Significance: The ability to assign biological sex to human skeletal remains is a fundamental requirement in archaeology, paleoanthropology, and medico-legal sciences. While DNA sequencing can be used, it is expensive, time-consuming, and often fails due to the poor quality of the remaining DNA. An easier, more reliable, and consistently applicable method is needed. We present a method for sex determination of human remains using peptides retrieved from tooth enamel. Amelogenin is an enamel-forming protein encoded for by both chromosomes X and Y, with slight differences in their amino acid sequences. Peptides with these differences were identified by nanoflow liquid chromatography mass spectrometry and found to correctly assign sex to archaeological human remains of various chronological ages, from hundreds to thousands of years old.

Cells subjected to sustained high osmolarity almost universally respond by accumulating compatible organic osmolytes that, in contrast to inorganic ions, are not deleterious even at high intracellular concentrations. Their accumulation from the external environment by known organic osmolyte transporters, such as the four identified in mammals, occurs only slowly in response to sustained high osmolarity, by synthesis of new transporter proteins. Most cells, however, are not subject to high or varying osmolarity, and it is not clear whether organic osmolytes are generally required at normal osmolarities or how they are regulated. The fertilized egg of the mouse is protected in the oviduct from perturbations in osmolarity. However, deleterious effects of osmotic stress were evidentin vitroeven at normal oviductal osmolarity. Glycine was found to protect development, indicating that early mouse embryos may use glycine as an organic osmolyte at physiological osmolarity. We have now found that GLYT1, a glycine transporter of the neurotransmitter transporter gene family, functions as the organic osmolyte transporter that mediates the osmotically regulated accumulation of glycine and regulates cell volume in early embryos. Furthermore, osmotic stimulation of GLYT1 transport was immediate, without a requirement for protein synthesis, implying regulation different from known organic osmolyte transporters. Thus, GLYT1 appears to have a previously unidentified role as an organic osmolyte transporter that functions in acute organic osmolyte and volume homeostasis near normal osmolarity.

Glioblastoma (GBM) is one of the most aggressive solid tumors for which treatment options and biomarkers are limited. Small extracellular vesicles (sEVs) produced by both GBM and stromal cells are central in the inter-cellular communication that is taking place in the tumor bulk. As tumor sEVs are accessible in biofluids, recent reports have suggested that sEVs contain valuable biomarkers for GBM patient diagnosis and follow-up. The aim of the current study was to describe the protein content of sEVs produced by different GBM cell lines and patient-derived stem cells. Our results reveal that the content of the sEVs mirrors the phenotypic signature of the respective GBM cells, leading to the description of potential informative sEV-associated biomarkers for GBM subtyping, such as CD44. Overall, these data could assist future GBM in vitro studies and provide insights for the development of new diagnostic and therapeutic methods as well as personalized treatment strategies.

Tooth enamel is the hardest, densest and most mineralized tissue in vertebrates.

Inflammation, characterized by the activation of both resident and infiltrated immune cells, is accompanied by increased production of oxidizing and nitrating species. Nitrogen dioxide, the proximal nitrating species formed under these conditions, reacts with unsaturated fatty acids to yield nitroalkene derivatives. These electrophilic products modulate protein function via post-translational modification of susceptible nucleophilic amino acids. Nitroalkenes react with Keap1 to instigate Nrf2 signaling, activate heat shock response gene expression, and inhibit NF-κB-mediated signaling, inducing net anti-inflammatory and tissue-protective metabolic responses. We report the purification and characterization of a NADPH-dependent liver enzyme that reduces the nitroalkene moiety of nitro-oleic acid, yielding the inactive product nitro-stearic acid. Prostaglandin reductase-1 (PtGR-1) was identified as a nitroalkene reductase by protein purification and proteomic studies. Kinetic measurements, inhibition studies, immunological and molecular biology approaches as well as clinical analyses confirmed this identification. Overexpression of PtGR-1 in HEK293T cells promoted nitroalkene metabolism to inactive nitroalkanes, an effect that abrogated the Nrf2-dependent induction of heme oxygenase-1 expression by nitro-oleic acid. These results situate PtGR-1 as a critical modulator of both the steady state levels and signaling activities of fatty acid nitroalkenes in vivo.

We recently developed a sensitive assay for 3′,5′-cAMP using high-performance liquid chromatography-tandem mass spectrometry. Using this assay, we investigated the release of 3′,5′-cAMP from isolated, perfused rat kidneys. To our surprise, we observed a dominant chromatographic peak that was because of an endogenous substance that had the same parent ion as 3′,5′-cAMP and that fragmented to the same daughter ion (adenine) as 3′,5′-cAMP. However, the retention time of this unknown was approximately 2.9 min, compared with 6.3 min for authentic 3′,5′-cAMP. We hypothesized that the unknown substance was an isomer of 3′,5′-cAMP. The unknown substance had the same retention time and mass spectral properties as authentic 2′,3′-cAMP. Renal venous secretion of 2′,3′-cAMP was greater in kidneys from 20-week-old genetically hypertensive rats compared with age-matched normotensive rats (12.49 ± 2.14 versus 5.32 ± 1.97 ng/min/g kidney weight, respectively; n = 18). Isoproterenol (1 μM; β-adrenoceptor agonist) increased renal venous 3′,5′-cAMP secretion (approximately 690% of control) but had no effect on 2′,3′-cAMP production. In contrast, rapamycin (0.2 μM; activator of mRNA turnover) and iodoacetate + 2,4-dinitrophenol (50 μM; metabolic inhibitors) increased the renal venous secretion of 2′,3′-cAMP (approximately 1000 and 4100% of control, respectively) while simultaneously decreasing the renal venous secretion of 3′,5′-cAMP. In conclusion, 2′,3′-cAMP is a naturally occurring isomer of 3′,5′-cAMP that is: 1) not made by adenylyl cyclase; 2) released from kidneys into the extracellular compartment; 3) released more by kidneys from rats with long-standing hypertension; 4) derived from mRNA turnover; and 5) increased by energy depletion.