Adhesion molecules and inflammatory injury

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/154291/1/fsb2008008008.pd

Role of lateral cell–cell border location and extracellular/transmembrane domains in PECAM/CD31 mechanosensation

Phosphorylation of tyrosine residues on platelet–endothelial cell adhesion molecule-1 (PECAM-1), followed by signal trans- 13 duction events, has been described in endothelial cells following exposure to hyperosmotic and fluid shear stress. However, it is 14 unclear whether PECAM-1 functions as a primary mechanosensor in this process. Utilizing a PECAM-1–null EC-like cell line, we 15 examined the importance of cellular localization and the extracellular and transmembrane domains in PECAM-1 phosphorylation 16 responses to mechanical stress. Tyrosine phosphorylation of PECAM-1 was stimulated in response to mechanical stress in null cells 17 transfected either with full length PECAM-1 or with PECAM-1 mutants that do not localize to the lateral cell–cell adhesion site and 18 that do not support homophilic binding between PECAM-1 molecules. Furthermore, null cells transfected with a construct that 19 contains the intact cytoplasmic domain of PECAM-1 fused to the extracellular and transmembrane domains of the interleukin-2 20 receptor also underwent mechanical stress-induced PECAM-1 tyrosine phosphorylation. These findings suggest that mechano- 21 sensitive PECAM-1 may lie downstream of a primary mechanosensor that activates a tyrosine kinase

Targeting fibroblast activation protein in tumor stroma with chimeric antigen receptor T cells can inhibit tumor growth and augment host immunity without severe toxicity.

The majority of chimeric antigen receptor (CAR) T-cell research has focused on attacking cancer cells. Here, we show that targeting the tumor-promoting, nontransformed stromal cells using CAR T cells may offer several advantages. We developed a retroviral CAR construct specific for the mouse fibroblast activation protein (FAP), comprising a single-chain Fv FAP [monoclonal antibody (mAb) 73.3] with the CD8α hinge and transmembrane regions, and the human CD3ζ and 4-1BB activation domains. The transduced muFAP-CAR mouse T cells secreted IFN-γ and killed FAP-expressing 3T3 target cells specifically. Adoptively transferred 73.3-FAP-CAR mouse T cells selectively reduced FAP(hi) stromal cells and inhibited the growth of multiple types of subcutaneously transplanted tumors in wild-type, but not FAP-null immune-competent syngeneic mice. The antitumor effects could be augmented by multiple injections of the CAR T cells, by using CAR T cells with a deficiency in diacylglycerol kinase, or by combination with a vaccine. A major mechanism of action of the muFAP-CAR T cells was the augmentation of the endogenous CD8(+) T-cell antitumor responses. Off-tumor toxicity in our models was minimal following muFAP-CAR T-cell therapy. In summary, inhibiting tumor growth by targeting tumor stroma with adoptively transferred CAR T cells directed to FAP can be safe and effective, suggesting that further clinical development of anti-human FAP-CAR is warranted

The Synthetic Lignan Secoisolariciresinol Diglucoside Prevents Asbestos-Induced NLRP3 Inflammasome Activation in Murine Macrophages

Background. The interaction of asbestos with macrophages drives two key processes that are linked to malignancy: (1) the generation of reactive oxygen species (ROS)/reactive nitrogen species (RNS) and (2) the activation of an inflammation cascade that drives acute and chronic inflammation, with the NLRP3 inflammasome playing a key role. Synthetic secoisolariciresinol diglucoside (SDG), LGM2605, is a nontoxic lignan with anti-inflammatory and antioxidant properties and was evaluated for protection from asbestos in murine peritoneal macrophages (MF). Methods. MFs were exposed to crocidolite asbestos ± LGM2605 given 4 hours prior to exposure and evaluated at various times for NLRP3 expression, secretion of inflammasome-activated cytokines (IL-1β and IL-18), proinflammatory cytokines (IL-6, TNFα, and HMGB1), NF-κB activation, and levels of total nitrates/nitrites. Results. Asbestos induces a significant (p<0.0001) increase in the NLRP3 subunit, release of proinflammatory cytokines, NLRP3-activated cytokines, NF-κB, and levels of nitrates/nitrites. LGM2605 significantly reduced NLRP3 ranging from 40 to 81%, IL-1β by 89–96%, and TNFα by 67–78%, as well as activated NF-κB by 48-49% while decreasing levels of nitrates/nitrites by 85–93%. Conclusions. LGM2605 reduced asbestos-induced NLRP3 expression, proinflammatory cytokine release, NF-κB activation, and nitrosative stress in MFs supporting its possible use in preventing the asbestos-induced inflammatory cascade leading to malignancy

The HDAC inhibitor panobinostat (LBH589) inhibits mesothelioma and lung cancer cells in vitro and in vivo with particular efficacy for small cell lung cancer

Lung cancer is the leading cause of cancer deaths in the United States. Current therapies are inadequate. Histone deacetylase inhibitors (HDACi) are a recently developed class of anticancer agents that cause increased acetylation of core histones and nonhistone proteins leading to modulation of gene expression and protein activityin - volved in cancer cell growth and survival pathways. We examined the efficacyof the HDACi panobinostat (LBH589) in a wide range of lung cancers and mesotheliomas. Panobinostat was cytotoxic in almost all 37 cancer cell lines tested. IC50 and LD50 values were in the low nmol/L range (4–470 nmol/L; median, 20 nmol/L). Small cell lung cancer (SCLC) cell lines were among the most sensitive lines, with LD50 values consistently <25 nmol/L. In lung cancer and mesothelioma animal models, panobinostat significantlyde creased tumor growth byan average of 62% when compared with vehicle control. Panobinostat was equallye ffective in immunocompetent and severe combined immunodeficiencymic e, indicating that the inhibition of tumor growth by panobinostat was not due to direct immunologic effects.Panobinostat was, however, particularlyeffective in SCLC xenografts, and the addition of the chemotherapyag ent etoposide augmented antitumor effects. Protein analysis of treated tumor biopsies revealed elevated amounts of cell cycle regulators such as p21 and proapoptosis factors, such as caspase 3 and 7 and cleaved poly[ADP-ribose] polymerase, coupled with decreased levels of antiapoptotic factors such as Bcl-2 and Bcl-XL. These studies together suggest that panobinostat maybe a useful adjunct in the treatment of thoracic malignancies, especiallySCLC

Human Skin/SCID Mouse Chimeras as an In Vivo Model for Human Cutaneous Mast Cell Hyperplasia

Human skin xenografted to mice with severe combined immunodeficiency syndrome (SCID) was evaluated to determine the integrity and fate of human dermal mast cells. There was an approximately 3-fold increase in number of dermal mast cells by 3 mo after engraftment (p < 0.05). These cells were responsive to conventional mast cell secretagogues and were confirmed to be of human origin by ultrastructural characterization of granule substructure and by reactivity for the human mast cell proteinase, chymase. CD1a+ Langerhans cells, also bone marrow–derived cells, failed to show evidence of concomitant hyperplasia, and increased mast cell number was not associated with alterations in number of dermal vascular profiles identified immunohistochemically for human CD31. RT-PCR analysis demonstrated human but not murine stem cell factor (SCF; also termed mast cell growth factor, c-kit ligand) mRNA in xenografts. Epidermal reactivity for stem cell factor protein shifted from a cytoplasmic pattern to an intercellular pattern by 3 mo after engraftment, suggesting a secretory phenotype, as previously documented for human cutaneous mastocytosis. The majority (>90%) of mast cells demonstrated membrane reactivity for human SCF at the time points of peak hyperplasia. These data establish SCID mouse recipients of human skin xenografts as a potential in vivo model for cutaneous mast cell hyperplasia

Peripheral Immune Cell Gene Expression Predicts Survival of Patients with Non-Small Cell Lung Cancer

Prediction of cancer recurrence in patients with non-small cell lung cancer (NSCLC) currently relies on the assessment of clinical characteristics including age, tumor stage, and smoking history. A better prediction of early stage cancer patients with poorer survival and late stage patients with better survival is needed to design patient-tailored treatment protocols. We analyzed gene expression in RNA from peripheral blood mononuclear cells (PBMC) of NSCLC patients to identify signatures predictive of overall patient survival. We find that PBMC gene expression patterns from NSCLC patients, like patterns from tumors, have information predictive of patient outcomes. We identify and validate a 26 gene prognostic panel that is independent of clinical stage. Many additional prognostic genes are specific to myeloid cells and are more highly expressed in patients with shorter survival. We also observe that significant numbers of prognostic genes change expression levels in PBMC collected after tumor resection. These post-surgery gene expression profiles may provide a means to re-evaluate prognosis over time. These studies further suggest that patient outcomes are not solely determined by tumor gene expression profiles but can also be influenced by the immune response as reflected in peripheral immune cells

Novel variants in the PRDX6 Gene and the risk of Acute Lung Injury following major trauma

<p>Abstract</p> <p>Background</p> <p>Peroxiredoxin 6 (<it>PRDX6</it>) is involved in redox regulation of the cell and is thought to be protective against oxidant injury. Little is known about genetic variation within the PRDX6 gene and its association with acute lung injury (ALI). In this study we sequenced the <it>PRDX6 </it>gene to uncover common variants, and tested association with ALI following major trauma.</p> <p>Methods</p> <p>To examine the extent of variation in the <it>PRDX6 </it>gene, we performed direct sequencing of the 5' UTR, exons, introns and the 3' UTR in 25 African American cases and controls and 23 European American cases and controls (selected from a cohort study of major trauma), which uncovered 80 SNPs. <it>In silico </it>modeling was performed using Patrocles and Transcriptional Element Search System (TESS). Thirty seven novel and tagging SNPs were tested for association with ALI compared with ICU at-risk controls who did not develop ALI in a cohort study of 259 African American and 254 European American subjects that had been admitted to the ICU with major trauma.</p> <p>Results</p> <p>Resequencing of critically ill subjects demonstrated 43 novel SNPs not previously reported. Coding regions demonstrated no detectable variation, indicating conservation of the protein. Block haplotype analyses reveal that recombination rates within the gene seem low in both Caucasians and African Americans. Several novel SNPs appeared to have the potential for functional consequence using <it>in silico </it>modeling. Chi²analysis of ALI incidence and genotype showed no significant association between the SNPs in this study and ALI. Haplotype analysis did not reveal any association beyond single SNP analyses.</p> <p>Conclusions</p> <p>This study revealed novel SNPs within the <it>PRDX6 </it>gene and its 5' and 3' flanking regions via direct sequencing. There was no association found between these SNPs and ALI, possibly due to a low sample size, which was limited to detection of relative risks of 1.93 and above. Future studies may focus on the role of <it>PRDX6 </it>genetic variation in other diseases, where oxidative stress is suspected.</p