Inhibition of CSF-1R Supports T-Cell Mediated Melanoma Therapy

Tumor associated macrophages (TAM) can promote angiogenesis, invasiveness and immunosuppression. The cytokine CSF-1 (or M-CSF) is an important factor of TAM recruitment and differentiation and several pharmacological agents targeting the CSF-1 receptor (CSF-1R) have been developed to regulate TAM in solid cancers. We show that the kinase inhibitor PLX3397 strongly dampened the systemic and local accumulation of macrophages driven by B16F10 melanomas, without affecting Gr-1⁺ myeloid derived suppressor cells. Removal of intratumoral macrophages was remarkably efficient and a modest, but statistically significant, delay in melanoma outgrowth was observed. Importantly, CSF-1R inhibition strongly enhanced tumor control by immunotherapy using tumor-specific CD8 T cells. Elevated IFNγ production by T cells was observed in mice treated with the combination of PLX3397 and immunotherapy. These results support the combined use of CSF-1R inhibition with CD8 T cell immunotherapy, especially for macrophage-stimulating tumors.     

Establishment and Assessment of a New Human Embryonic Stem Cell‐Based Biomarker Assay for Developmental Toxicity Screening

A metabolic biomarker‐based in vitro assay utilizing human embryonic stem (hES) cells was developed to identify the concentration of test compounds that perturbs cellular metabolism in a manner indicative of teratogenicity. This assay is designed to aid the early discovery‐phase detection of potential human developmental toxicants. In this study, metabolomic data from hES cell culture media were used to assess potential biomarkers for development of a rapid in vitro teratogenicity assay. hES cells were treated with pharmaceuticals of known human teratogenicity at a concentration equivalent to their published human peak therapeutic plasma concentration. Two metabolite biomarkers (ornithine and cystine) were identified as indicators of developmental toxicity. A targeted exposure‐based biomarker assay using these metabolites, along with a cytotoxicity endpoint, was then developed using a 9‐point dose–response curve. The predictivity of the new assay was evaluated using a separate set of test compounds. To illustrate how the assay could be applied to compounds of unknown potential for developmental toxicity, an additional 10 compounds were evaluated that do not have data on human exposure during pregnancy, but have shown positive results in animal developmental toxicity studies. The new assay identified the potential developmental toxicants in the test set with 77% accuracy (57% sensitivity, 100% specificity). The assay had a high concordance (≥75%) with existing in vivo models, demonstrating that the new assay can predict the developmental toxicity potential of new compounds as part of discovery phase testing and provide a signal as to the likely outcome of required in vivo tests.     

Parental Transfer of the Antimicrobial Protein LBP/BPI Protects Biomphalaria glabrata Eggs against Oomycete Infections

Vertebrate females transfer antibodies via the placenta, colostrum and milk or via the egg yolk to protect their immunologically immature offspring against pathogens. This evolutionarily important transfer of immunity is poorly documented in invertebrates and basic questions remain regarding the nature and extent of parental protection of offspring. In this study, we show that a lipopolysaccharide binding protein/bactericidal permeability increasing protein family member from the invertebrate Biomphalaria glabrata (BgLBP/BPI1) is massively loaded into the eggs of this freshwater snail. Native and recombinant proteins displayed conserved LPS-binding, antibacterial and membrane permeabilizing activities. A broad screening of various pathogens revealed a previously unknown biocidal activity of the protein against pathogenic water molds (oomycetes), which is conserved in human BPI. RNAi-dependent silencing of LBP/BPI in the parent snails resulted in a significant reduction of reproductive success and extensive death of eggs through oomycete infections. This work provides the first functional evidence that a LBP/BPI is involved in the parental immune protection of invertebrate offspring and reveals a novel and conserved biocidal activity for LBP/BPI family members.     

Effect of silencing the two major tomato fruit pectin methylesterase isoforms on cell wall pectin metabolism

Post‐harvest storage is largely limited by fruit softening, a result of cell wall degradation. Pectin methylesterase (PE) (EC 3.1.1.11) is a major hydrolase responsible for pectin de‐esterification in the cell wall, a response to fruit ripening. Two major PE isoforms, PE1 and PE2, have been isolated from tomato (Solanum lycopersicon) pericarp tissue and both have previously been down‐regulated using antisense suppression. In this paper, PE1 and PE2 double antisense tomato plants were successfully generated through crossing the two single antisense lines. In the double antisense fruit, approximately 10% of normal PE activity remained and ripening associated pectin de‐esterification was almost completely blocked. However, double antisense fruit softened normally during ripening. In tomato fruit, the PE1 isoform was found to contribute little to total PE activity and have little effect on the degree of esterification of pectin. In contrast, the other dominant fruit isoform, PE2, has a major impact on de‐esterification of total pectin. PE2 appears to act on non‐CDTA‐soluble pectin during ripening and on CDTA‐soluble pectin before the start of ripening in a potentially block‐wise fashion.     

Associations between COPD related manifestations: a cross-sectional study

Background

Cardiovascular disease, osteoporosis and emphysema are associated with COPD. Associations between these factors and whether they predict all-cause mortality in COPD patients are not well understood. Therefore, we examined associations between markers of cardiovascular disease (coronary artery calcification [CAC], thoracic aortic calcification [TAC] and arterial stiffness), bone density (bone attenuation of the thoracic vertebrae), emphysema (PI-950 and 15th percentile) and all-cause mortality in a COPD cohort.

Methods

We assessed CAC, TAC, bone attenuation of the thoracic vertebrae, PI-950 and 15th percentile on low-dose chest computed tomography in COPD subjects. We measured arterial stiffness as carotid-radial pulse wave velocity (PWV), and identified deaths from the national register.

Results

We studied 119 COPD subjects; aged 67.8 ±7.3, 66% were males and mean FEV₁% predicted was 46.0 ±17.5. Subjects were classified into three pre-specificed groups: CAC = 0 (n = 14), 0 < CAC ≤ 400 (n = 41) and CAC > 400 (n = 64). Subjects with higher CAC were more likely to be older (p < 0.001) and male (p = 0.03), and more likely to have higher systolic blood pressure (p = 0.001) and a history of hypertension (p = 0.002) or ischemic heart disease (p = 0.003). Higher CAC was associated with higher PWV (OR 1.62, p = 0.04) and lower bone attenuation (OR 0.32, p = 0.02), but not with 15th percentile, after adjustment for age, sex and pack-years of smoking. In a Cox proportional hazards model, CAC, TAC and 15th percentile predicted all-cause mortality (HR 2.01, 2.09 and 0.66, respectively).

Conclusions

Increased CAC was associated with increased arterial stiffness and lower bone density in a COPD cohort. In addition, CAC, TAC and extent of emphysema predicted all-cause mortality.

Trial registration

Lothian NHS Board, Lothian Research Ethics Committee, LREC/2003/8/28.     

The quantitative genetic basis of sex ratio variation in Nasonia vitripennis: a QTL study

Our understanding of how natural selection should shape sex allocation is perhaps more developed than for any other trait. However, this understanding is not matched by our knowledge of the genetic basis of sex allocation. Here, we examine the genetic basis of sex ratio variation in the parasitoid wasp Nasonia vitripennis, a species well known for its response to local mate competition (LMC). We identified a quantitative trait locus (QTL) for sex ratio on chromosome 2 and three weaker QTL on chromosomes 3 and 5. We tested predictions that genes associated with sex ratio should be pleiotropic for other traits by seeing if sex ratio QTL co-occurred with clutch size QTL. We found one clutch size QTL on chromosome 1, and six weaker QTL across chromosomes 2, 3 and 5, with some overlap to regions associated with sex ratio. The results suggest rather limited scope for pleiotropy between these traits.     

Effect of postactivation potentiation on swimming starts in international sprint swimmers

The aim of this study was to investigate the effects of postactivation potentiation (PAP) on swim start performance (time to 15 m) in a group of international sprint swimmers. Nine international sprint swimmers (7 men and 2 women) volunteered and gave informed consent for this study, which was approved by the university ethics committee. Initially, swimmers performed a countermovement jump (CMJ) on a portable force platform (FP) at baseline and at the following time points ～15 seconds, 4, 8, 12, and 16 minutes after a PAP stimulus (1 set of 3 repetitions at 87% 1 repetition maximum [RM]) to individually determine the recovery time required to observe enhanced muscle performance. On 2 additional days, swimmers performed a swim start to 15 m under 50-m freestyle race conditions, which was preceded by either their individualized race specific warm-up or a PAP stimulus (1 set of 3 repetitions at 87% 1RM). Both trials were recorded on 2 cameras operating at 50 Hz with camera 1 located at the start and camera 2 at the 15-m mark. Peak vertical force (PVF) and peak horizontal force (PHF) were measured during all swim starts from a portable FP placed on top of the swim block. A repeated measures analysis of variance revealed a significant time effect with regard to power output (PO) (F = 20.963, p < 0.01) and jump height (JH) (F = 14.634, p < 0.01) with a paired comparison indicating a significant increase in PO and JH after 8 minutes of recovery from the PAP stimulus. There was a significant increase in both PHF and PVF after the PAP stimulus compared to the swim-specific warm-up during the swim start (PHF 770 ± 228 vs. 814 ± 263 N, p = 0.018; PVF: 1,462 ± 280 vs. 1,518 ± 311 N, p = 0.038); however, time to 15 m was the same when both starts were compared (7.1 ± 0.8 vs. 7.1 ± 0.8 seconds, p = 0.447). The results from this study indicate that muscle performance during a CMJ is enhanced after a PAP stimulus providing adequate recovery (～8 minutes) is given between the 2 activities. In addition, this study demonstrated that swimmers performed equally well in terms of time to 15 m when a PAP stimulus was compared to their individualized race specific warm-up and indicates that PAP may be a useful addition to a warm-up protocol before races. However, more research is required to fully understand the role PAP plays in swim performance.     

Iduna protects the brain from glutamate excitotoxicity and stroke by interfering with poly(ADP-ribose) polymer-induced cell death

Glutamate acting on N-methyl-D-aspartate (NMDA) receptors induces neuronal injury following stroke, through activation of poly(ADP-ribose) polymerase-1 (PARP-1) and generation of the death molecule poly(ADP-ribose) (PAR) polymer. Here we identify Iduna, a previously undescribed NMDA receptor-induced survival protein that is neuroprotective against glutamate NMDA receptor-mediated excitotoxicity both in vitro and in vivo and against stroke through interfering with PAR polymer-induced cell death (parthanatos). Iduna's protective effects are independent and downstream of PARP-1 activity. Iduna is a PAR polymer-binding protein, and mutation at the PAR polymer binding site abolishes the PAR binding activity of Iduna and attenuates its protective actions. Iduna is protective in vivo against NMDA-induced excitotoxicity and middle cerebral artery occlusion-induced stroke in mice. To our knowledge, these results define Iduna as the first known endogenous inhibitor of parthanatos. Interfering with PAR polymer signaling could be a new therapeutic strategy for the treatment of neurologic disorders.