Human Properdin Opsonizes Nanoparticles and Triggers a Potent Pro-inflammatory Response by Macrophages without Involving Complement Activation.

Development of nanoparticles as tissue-specific drug delivery platforms can be considerably influenced by the complement system because of their inherent pro-inflammatory and tumorigenic consequences. The complement activation pathways, and its recognition subcomponents, can modulate clearance of the nanoparticles and subsequent inflammatory response and thus alter the intended translational applications. Here, we report, for the first time, that human properdin, an upregulator of the complement alternative pathway, can opsonize functionalized carbon nanotubes (CNTs) via its thrombospondin type I repeat (TSR) 4 and 5. Binding of properdin and TSR4+5 is likely to involve charge pattern/polarity recognition of the CNT surface since both carboxymethyl cellulose-coated carbon nanotubes (CMC-CNT) and oxidized (Ox-CNT) bound these proteins well. Properdin enhanced the uptake of CMC-CNTs by a macrophage cell line, THP-1, mounting a robust pro-inflammatory immune response, as revealed by qRT-PCR, multiplex cytokine array, and NF-κB nuclear translocation analyses. Properdin can be locally synthesized by immune cells in an inflammatory microenvironment, and thus, its interaction with nanoparticles is of considerable importance. In addition, recombinant TSR4+5 coated on the CMC-CNTs inhibited complement consumption by CMC-CNTs, suggesting that nanoparticle decoration with TSR4+5, can be potentially used as a complement inhibitor in a number of pathological contexts arising due to exaggerated complement activation.

Pulmonary surfactant protein SP-D opsonises carbon nanotubes and augments their phagocytosis and subsequent pro-inflammatory immune response.

Carbon nanotubes (CNTs) are increasingly being developed for use in biomedical applications, including drug delivery. One of the most promising applications under evaluation is in treating pulmonary diseases such as tuberculosis. Once inhaled or administered, the nanoparticles are likely to be recognised by innate immune molecules in the lungs such as hydrophilic pulmonary surfactant proteins. Here, we set out to examine the interaction between surfactant protein D (SP-D), a key lung pattern recognition molecule and CNTs, and possible downstream effects on the immune response via macrophages. We show here that a recombinant form of human SP-D (rhSP-D) bound to oxidised and carboxymethyl cellulose (CMC) coated CNTs via its C-type lectin domain and enhanced phagocytosis by U937 and THP-1 macrophages/monocytic cell lines, together with an increased pro-inflammatory response, suggesting that sequestration of SP-D by CNTs in the lungs can trigger an unwanted and damaging immune response. We also observed that functionalised CNTs, opsonised with rhSP-D, continued to activate complement via the classical pathway, suggesting that C1q, which is the recognition sub-component of the classical pathway, and SP-D have distinct pattern recognition sites on the CNTs. Consistent with our earlier reports, complement deposition on the rhSP-D opsonised CNTs led to dampening of the pro-inflammatory immune response by THP-1 macrophages, as evident from qPCR, cytokine array and NF-κB nuclear translocation analyses. This study highlights the importance of understanding the interplay between innate immune humoral factors including complement in devising nanoparticle based drug delivery strategies.

Systematic review of alcohol screening tools for use in the emergency department.

To ascertain which alcohol screening tool is most accurate in identifying alcohol misuse in patients in the emergency department a systematic review of diagnostic cohort studies of appropriate alcohol screening tools was performed. A thorough search of medical databases and relevant peer journals was conducted. Citation and author tracking was also utilised due to an initial paucity of relevant literature. Seven relevant papers were identified from this search, which allowed a review of the quality of the following alcohol screening tools: the fast alcohol screening tool (FAST), the Paddington alcohol test (PAT), the rapid alcohol problem screen (RAPS-4) and the TWEAK (where TWEAK is an acronym of the first letter of the key words in the questions of this screening tool: tolerance, worried, eye-opener, amnesia, K (cut-down)). The most sensitive screening tool within this review appears to be the FAST (93-94%), which has a specificity of 86-88% with a positive predicted value of 86-87%. Although the FAST appears to be the best for accurately identifying alcohol misuse within emergency department patients, it was assessed as a universal screening tool, and it may not be feasible (time or cost) to screen all who present to this service. In contrast, the PAT has been developed to be used on a select population within the emergency department and has already been shown to be cost-effective.

Effect of 24-h alcohol licensing on emergency departments: the South Yorkshire experience.

BACKGROUND: The alcohol Licensing Act (2003) was introduced to England and Wales on 23 November 2005. A single-centre study in 2007 from St Thomas's Hospital concluded that their alcohol-related attendances had significantly increased after the implementation of this new Act. This study aimed to assess whether this finding was reproduced in other hospitals. METHOD: A retrospective cohort study, reviewing anonymised routine data from four emergency departments (ED) in South Yorkshire, was undertaken. The study population was adults (over the age of 18 years) attending the ED with injuries or illnesses directly related to alcohol in the 12 months before and after the implementation of the Licensing Act (2003). The primary outcome was the number of these alcohol-related attendances. Secondary outcomes assessed whether there was any change in the timing of these presentations. RESULTS: Alcohol-related attendances, as detected by clinical coding, increased from 0.6% to 0.7% as a proportion of all attendances (95% CI 0.1 to 0.2, p<0.001). They increased by 0.4% at the Northern General Hospital and by 0.1% at Barnsley Hospital, decreased by 0.2% at Doncaster Royal Infirmary and did not significantly change at Rotherham General Hospital. The secondary outcome showed an unaltered peak time of 01:00 hours for alcohol-related attendances. CONCLUSION: Trends in alcohol-related attendances after the implementation of the Licensing Act (2003) varied across South Yorkshire hospitals and probably reflect local factors rather than any consistent impact from the Act.

An in vitro based investigation of the cytotoxic effect of water extracts of the Chinese herbal remedy LD on cancer cells.

BACKGROUND: Long Dan Xie Gan Wan (LD), a Chinese herbal remedy formulation, is traditionally used to treat a range of conditions, including gall bladder diseases, hepatitis, hyperthyroidism, migraines but it is not used for the management or treatment of cancer. However some of its herbal constituents, specifically Radix bupleuri, Radix scutellariae and Rhizoma alismatis have been shown to inhibit the growth of cancer cells. Thus, the aim of the study was to investigate the impact of LD on cancer cells in vitro. METHODS: HL60 and HT29 cancer cell lines were exposed to water extracts of LD (1:10, 1:50, 1:100 and/or 1:1000 prepared from a 3 mg/30 ml stock) and for both cell lines growth, apoptotic induction, alterations in cell cycle characteristics and genotoxicity were investigated. The specificity of the action of LD on these cancer cell lines was also investigated by determining its effect on human peripheral blood lymphocytes. Preliminary chemical analysis was carried out to identify cytotoxic constituents of LD using HPLC and LCMS. RESULTS: LD was significantly cytotoxic to, and induced apoptosis in, both cell lines. Apoptotic induction appeared to be cell cycle independent at all concentrations of LD used (1:10, 1:50 and 1:100) for the HL60 cell lines and at 1:10 for the HT29 cell line. At 1:50 and 1:100 apoptotic induction by LD appeared to be cell cycle dependent. LD caused significant genotoxic damage to both cell lines compared to their respective controls. The specificity study showed that LD exerted a moderate cytotoxic action against non-proliferating and proliferating blood lymphocytes but not apoptosis. Chemical analysis showed that a number of fractions were found to exert a significant growth inhibitory effect. However, the molecular weights of compounds within these fractions did not correspond to those from the herbal constituents of LD. CONCLUSION: It is possible that LD may have some chemotherapeutic potential. However, further studies are required to determine its cytotoxic constituents.

Apoptotic effect of Oldenlandia diffusa on the leukaemic cell line HL60 and human lymphocytes.

Oldenlandia diffusa is traditionally prescribed in the treatment of a number of cancers and studies suggest that it exerts a cytotoxic action specific to cancer cells. To further investigate this suggested action, the effect(s) of Oldenlandia diffusa on leukaemic cells (HL60) and stimulated and unstimulated human blood lymphocytes (PBLs) was investigated. For the HL60s, cell growth, apoptotic induction, alterations in cell cycle characteristics and genotoxicity were investigated. For the PBLs, apoptotic induction and alterations in cell cycle characteristics were investigated. Preliminary chemical analysis to identify the cytotoxic constituents of Oldenlandia diffusa was also carried out. Results showed that Oldenlandia diffusa significantly inhibited the growth of the HL60s and induced apoptosis in a cell cycle-independent fashion, possibly through the induction of genotoxic damage. Oldenlandia diffusa did not induce apoptosis in the PBLs however progression through the cell cycle was not evident (in stimulated PBLs) suggesting some degree of cytotoxicity. Preliminary chemical analysis indicated that a number of compounds appear to be responsible for the cytotoxic action of Oldenlandia diffusa. These results indicate that HL60s are more sensitive to Oldenlandia diffusa than stimulated PBLs and thus support a cytotoxic action for Oldenlandia diffusa that has some degree of specificity.