Mapping bone microstructure trace element composition: Sr,Ba, Cu,and Pb in an early modern Danish skeleton

Objectives

A Geographical Information System (GIS) approach enhances the acquisition, management, and analysis of trace element data from cortical bone. A high-resolution spatial dimension expands the research potential of Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS) data from cortical bone cross-sections. The chemical characterization of hundreds of osteons, notably sequences of superimposed osteons, permits more exacting studies of individual life histories than is possible with analyses of bulk bone samples.

Methods

A GIS procedure was used to estimate Sr, Ba, Pb, and Cu concentrations, originally generated through LA-ICP-MS, for bone microstructural features, notably fragmentary and intact osteons, in a human femoral cross-section. The skeleton is from Ribe, Denmark, and dates to the early modern period.

Results

Postmortem chemical alteration was limited to the bone's outer and inner margins. Two dietary indicators, Sr and Ba, and two socioeconomic indicators, Pb and Cu, measured for individual osteons were correlated with one another. Osteon sequences indicate concentrations of all four elements increased late in life for this individual.

Conclusions

The application of GIS procedures expedites fine-grained analyses of variation in the distribution of trace elements in bone microstructure identifiable in cortical bone cross-sections. It provides an efficient means of extracting the most information possible from LA-ICP-MS data about the lives of people in the past. Combining the two procedures makes it easier to track exposure to elements such as Pb across the part of an individual's life represented by osteon sequences.

     

Geoducks (Panopea abrupta) as isotopic bioarchives of seasonality in the climate of British Columbia

Geochemical analyses of geoduck shells (Panopea abrupta) have yielded new high-resolution isotope records of individual growth increments. The geoducks, which were collected from Parry Passage at Langara Island, British Columbia, are the most northern specimens that have yet been analyzed for stable oxygen and carbon isotopes. The isotope records represent the entire shell growth during the calendar years 1918, 1987, and 1991. The analyses showed that values of δ¹⁸O_shell and δ¹³C_shell vary within their growth increments. Temperature derived from δ¹⁸O_shell is consistent with modern instrumental data of sea-surface temperature. High-resolution δ¹⁸O records from individual growth increments substantiate the assumption that they are formed annually. The instrumental data indicate that the growth season was from May through October/November. The sea temperature became gradually warmer during the beginning of the growth season, and decreased steeply during autumn. Similar subannual trends can be recognized in shells growing almost a century ago. Correlation between δ¹⁸O_shell and δ¹³C_shell is negative for 1917–1919 and 1986–1988, whereas the correlation is positive for 1990–1992. Differences in correlation may be related to the composition of available food particles, i.e., algal blooms and plant debris. The food composition is partially affected by the Pacific Decadal Oscillation (PDO). These findings have major implications for the understanding of the interpretation of carbon isotopes in marine bivalves. Because of their spatial and temporal distribution, the geoduck shells are highly suitable for oceanographic reconstructions using stable-isotope analysis and sclerochronology. Furthermore, the large shell size and thickness and their deeply buried life position suggest a high preservation potential in ancient deposits. It is well known that the genus Panopea has a long-range fossil record.     

Comparative Analysis of Protocols to Induce Human CD4+Foxp3+ Regulatory T Cells by Combinations of IL-2, TGF-beta,Retinoic Acid,Rapamycin and Butyrate

Regulatory T cells (Tregs) suppress other immune cells and are critical mediators of peripheral tolerance. Therapeutic manipulation of Tregs is subject to numerous clinical investigations including trials for adoptive Treg transfer. Since the number of naturally occurring Tregs (nTregs) is minute, it is highly desirable to develop a complementary approach of inducing Tregs (iTregs) from naïve T cells. Mouse studies exemplify the importance of peripherally induced Tregs as well as the applicability of iTreg transfer in different disease models. Yet, procedures to generate iTregs are currently controversial, particularly for human cells. Here we therefore comprehensively compare different established and define novel protocols of human iTreg generation using TGF-β in combination with other compounds. We found that human iTregs expressed several Treg signature molecules, such as Foxp3, CTLA-4 and EOS, while exhibiting low expression of the cytokines Interferon-γ, IL-10 and IL-17. Importantly, we identified a novel combination of TGF-β, retinoic acid and rapamycin as a robust protocol to induce human iTregs with superior suppressive activity in vitro compared to currently established induction protocols. However, iTregs generated by these protocols did not stably retain Foxp3 expression and did not suppress in vivo in a humanized graft-versus-host-disease mouse model, highlighting the need for further research to attain stable, suppressive iTregs. These results advance our understanding of the conditions enabling human iTreg generation and may have important implications for the development of adoptive transfer strategies targeting autoimmune and inflammatory diseases.     

Voluntary Exercise Prevents Cisplatin-Induced Muscle Wasting during Chemotherapy in Mice

Loss of muscle mass related to anti-cancer therapy is a major concern in cancer patients, being associated with important clinical endpoints including survival, treatment toxicity and patient-related outcomes. We investigated effects of voluntary exercise during cisplatin treatment on body weight, food intake as well as muscle mass, strength and signalling. Mice were treated weekly with 4 mg/kg cisplatin or saline for 6 weeks, and randomized to voluntary wheel running or not. Cisplatin treatment induced loss of body weight (29.8%, P<0.001), lean body mass (20.6%, P = 0.001), as well as anorexia, impaired muscle strength (22.5% decrease, P<0.001) and decreased glucose tolerance. In addition, cisplatin impaired Akt-signalling, induced genes related to protein degradation and inflammation, and reduced muscle glycogen content. Voluntary wheel running during treatment attenuated body weight loss by 50% (P<0.001), maintained lean body mass (P<0.001) and muscle strength (P<0.001), reversed anorexia and impairments in Akt and protein degradation signalling. Cisplatin-induced muscular inflammation was not prevented by voluntary wheel running, nor was glucose tolerance improved. Exercise training may preserve muscle mass in cancer patients receiving cisplatin treatment, potentially improving physical capacity, quality of life and overall survival.     

Phosphoproteomic Analysis of FLCN Inactivation Highlights Differential Kinase Pathways and Regulatory TFEB Phosphoserines

In Birt–Hogg–Dubé (BHD) syndrome, germline loss-of-function mutations in the Folliculin (FLCN) gene lead to an increased risk of renal cancer. To address how FLCN inactivation affects cellular kinase signaling pathways, we analyzed comprehensive phosphoproteomic profiles of FLCN^POS and FLCN^NEG human renal tubular epithelial cells (RPTEC/TERT1). In total, 15,744 phosphorylated peptides were identified from 4329 phosphorylated proteins. INKA analysis revealed that FLCN loss alters the activity of numerous kinases, including tyrosine kinases EGFR, MET, and the Ephrin receptor subfamily (EPHA2 and EPHB1), as well their downstream targets MAPK1/3. Validation experiments in the BHD renal tumor cell line UOK257 confirmed that FLCN loss contributes to enhanced MAPK1/3 and downstream RPS6K1/3 signaling. The clinically available MAPK inhibitor Ulixertinib showed enhanced toxicity in FLCN^NEG cells. Interestingly, FLCN inactivation induced the phosphorylation of PIK3CD (Tyr524) without altering the phosphorylation of canonical Akt1/Akt2/mTOR/EIF4EBP1 phosphosites. Also, we identified that FLCN inactivation resulted in dephosphorylation of TFEB Ser109, Ser114, and Ser122, which may be linked to increased oxidative stress levels in FLCN^NEG cells. Together, our study highlights differential phosphorylation of specific kinases and substrates in FLCN^NEG renal cells. This provides insight into BHD-associated renal tumorigenesis and may point to several novel candidates for targeted therapies.     

An Ancient Duplication of Exon 5 in the Snap25 Gene Is Required for Complex Neuronal Development/Function

Alternative splicing is an evolutionary innovation to create functionally diverse proteins from a limited number of genes. SNAP-25 plays a central role in neuroexocytosis by bridging synaptic vesicles to the plasma membrane during regulated exocytosis. The SNAP-25 polypeptide is encoded by a single copy gene, but in higher vertebrates a duplication of exon 5 has resulted in two mutually exclusive splice variants, SNAP-25a and SNAP-25b. To address a potential physiological difference between the two SNAP-25 proteins, we generated gene targeted SNAP-25b deficient mouse mutants by replacing the SNAP-25b specific exon with a second SNAP-25a equivalent. Elimination of SNAP-25b expression resulted in developmental defects, spontaneous seizures, and impaired short-term synaptic plasticity. In adult mutants, morphological changes in hippocampus and drastically altered neuropeptide expression were accompanied by severe impairment of spatial learning. We conclude that the ancient exon duplication in the Snap25 gene provides additional SNAP-25-function required for complex neuronal processes in higher eukaryotes.     

Lung Protective Ventilation Induces Immunotolerance and Nitric Oxide Metabolites in Porcine Experimental Postoperative Sepsis

Low tidal volume ventilation is beneficial in patients with severe pulmonary dysfunction and would, in theory, reduce postoperative complications if implemented during routine surgery. The study aimed to investigate whether low tidal volume ventilation and high positive end-expiratory pressure (PEEP) in a large animal model of postoperative sepsis would attenuate the systemic inflammatory response and organ dysfunction. Thirty healthy pigs were randomized to three groups: Group Prot-7h, i.e. protective ventilation for 7 h, was ventilated with a tidal volume of 6 mL x kg^-1 for 7 h; group Prot-5h, i.e. protective ventilation for 5 h, was ventilated with a tidal volume of 10 mL x kg^-1 for 2 h, after which the group was ventilated with a tidal volume of 6 mL x kg^-1; and a control group that was ventilated with a tidal volume of 10 mL x kg^-1 for 7 h. In groups Prot-7h and Prot-5h PEEP was 5 cmH₂O for 2 h and 10 cmH₂O for 5 h. In the control group PEEP was 5 cmH₂O for the entire experiment. After surgery for 2 h, postoperative sepsis was simulated with an endotoxin infusion for 5 h. Low tidal volume ventilation combined with higher PEEP led to lower levels of interleukin 6 and 10 in plasma, higher PaO₂/FiO2, better preserved functional residual capacity and lower plasma troponin I as compared with animals ventilated with a medium high tidal volume and lower PEEP. The beneficial effects of protective ventilation were seen despite greater reductions in cardiac index and oxygen delivery index. In the immediate postoperative phase low V_T ventilation with higher PEEP was associated with reduced ex vivo plasma capacity to produce TNF-α upon endotoxin stimulation and higher nitrite levels in urine. These findings might represent mechanistic explanations for the attenuation of systemic inflammation and inflammatory-induced organ dysfunction.