioactive lipid that transmits potent signals through a family of G protein coupled receptors with resultant anti-apoptotic and pro-angiogenic effects. We have recently reported that lymphoblastoid B cell lines (LCLs) from rheumatoid arthritis (RA) patients are resistant to Fas-mediated cell death due to over-production of S1P, secondary to over-activity of sphingosine kinase-1 (SphK1). Here we investigated the signaling events that S1P triggers in those cells. Our results show that RA-derived LCLs display increased constitutive enzymatic activity of phosphatidylinositol 3-kinase (PI3K). Incubation of LCLs with a PI3K inhibitor wortmannin reversed PI3K over-activity and the resistance to Fas-mediated cell death. Incubation of RA LCLs with nanomolar concentration of S1P triggered exaggerated activation of both SphK and PI3K in RA LCLs compared to control cells. PI3K was mapped upstream of SphK, since wortmannin could block SphK activation by S1P. S1P signaling effect could be blocked by the Gi/G0 protein inhibitor, pertussis toxin and by an inhibitor of S1P-receptor interaction, suramin. S1P receptor expression levels did not appear to be the cause of disparate S1P-triggered signaling, since LCLs from RA patients and their healthy twin controls did not show statistically significant differences in the expression levels of the five known S1P receptors, as determined by quantitative real time reverse transcription-polymerase chain reaction analyses. Thus, we conclude that Fas death signaling aberration in RA LCLs is caused by extracellular S1P, which triggers PI3K-dependent SphK over-activity through a Gi protein-coupled receptor-mediated signaling cascade.

Sphingosine kinase 1-mediated inhibition of Fas death signaling in rheumatoid arthritis B lymphoblastoid cells.

OBJECTIVE: It is becoming increasingly apparent that B cells play an important role in the pathogenesis of rheumatoid arthritis (RA). Due to the scarcity of B cells in RA, it has been technically difficult to functionally characterize B cell apoptosis in this disease. As a necessary first step to identify candidate aberrations, we investigated Fas-mediated signaling events in immortalized peripheral blood B lymphoblastoid cell lines (LCLs) from patients with RA and controls. METHODS: Cell death was determined by the MTS assay, and apoptosis was detected by the TUNEL assay and DNA laddering. Proteolytic activation of caspase 3 was determined by immunoblotting, and its enzymatic activity was determined by a fluorometric technique. Messenger RNA (mRNA) expression was quantified by real-time polymerase chain reaction (PCR) analysis. The functional role of sphingosine kinase (SPHK) was determined by measuring its enzymatic activity, by quantifying the levels of its product, sphingosine 1-phosphate (S1P), and by investigating the ability of the SPHK inhibitor N,N-dimethylsphingosine and isozyme-specific small interfering RNA (siRNA) oligonucleotides to reverse signaling aberrations. RESULTS: LCLs from patients with RA displayed disease-specific Fas-mediated signal transduction impairment with consequent resistance to cell death. RA LCLs displayed high constitutive SPHK activity and increased levels of S1P. Real-time PCR analysis showed higher SPHK-1 mRNA expression levels in RA patients compared with paired controls. Increased SPHK-1 (but not SPHK-2) mRNA levels were observed in synovial tissue from RA patients. Competitive inhibitors of SPHK reversed the resistance of RA LCLs to Fas-induced apoptosis. Additionally, resistance to Fas-mediated signaling was reversed by siRNA oligonucleotides specific for SPHK-1 but not by oligonucleotides specific for SPHK-2. CONCLUSION: These findings demonstrate disease-specific resistance to Fas-mediated death signaling in patients with RA and implicate increased SPHK-1 activity as the cause of this aberration.

Sphingosine-1-phosphate induces early response gene expression in C6 glioma cells.

Sphingosine-1-phosphate (S1P) caused dose-dependent and time-dependent increases in c-fos mRNA. Pretreatment with pertussis toxin (PTX; 100 ng/mlx24 h) reduced c-fos activation by S1P (100 microM-187+/-6% vs. 411+/-27%) and lysophosphatidic acid (LPA; 100 microM-90+/-34% vs. 188+/-41%), but not by sphingosylphosphorylcholine (SPC; 100 microM-390+/-47% vs. 420+/-44%). RT-PCR analysis and sequencing demonstrated the presence of previously unidentified LPA-responsive Endothelial Differentiation Gene (EDG) receptor mRNAs in C6 cells: EDG-2 and EDG-4.