Activation of NALP1 inflammasomes in rats with adjuvant arthritis; a novel therapeutic target of carboxyamidotriazole in a model of rheumatoid arthritis

Background and Purpose

Pro-inflammatory cytokines are important in rheumatoid arthritis (RA) and their production is mainly regulated by NF-κB and inflammasomes. Carboxyamidotriazole (CAI) exhibits potent anti-inflammatory activities by decreasing cytokines. Here, we have investigated NACHT, LRR and PYD domains-containing protein (NALP) inflammasomes in a rat model of RA and explored the therapeutic effects of CAI in this model and the involvement of NF-κB and inflammasomes in the actions of CAI.

Experimental Approach

The anti-arthritic effects of CAI were assessed in the adjuvant arthritis (AA) model in rats, using radiological and histological techniques. NALP1 and NALP3 inflammasomes, NF-κB pathway and pro-inflammatory cytokines levels were measured with Western blots, immunohistochemistry and elisa.

Key Results

CAI decreased the arthritis index, improved radiological and histological changes, and reduced synovial IL-1β, IL-6, IL-18 and TNF-α levels in rats with AA. Compared with normal rats, the 70 kDa NALP1 isoform was up-regulated, NALP3 was down-regulated, and levels of the 165 kDa NALP1 isoform and the adaptor protein ASC were unchanged in synovial tissue from AA rats. CAI reduced the 70 kDa NALP1 isoform and restored NALP3 levels in AA rats; CAI inhibited caspase-1 activation in AA synovial tissue, but not its enzymic activity in vitro. In addition, CAI reduced expression of p65 NF-κB subunit and IκBα phosphorylation and degradation in AA rats.

Conclusion and Implications

NALP1 inflammasomes were activated in synovial tissues from AA rats and appeared to be a novel therapeutic target for RA. CAI could have therapeutic value in RA by inhibiting activation of NF-κB and NALP1 inflammasomes and by decreasing pro-inflammatory cytokines.     

Design of melt-recyclable poly(ε-caprolactone)-based supramolecular shape-memory nanocomposites

A novel poly(epsilon-caprolactone) (PCL) supramolecular network exhibiting shape-memory behavior was successfully constructed with pendant UPy units that are highly able to dimerize. The dynamic network was obtained by a simple and versatile strategy consisting of chain-extension reaction between α,ω-dihydroxyoligoPCL and hydroxylated UPy units in the presence of hexamethylene diisocyanate as a coupling agent and further intermolecular dimerization of the UPy along the polyurethane backbone. ¹H NMR analyses confirmed the dynamic features of the system, and DMTA in tensile mode was investigated to assess the SMP properties. Recyclability was also assessed by taking advantage of these supramolecular networks. Further addition of cellulose nanocrystals into the polymer network enabled adjustment of the extent of the net-points and therefore the SMP features. As confirmed by dispersion tests in solution and SEM observations, these bio-based nanofillers were homogeneously distributed in the network via supramolecular interaction between the hydroxyl groups present on their surface and UPy moieties along the polyurethane backbone. Thus, the here developed nanomaterials might reveal applicability in areas where a combination of SMP and biocompatibility is needed.

Novel melt-recyclable poly(ε-caprolactone)/cellulose nanocrystals supramolecular nanocomposite networks with shape-memory behavior have been successfully constructed by playing with UPy chemistry.     

HepaRG culture in tethered spheroids as an in vitro three‐dimensional model for drug safety screening

Conventional two‐dimensional (2D) monolayer cultures of HepaRG cells allow in vitro maintenance of many liver‐specific functions. However, cellular dedifferentiation and functional deterioration over an extended culture period in the conventional 2D HepaRG culture have hampered its applications in drug testing. To address this issue, we developed tethered spheroids of HepaRG cells on Arg–Gly–Asp (RGD) and galactose‐conjugated substratum with an optimized hybrid ratio as an in vitro three‐dimensional (3D) human hepatocyte model. The liver‐specific gene expression level and drug metabolizing enzyme activities in HepaRG‐tethered spheorids were markedly higher than those in 2D cultures throughout the culture period of 7 days. The inducibility of three major cytochrome P450 (CYP) enzymes, namely CYP1A2, CYP2B6 and CYP3A4, was improved in both mRNA and activity level in tethered spheroids. Drug‐induced cytotoxic responses to model hepatotoxins (acetaminophen, chlorpromazine and ketoconazole) in tethered spheroids were comparable to 2D cultures as well as other studies in the literature. Our results suggested that the HepaRG‐tethered spheroid would be an alternative in vitro model suitable for drug safety screening. Copyright © 2014 John Wiley & Sons, Ltd.     

Quantitative toxicoproteomic analysis of zebrafish embryos exposed to a retinoid X receptor antagonist UVI3003

Retinoid X receptor (RXR) antagonists, including some environmental endocrine disruptors, have a teratogenic effect on vertebrate embryos. To investigate the toxicological mechanism on the protein expression level, a quantitative proteomic study was conducted to analyze the proteome alterations of zebrafish (Danio rerio) embryos exposed to gradient concentrations of a representative RXR antagonist UVI3003. Using isobaric Tags for Relative and Absolute Quantitation (iTRAQ) labeling coupled nano high‐performance liquid chromatography‐tandem mass spectrometry (nano HPLC‐MS/MS), in total 6592 proteins were identified, among which 195 proteins were found to be differentially expressed by more than a two‐fold change in exposed groups compared with the control. Gene ontology analysis showed that these differential proteins were mostly involved in anatomical structure development, biosynthetic process, ion binding and oxidoreductase activity. Moreover, the biological pathways of translation, lipoprotein metabolism, cell survival and gluconeogenesis were intensively inhibited after exposure. Some significantly downregulated proteins such as apolipoprotein A‐I and vitellogenin and upregulated proteins such as calcium activated nucleotidase 1b, glutathione S‐transferase and glucose 6‐dehydrogenases showed a strong dose‐dependent response. The results provided new insight into the molecular details of RXR antagonist‐induced teratogenicity and added novel information of pathways and potential biomarkers for evaluation of RXR interfering activity. Copyright © 2015 John Wiley & Sons, Ltd.     

Effect of inflammatory factor‐induced cyclo‐oxygenase expression on the development of reperfusion‐related no‐reflow phenomenon in acute myocardial infarction

No reflow after reperfusion therapy for myocardial infarction is a strong predictor of clinical outcome. Increased levels of inflammatory factors, including C‐reactive protein (CRP), in patients with acute myocardial infarction (AMI) undergoing primary percutaneous coronary intervention (PCI) may affect myocardial perfusion. However, why the no‐reflow phenomenon increases in inflammation stress after PCI is not clear. The aim of the present study was to determine the effects and molecular mechanisms underlying the effects of CRP on the expression of cyclo‐oxygenase (COX) on the development of the no‐reflow phenomenon. There was a significant increase in plasma levels of CRP and interleukin (IL)‐6 in no‐reflow patients, suggesting that inflammatory factors play an important role in the development of the no‐reflow phenomenon. The mechanisms involved were further evaluated after reperfusion in a rat model mimicking the no‐reflow phenomenon. Compared with normal reflow rats, there were significant increases in both COX‐1 and COX‐2 in cardiac tissue from no‐reflow rats. The COX inhibitor indomethacin (5 mg/kg, i.p.) significantly reduced the no‐reflow area. In another series of experiments, human coronary artery endothelial cells (HCAEC) were treated with CRP at clinically relevant concentrations (5–25 μg/mL). C‐Reactive protein significantly increased COX‐1 and COX‐2 levels in a time‐ and concentration‐dependent manner. In addition, extracellular signal‐regulated kinase (ERK) and Jun N‐terminal kinase (JNK) were activated in CRP (5, 10, 25 μg/mL)‐treated HCAEC cultures. Furthermore, the ERK inhibitor pd98059 (30 μmol/L) and the JNK inhibitor sp600125 (10 μmol/L) blocked CRP‐induced COX‐1 and COX‐2 expression for 12 h. Together, the findings of the present study suggest that CRP can promote the development of the no‐reflow phenomenon by increasing COX‐1 and COX‐2 expression, which is regulated, in part, via ERK and JNK activity.     

Higher intake of carotenoid is associated with a lower risk of colorectal cancer in Chinese adults: a case–control study

Purpose

The associations between specific carotenoid intake and colorectal cancer risk remain inconsistent. The aim of this study was to examine the association between specific dietary carotenoid intake with colorectal cancer risk in Chinese adults.

Method

From July 2010 to October 2013, 845 eligible colorectal cancer cases and 845 frequency-matched controls (age and sex) completed in-person interviews. A validated food frequency questionnaire was used to estimate dietary intake. Multivariate logistical regression models were used to calculate the odds ratio (OR) and 95 % confidence intervals (95 % CIs) of colorectal cancer risk after adjusting for various confounders.

Results

A strong inverse association was found between β-cryptoxanthin intake and colorectal cancer risk. Compared with the lowest quartile, the highest quartile intake showed a risk reduction of 77 % (OR 0.23, 95 % CI 0.17–0.33, P _trend < 0.01) after adjustment for various confounding variables. The inverse associations were also observed for α-carotene (OR 0.50, 95 % CI 0.37–0.68, P _trend < 0.01), β-carotene (OR 0.67, 95 % CI 0.49–0.91, P _trend < 0.01), and lycopene (OR 0.51, 95 % CI 0.37–0.70, P _trend < 0.01). There was no statistically significant association between lutein/zeaxanthin intake and colorectal cancer risk. These findings were consistent across cancer site, sources of controls, and smoking status. The inverse associations between dietary α-carotene, β-cryptoxanthin, and lycopene intake and colorectal cancer risk were found in both males and females, while inverse associations between β-carotene intake and colorectal cancer risk were only observed in males.

Conclusions

Consumption of α-carotene, β-carotene, β-cryptoxanthin, and lycopene was inversely associated with colorectal cancer risk. No significant association was found between lutein/zeaxanthin intake and colorectal cancer risk.

     

Photoluminescence mechanism and applications of Zn-doped carbon dots

Heteroatom-doped carbon dots (CDs) with excellent optical characteristics and negligible toxicity have emerged in many applications including bioimaging, biosensing, photocatalysis, and photothermal therapy. The metal-doping of CDs using various heteroatoms results in an enhancement of the photophysics but also imparts them with multifunctionality. However, unlike nonmetal doping, typical metal doping results in low fluorescence quantum yields (QYs), and an unclear photoluminescence mechanism. In this contribution, we detail results concerning zinc doped CDs (Zn-CDs) with QYs of up to 35%. The zinc ion charges serve as a surface passivating agent and prevent the aggregation of graphene π–π stacking, leading to an increase in the QY of the Zn-CDs. Structural and chemical investigations using spectroscopic and first principle simulations further revealed the effects of zinc doping on the CDs. The robust Zn-CDs were used for the ultra-trace detection of Hg²⁺ with a detection limit of 0.1 μM, and a quench mechanism was proposed. The unique optical properties of the Zn-CDs have promise for use in applications such as in vivo sensing and future phototherapy applications.

Zinc ions, acting as a surface passivating agent, prevented the aggregation of graphene π–π stacking and increased the quantum yield of Zn-carbon dots.     

Reduction responsive and surface charge switchable polyurethane micelles with acid cleavable crosslinks for intracellular drug delivery

Previously we synthesized redox sensitive polyurethane micelles, core crosslinked by diisocyanates (PU-CCL). To improve the intracellular drug release and tumor cellular toxicity of anticancer drugs loaded into polyurethane micelles, we now describe redox sensitive polyurethane micelles with tunable surface charge switchabilities, crosslinked with pH cleavable Schiff bonds, as anticancer drug carriers. Different amounts of 1,6-diaminohexane were connected onto the pendant carboxyl groups of amphiphilic multi-blocked polyurethane (PU-SS-COOH), resulting in polyurethanes with various ratios of pendant carboxyl and amine groups (denoted as PU-SS-COOH-NH₂-1, PU-SS-COOH-NH₂-2 and PU-SS-COOH-NH₂-3). The surface charge switched as the pH was increased for PU-SS-COOH-NH₂-1, PU-SS-COOH-NH₂-2 and PU-SS-COOH-NH₂-3. Then the PU-SS-COOH-NH₂-3 micelles, dissolved in water, were crosslinked by glutaraldehyde resulting in surface charge switchable and reduction responsive polyurethane micelles with acid cleavable crosslinks (PU-ACCL). The crosslinked polyurethane micelles (PU-ACCL) demonstrated superior particle stability in phosphate buffered saline (PBS, pH = 7.4) solution without reducing agents, whereas the drug release rate was markedly accelerated by the addition of glutathione (GSH). Notably, the drug release from PU-ACCL was further accelerated in acidic fluid as the result of acid induced cleavage of the crosslinks. In vitro cytotoxicity studies demonstrated that doxorubicin (DOX)-loaded PU-ACCL micelles displayed increased cytotoxicity against tumor cells which was comparable to that obtained for DOX loaded into uncrosslinked polyurethane micelles. The reduction responsive and surface charge switchable polyurethane micelles with acid cleavable crosslinks, which have superior extracellular stability and provide rapid intracellular drug release, may hold great potential as a bio-triggered drug delivery system for cancer therapy.

Previously we synthesized redox sensitive polyurethane micelles, core crosslinked by diisocyanates (PU-CCL).