Effect of diets with different crude protein levels on ammonia and greenhouse gas emissions from a naturally ventilated dairy housing.

Less crude protein (CP) in the diet can reduce nitrogen excretion of dairy cattle and lower their ammonia (NH3) and nitrous oxide (N2O) formation potential. The diet composition might also affect emissions of methane (CH4) and carbon dioxide (CO2). However, previous studies did not investigate the effect of diets with different CP levels that are customary practice in Switzerland on NH3 and greenhouse gas emissions on a practical scale. In a case-control approach, we quantified the emissions (NH3, N2O, CH4, CO2) in two separate but identical compartments of a naturally ventilated cubicle housing for lactating dairy cows over six days by using a tracer ratio method. Cows in one compartment received a diet with 116 g CP per kilogram dry matter (DM), in the other compartment with 166 g CP kg-1 DM. Subsequently, diets were switched for a second 6-day measurement phase. The results showed that the diet, aside from outside temperature and wind speed in the housing, was driving NH3 and N2O emissions. NH3 and N2O emission reduction per livestock unit (LU) was on average 46 % and almost 20 %, respectively, for the diet with low CP level compared to the higher CP level. In addition, strong relationships were observed between the CP content of the diet, N excretion in the urine and the milk urea content. An increased temperature or wind speed led to a clear increase in NH3 emissions. Differences in CH4 and CO2 emissions per LU indicated a significant influence of the diet, which cannot be attributed to the CP content. Our herd-level study demonstrated that a significant reduction in NH3 and N2O emissions related to LU, energy-corrected milk as well as DM intake can be achieved by lowering the CP content in the diet.

Monounsaturated and Diunsaturated Fatty Acids Sensitize Cervical Cancer to Radiation Therapy.

Obesity induces numerous physiological changes that can impact cancer risk and patient response to therapy. Obese patients with cervical cancer have been reported to have superior outcomes following chemoradiotherapy, suggesting that free fatty acids (FFA) might enhance response to radiotherapy. Here, using preclinical models, we show that monounsaturated and diunsaturated FFAs (uFFA) radiosensitize cervical cancer through a novel p53-dependent mechanism. UFFAs signaled through PPARγ and p53 to promote lipid uptake, storage, and metabolism after radiotherapy. Stable isotope labeling confirmed that cervical cancer cells increase both catabolic and anabolic oleate metabolism in response to radiotherapy, with associated increases in dependence on mitochondrial fatty acid oxidation for survival. In vivo, supplementation with exogenous oleate suppressed tumor growth in xenografts after radiotherapy, an effect that could be partially mimicked in tumors from high fat diet-induced obese mice. These results suggest that supplementation with uFFAs may improve tumor responses to radiotherapy, particularly in p53 wild-type tumors. SIGNIFICANCE: Metabolism of monounsaturated and diunsaturated fatty acids improves the efficacy of radiotherapy in cancer through modulation of p53 activity. See related commentary by Jungles and Green, p. 4513.

Stromal Reprogramming by FAK Inhibition Overcomes Radiation Resistance to Allow for Immune Priming and Response to Checkpoint Blockade.

The effects of radiotherapy (RT) on tumor immunity in pancreatic ductal adenocarcinoma (PDAC) are not well understood. To better understand if RT can prime antigen-specific T-cell responses, we analyzed human PDAC tissues and mouse models. In both settings, there was little evidence of RT-induced T-cell priming. Using in vitro systems, we found that tumor-stromal components, including fibroblasts and collagen, cooperate to blunt RT efficacy and impair RT-induced interferon signaling. Focal adhesion kinase (FAK) inhibition rescued RT efficacy in vitro and in vivo, leading to tumor regression, T-cell priming, and enhanced long-term survival in PDAC mouse models. Based on these data, we initiated a clinical trial of defactinib in combination with stereotactic body RT in patients with PDAC (NCT04331041). Analysis of PDAC tissues from these patients showed stromal reprogramming mirroring our findings in genetically engineered mouse models. Finally, the addition of checkpoint immunotherapy to RT and FAK inhibition in animal models led to complete tumor regression and long-term survival. SIGNIFICANCE: Checkpoint immunotherapeutics have not been effective in PDAC, even when combined with RT. One possible explanation is that RT fails to prime T-cell responses in PDAC. Here, we show that FAK inhibition allows RT to prime tumor immunity and unlock responsiveness to checkpoint immunotherapy. This article is highlighted in the In This Issue feature, p. 2711.

Glutaminase Inhibitors Induce Thiol-Mediated Oxidative Stress and Radiosensitization in Treatment-Resistant Cervical Cancers.

The purpose of this study was to determine if radiation (RT)-resistant cervical cancers are dependent upon glutamine metabolism driven by activation of the PI3K pathway and test whether PI3K pathway mutation predicts radiosensitization by inhibition of glutamine metabolism. Cervical cancer cell lines with and without PI3K pathway mutations, including SiHa and SiHa PTEN-/- cells engineered by CRISPR/Cas9, were used for mechanistic studies performed in vitro in the presence and absence of glutamine starvation and the glutaminase inhibitor, telaglenastat (CB-839). These studies included cell survival, proliferation, quantification of oxidative stress parameters, metabolic tracing with stable isotope-labeled substrates, metabolic rescue, and combination studies with L-buthionine sulfoximine (BSO), auranofin (AUR), and RT. In vivo studies of telaglenastat ± RT were performed using CaSki and SiHa xenografts grown in immune-compromised mice. PI3K-activated cervical cancer cells were selectively sensitive to glutamine deprivation through a mechanism that included thiol-mediated oxidative stress. Telaglenastat treatment decreased total glutathione pools, increased the percent glutathione disulfide, and caused clonogenic cell killing that was reversed by treatment with the thiol antioxidant, N-acetylcysteine. Telaglenastat also sensitized cells to killing by glutathione depletion with BSO, thioredoxin reductase inhibition with AUR, and RT. Glutamine-dependent PI3K-activated cervical cancer xenografts were sensitive to telaglenastat monotherapy, and telaglenastat selectively radiosensitized cervical cancer cells in vitro and in vivo These novel preclinical data support the utility of telaglenastat for glutamine-dependent radioresistant cervical cancers and demonstrate that PI3K pathway mutations may be used as a predictive biomarker for telaglenastat sensitivity.

Occurrence and Fate of Natural Estrogens in Swiss Cattle and Pig Slurry.

Natural estrogens act as endocrine disruptors. However, the fate of livestock farming derived natural estrogens (17α-estradiol, 17ß-estradiol, estrone, and estriol) in slurry is not well understood. In this study, we assessed the effects of on farm-storage on natural estrogen concentrations in slurry. Furthermore, we monitored pig and cattle slurry pits from major agricultural areas in Switzerland and determined natural estrogen concentrations therein. They were relatively stable over time, and mean concentrations ranged from 138 to 861 and 54 to 244 ng/L for cattle and pig slurries, respectively. 17α-Estradiol and estriol were the most prevalent estrogens in cattle and pig slurries, respectively. Based on livestock numbers, agricultural area, and estrogen concentrations in slurry, the estimated annual load of total natural estrogens applied on agricultural area amounted to 36 mg/ha. Our results indicate that slurry application is a relevant source of natural estrogens in the environment.

Investigation of Production Limits in Manufacturing Microstructured Surfaces Using Micro Coining.

The application of microstructured surfaces is one possible method to reduce friction in lubricated contacts between components with relative movement. Due to this, the energy efficiency and the occurring wear during the operating time of the final products could be decreased. To manufacture structured surfaces economically, a micro coining process was analyzed within this study. This process offers the potential for integration into the established manufacturing processes of different final products, such as tappets used in a valve train. Thus, large-scale production is enabled. To detect the manufacturing limits of the micro coining process, the manufacturing of the coining tools as well as the coining process needs to be investigated. Within this study, the achievable accuracy and the failure of cuboid and cylindrical microstructure elements with selected dimensions were analyzed. For both types of microstructures, the minimal lateral dimensions were detected. Besides the achievable accuracy, correlations between different geometrical dimensions of the micro elements are presented. Additionally, the aspect ratio is detected as the main cause of failure for the micro coining process. In general, the suitability of a coining process for manufacturing microstructured surfaces is proven.

Recognizing sepsis in the adult patient.

Patients in every health care setting are at risk for systemic inflammatory response syndrome, sepsis, severe sepsis, and even septic shock. The increasing incidence of sepsis, especially among older adults, its high mortality rate, and its subtle and rapid progression make prompt recognition and treatment imperative. Even though severe sepsis requires treatment in the ICU, the assessment of sepsis isn't solely the domain of the physician, critical care nurse, or ED nurse. Improving outcomes in patients with sepsis depends on every nurse involved in their care. The case study presented here is of a nursing home resident with unrecognized sepsis that progresses to severe sepsis-at which point not even seven days' treatment in the ICU could halt the progression to multiple organ failure.

The in vivo antitumoral effects of lipopolysaccharide against glioblastoma multiforme are mediated in part by Toll-like receptor 4.

OBJECTIVE: Toll-like receptor 4 (Tlr-4) mediates many biological effects of lipopolysaccharide (LPS), which has antitumoral effects on glioblastoma both in vivo and in vitro. However, the precise role of Tlr-4 in these antitumoral effects remains unknown. METHODS: The role of Tlr-4 in the antitumoral effect of LPS on glioblastomas was assessed in wild-type BALB/c mice and in Tlr-4 knockout (KO) BALB/c mice. Mice were implanted with DBT glioblastoma cells intracranially or subcutaneously, were treated with intratumoral LPS, and were assessed by histopathological examination for degrees of tumor progression and inflammation. Flow cytometry and Western blotting with antibodies to the Tlr-4 receptor and flow cytometry to the related CD14 moiety were performed to quantitate the expression levels of these two receptors by glioblastoma cells. RESULTS: For subcutaneous tumors, LPS caused near complete tumor elimination in wild-type mice, but only a 50% reduction in Tlr-4 KO mice. For mice implanted with intracranial glioblastomas, LPS increased survival times modestly in wild-type mice, but showed no benefit in the Tlr-4 KO mice. There were no histological differences among wild-type and Tlr-4 KO mice, except for tumor size. In both models, an early neutrophilic and later macrophage-rich inflammatory infiltrate were seen after LPS administration. Quantitative flow cytometry and Western blotting showed no Tlr-4 receptor or CD14 expression in murine and human glioblastoma cells in vitro, and Western blotting suggested that Tlr-4 effects are mediated by nontumoral elements such as microglia and inflammatory cells. CONCLUSION: LPS-induced antitumoral effects on glioblastoma multiforme are mediated, in part, by the Tlr-4 receptor. Further understanding of this process may lead to novel treatment strategies for this uniformly fatal disease.

Analysis of the antitumoral mechanisms of lipopolysaccharide against glioblastoma multiforme.

Our objective was to analyze the lipopolysaccharide (LPS) antitumoral effect upon glioblastoma, including whether the lipid A subunit alone can elicit glioblastoma regression, whether dexamethasone suppresses this response to LPS, whether B and T lymphocytes factor in this response, and whether this antitumoral effect of LPS provides resistance against subsequent challenge with glioblastoma. Mice (BALB/c, nude or SCID) implanted with s.c. DBT glioblastomas were treated with LPS (with or without dexamethasone) or with lipid A. A subset of BALB/c mice in which s.c. DBT glioblastomas had previously been eradicated using LPS were re-implanted with s.c. or intracranial (i.c.) DBT cells. For mice with s.c. tumors, mean tumor masses (MTM) were compared between groups. Survival was compared for mice with i.c. tumors. Lipid A caused near complete tumor regression of DBT glioblastomas in BALB/c mice (p<0.0001). Dexamethasone did not alter the antitumoral effect of LPS (p=0.48). LPS reduced the MTM of s.c. glioblastomas in T lymphocyte-deficient nude mice, but not as effectively as in immunocompetent mice. The antitumoral response to LPS for T and B lymphocyte-deficient SCID mice bearing DBT glioblastomas was similar to that for nude mice. Eradication of s.c. DBT glioblastoma in BALB/c provided partial resistance to subsequent challenge with s.c. or i.c. glioblastoma. We conclude that the LPS-mediated antitumoral response against glioblastoma is dependent upon the lipid A subunit of LPS, partially dependent upon T lymphocytes, independent of B lymphocytes, unaffected by dexamethasone and provides partial protection against subsequent challenges with glioblastoma.

Albumin-binding MR blood pool agents as MRI contrast agents in an intracranial mouse glioma model.

Intravenous MRI contrast agents are commonly used to improve the detection of intracranial tumors and other central nervous system (CNS) lesions for diagnosis and treatment planning. Two small-molecule, albumin-binding blood pool contrast agents (MP-2269 and MS-325) of potential clinical significance were evaluated at 1.5 Tesla in a mouse glioma model and compared with an extracellular contrast agent (OptiMARK). Tumor image contrast was significantly enhanced and long-lived following administration of 30 micromole/kg of the blood pool agents: specifically, contrast enhancement peaked slowly at 25-30 min following administration, remained constant for >3 hr, and returned to baseline within 20 hr. Comparable but "transient" enhancement was achieved using 100 micromole/kg OptiMARK: specifically, contrast enhancement peaked rapidly at 2-5 min following administration and then declined over 40 min. The blood pool contrast agents demonstrated an approximately threefold increased dose-effectiveness and a lengthened window of tumor contrast enhancement in comparison to commonly available extracellular contrast agents. This demonstrates the potential of alternative contrast-enhanced (CE) MRI examination protocols for tumor detection.