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

Deoxycholate induces COX-2 expression via Erk1/2-, p38-MAPK and AP-1-dependent mechanisms in esophageal cancer cells

<p>Abstract</p> <p>Background</p> <p>The progression from Barrett's metaplasia to adenocarcinoma is associated with the acquirement of an apoptosis-resistant phenotype. The bile acid deoxycholate (DCA) has been proposed to play an important role in the development of esophageal adenocarcinoma, but the precise molecular mechanisms remain undefined. The aim of this study was to investigate DCA-stimulated COX-2 signaling pathways and their possible contribution to deregulated cell survival and apoptosis in esophageal adenocarcinoma cells.</p> <p>Methods</p> <p>Following exposure of SKGT-4 cells to DCA, protein levels of COX-2, MAPK and PARP were examined by immunoblotting. AP-1 activity was assessed by mobility shift assay. DCA-induced toxicity was assessed by DNA fragmentation and MTT assay.</p> <p>Results</p> <p>DCA induced persistent activation of the AP-1 transcription factor with Fra-1 and JunB identified as the predominant components of the DCA-induced AP-1 complex. DCA activated Fra-1 via the Erk1/2- and p38 MAPK while Erk1/2 is upstream of JunB. Moreover, DCA stimulation mediated inhibition of proliferation with concomitant low levels of caspase-3-dependent PARP cleavage and DNA fragmentation. Induction of the anti-apoptotic protein COX-2 by DCA, via MAPK/AP-1 pathway appeared to balance the DCA mediated activation of pro-apoptotic markers such as PARP cleavage and DNA fragmentation. Both of these markers were increased upon COX-2 suppression by aspirin pretreatment prior to DCA exposure.</p> <p>Conclusion</p> <p>DCA regulates both apoptosis and COX-2-regulated cell survival in esophageal cells suggesting that the balance between these two opposing signals may determine the transformation potential of DCA as a component of the refluxate.</p

A multiscale systems perspective on cancer, immunotherapy, and Interleukin-12

Monoclonal antibodies represent some of the most promising molecular targeted immunotherapies. However, understanding mechanisms by which tumors evade elimination by the immune system of the host presents a significant challenge for developing effective cancer immunotherapies. The interaction of cancer cells with the host is a complex process that is distributed across a variety of time and length scales. The time scales range from the dynamics of protein refolding (i.e., microseconds) to the dynamics of disease progression (i.e., years). The length scales span the farthest reaches of the human body (i.e., meters) down to the range of molecular interactions (i.e., nanometers). Limited ranges of time and length scales are used experimentally to observe and quantify changes in physiology due to cancer. Translating knowledge obtained from the limited scales observed experimentally to predict patient response is an essential prerequisite for the rational design of cancer immunotherapies that improve clinical outcomes. In studying multiscale systems, engineers use systems analysis and design to identify important components in a complex system and to test conceptual understanding of the integrated system behavior using simulation. The objective of this review is to summarize interactions between the tumor and cell-mediated immunity from a multiscale perspective. Interleukin-12 and its role in coordinating antibody-dependent cell-mediated cytotoxicity is used illustrate the different time and length scale that underpin cancer immunoediting. An underlying theme in this review is the potential role that simulation can play in translating knowledge across scales

Interleukin-12 signalling pathways in human T lymphocytes

The aim of the work that constitutes the present PhD thesis was to elucidate the intracellular signalling pathways downstream of the cytokine Interleukin-12 (IL-12) in human T lymphocytes. IL-12 plays a key role in the onset of the cellular immune response by driving the differentiation of naive Th cells into Th1 cells. The important role of IL-12 in human immune responses has been emphasised by recent reports that mutations of the IL-12 receptor (IL-12R) are found in immunodeficient patients suffering from recurrent Mycoplasma sp. and Salmonella sp. infections. IL-12 exerts its function through interaction with the IL-12R that leads to the activation of the Janus kinases Tyk2 and Jak2. IL-12 also induces tyrosine phosphorylation and DNA binding of STAT4 (Signal Transducer and Activator of Transcription 4), an essential response for Th1 cell differentiation. We have extensively studied the regulation of the transcription factor STAT4 in response to IL-12, using PHA-activated human T cells isolated from healthy donors as a model. These studies include: a) characterisation of other possible signals that induced in response to IL-12, could regulate the activation of STAT4; b) a comparison between the characteristics of STAT4 activation in response to IL-12 and to Interferon-α (IFN-α); and c) study of the mechanism by which IL-12 induced STAT4 activation is potentiated by IL-2

Analysis of interleukin-2 signaling using affinity precipitations and polyacrylamide gel electrophoresis

Interleukin-2 (IL-2) was first discovered and characterized as T-cell growth factor (1). It is responsible for the growth of T cells and thus plays an important role in the proper functioning of the immune system. IL-2 has a high affinity receptor consisting of an α-, β-, and γ-chain. The γ- chain is shared with a number of other cytokines such as IL-3 and IL-7, which also play a role in cell growth (2). The IL-2 receptor β- chain also comprises part of the receptor for IL-15