MALDI-TOF peptidomic analysis of serum and post-prostatic massage urine specimens to identify prostate cancer biomarkers

Background

Lower urinary tract symptoms (LUTS) and prostate specific antigen-based parameters seem to have only a limited utility for the differential diagnosis of prostate cancer (PCa). MALDI-TOF/MS peptidomic profiling could be a useful diagnostic tool for biomarker discovery, although reproducibility issues have limited its applicability until now. The current study aimed to evaluate a new MALDI-TOF/MS candidate biomarker.

Methods

Within- and between-subject variability of MALDI-TOF/MS-based peptidomic urine and serum analyses were evaluated in 20 and 15 healthy donors, respectively. Normalizations and approaches for accounting below limit of detection (LOD) values were utilized to enhance reproducibility, while Monte Carlo experiments were performed to verify whether measurement error can be dealt with LOD data. Post-prostatic massage urine and serum samples from 148 LUTS patients were analysed using MALDI-TOF/MS. Regression-calibration and simulation and extrapolation methods were used to derive the unbiased association between peptidomic features and PCa.

Results

Although the median normalized peptidomic variability was 24.9%, the within- and between-subject variability showed that median normalization, LOD adjustment, and log₂ data transformation were the best combination in terms of reliability; in measurement error conditions, intraclass correlation coefficient was a reliable estimate when the LOD/2 was substituted for below LOD values. In the patients studied, 43 peptides were shared by the urine and serum, and several features were found to be associated with PCa. Only few serum features, however, show statistical significance after the multiple testing procedures were completed. Two serum fragmentation patterns corresponded to the complement C4-A.

Conclusions

MALDI-TOF/MS serum peptidome profiling was more efficacious with respect to post-prostatic massage urine analysis in discriminating PCa.

     

Emerging new crown symptoms on Castanea sativa (Mill.): Attempting to model interactions among pests and fungal pathogens

In the 2015–2016 growing seasons, two novel symptoms were assessed on the crown of trees in orchards and coppices of chestnut groves in Central Italy. The first symptom was flagging of annual shoots with green leaves undergoing sudden wilt and turning brown later in the season. The second symptom consisted of leaves on annual shoots turning yellow before wilting in absence of flagging represented the second symptom. Samples were collected along transects in early summer, late summer and winter, and processed in the laboratory. The flagging symptom was associated in early summer with the presence of C. parasitica in cryptic dried buds on stems from the previous year's growth. The pathogen was also found in dormant buds in winter, suggesting that the infection could take place in summer during the Chinese gall wasp oviposition period. Cryphonectria parasitica was also isolated from abandoned galls in winter supporting the hypothesis that galls are a potential source of inoculum for crown infections. Aetiology of yellowing was not clarified and no fungal taxa were specifically associated with this symptom. Gnomoniopsis castanea, C. parasitica and, in early summer, Colletotrichum acutatum were the most abundant fungal taxa isolated from chestnut shoots and buds.     

Proteomic identification of altered protein O-GlcNAcylation in a triple transgenic mouse model of Alzheimer's disease

PET scan analysis demonstrated the early reduction of cerebral glucose metabolism in Alzheimer disease (AD) patients that can make neurons vulnerable to damage via the alteration of the hexosamine biosynthetic pathway (HBP). Defective HBP leads to flawed protein O-GlcNAcylation coupled, by a mutual inverse relationship, with increased protein phosphorylation on Ser/Thr residues. Altered O-GlcNAcylation of Tau and APP have been reported in AD and is closely related with pathology onset and progression. In addition, type 2 diabetes patients show an altered O-GlcNAcylation/phosphorylation that might represent a link between metabolic defects and AD progression. Our study aimed to decipher the specific protein targets of altered O-GlcNAcylation in brain of 12-month-old 3×Tg-AD mice compared with age-matched non-Tg mice. Hence, we analysed the global O-GlcNAc levels, the levels and activity of OGT and OGA, the enzymes controlling its cycling and protein specific O-GlcNAc levels using a bi-dimensional electrophoresis (2DE) approach. Our data demonstrate the alteration of OGT and OGA activation coupled with the decrease of total O-GlcNAcylation levels. Data from proteomics analysis led to the identification of several proteins with reduced O-GlcNAcylation levels, which belong to key pathways involved in the progression of AD such as neuronal structure, protein degradation and glucose metabolism. In parallel, we analysed the O-GlcNAcylation/phosphorylation ratio of IRS1 and AKT, whose alterations may contribute to insulin resistance and reduced glucose uptake. Our findings may contribute to better understand the role of altered protein O-GlcNAcylation profile in AD, by possibly identifying novel mechanisms of disease progression related to glucose hypometabolism.     

Two-Photon Bidirectional Control and Imaging of Neuronal Excitability with High Spatial Resolution In Vivo

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Pas1 is a common lung cancer susceptibility locus in three mouse strains

Inherited predisposition to lung cancer is a phenotypic trait shared by different mouse inbred strains that show either a high or an intermediate predisposition. Other strains are instead genetically resistant. The Pas1 locus is the major determinant of lung cancer predisposition in the A/J strain (Gariboldi et al. 1993). To define the determinants of susceptibility to lung tumorigenesis in the highly susceptible SWR/J and in the intermediately susceptible BALB/c mice, we analyzed (BALB/c × SWR/J)F₂ and (BALB/c × C3H/He)F₂ crosses by genetic linkage experiments. The present results provide unequivocal evidence that the same Pas1/+ allele that leads to lung cancer predisposition is shared by A/J, SWR/J, and BALB/c strains. The intermediate susceptibility of the BALB/c strain would result by interaction of Pas1 locus with lung cancer resistance loci. Received: 18 April 1997 / Accepted: 15 June 1997     

8-Oxo-7,8-dihydro-2′-deoxyguanosine and other lesions along the coding strand of the exon 5 of the tumour suppressor gene P53 in a breast cancer case-control study

The next-generation sequencing studies of breast cancer have reported that the tumour suppressor P53 (TP53) gene is mutated in more than 40% of the tumours. We studied the levels of oxidative lesions, including 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG), along the coding strand of the exon 5 in breast cancer patients as well as in a reactive oxygen species (ROS)-attacked breast cancer cell line using the ligation-mediated polymerase chain reaction technique. We detected a significant ‘in vitro’ generation of 8-oxodG between the codons 163 and 175, corresponding to a TP53 region with high mutation prevalence, after treatment with xanthine plus xanthine oxidase, a ROS-generating system. Then, we evaluated the occurrence of oxidative lesions in the DNA-binding domain of the TP53 in the core needle biopsies of 113 of women undergoing breast investigation for diagnostic purpose. An increment of oxidative damage at the −G− residues into the codons 163 and 175 was found in the cancer cases as compared to the controls. We found significant associations with the pathological stage and the histological grade of tumours. As the major news of this study, this largest analysis of genomic footprinting of oxidative lesions at the TP53 sequence level to date provided a first roadmap describing the signatures of oxidative lesions in human breast cancer. Our results provide evidence that the generation of oxidative lesions at single nucleotide resolution is not an event highly stochastic, but causes a characteristic pattern of DNA lesions at the site of mutations in the TP53, suggesting causal relationship between oxidative DNA adducts and breast cancer.     

The impact of <Emphasis Type="Italic">Carpobrotus</Emphasis> cfr. <Emphasis Type="Italic">acinaciformis</Emphasis> (L.) L. Bolus on soil nutrients,microbial communities structure and native plant communities in Mediterranean ecosystems

Background and aims

Carpobrotus spp. are amongst the most impactful and widespread plant invaders of Mediterranean habitats. Despite the negative ecological impacts on soil and vegetation that have been documented, information is still limited about the effect by Carpobrotus on soil microbial communities. We aimed to assess the changes in the floristic, soil and microbial parameters following the invasion by Carpobrotus cfr. acinaciformis within an insular Mediterranean ecosystem.

Methods

Within three study areas a paired-site approach, comparing an invaded vs. a non-invaded plot, was established. Within each plot biodiversity indexes, C and N soil content, pH and microbial biomass and structure (bacterial and fungal) were assessed.

Results

Invaded plots showed a decrease of α-species richness and diversity. The least represented plant species in invaded plots were those related to grassland habitats. In all invaded soils, a significant increase of carbon and nitrogen content and a significant decrease of pH were registered. Carpobrotus significantly increased bacterial and fungal biomass and altered soil microbial structure, particularly favoring fungal growth.

Conclusions

Carpobrotus may deeply impact edaphic properties and microbial communities and, in turn, these strong modifications probably increase its invasive potential and its ability to overcome native species, by preventing their natural regeneration.

     

Radiocarbon-Based Assessment of Heterotrophic Soil Respiration in Two Mediterranean Forests

The amount of soil organic carbon (SOC) released into the atmosphere as carbon dioxide (CO₂), which is referred to as heterotrophic respiration (Rh), is technically difficult to measure despite its necessity to the understanding of how to protect and increase soil carbon stocks. Within this context, the aim of this study is to determine Rh in two Mediterranean forests dominated by pine and oak using radiocarbon measurements of the bulk SOC from different soil layers. The annual Rh was 3.22 Mg C ha^?1 y^?1 under pine and 3.13 Mg C ha^?1 y^?1 under oak, corresponding to 38 and 31% of the annual soil respiration, respectively. The accuracy of the Rh values was evaluated by determining the net primary production (NPP), as the sum of the Rh and the net ecosystem production measured by eddy covariance, then comparing it with the NPP obtained through independent biometric measurements. No significant differences were observed, which suggested the suitability of our methodology to infer Rh. Assuming the C inputs to soil to consist exclusively of the aboveground and belowground litter and the C output exclusively of the Rh, both soils were C sinks, which is consistent with a previous modeling study that was performed in the same stands. In conclusion, radiocarbon analysis of bulk SOC provided a reliable estimate of the average annual amount of soil carbon released to the atmosphere; hence, its application is convenient for calculating Rh because it utilizes only a single soil sampling and no time-consuming monitoring activities.