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At least 73 records · Page 4

Osteoporosis Treatments Affect Bone Matrix Maturation in a Rat Model of Induced Cortical Remodeling

ABSTRACT To test how osteoporosis drugs affect bone matrix maturation during cortical bone remodeling, 72 pregnant rats were switched from a 0.4% to a 0.01% calcium diet at parturition for a 23‐day lactation period. At weaning, eight dams were sacrificed to establish baseline values, while the remaining dams were returned to 0.4% calcium and treated with vehicle (saline), sodium fluoride (NaF), zoledronic acid (ZA), or sclerostin antibody (Scl‐Ab) for either 7 or 28 days (eight animals per group per time point). Femora were examined by μCT, dynamic histomorphometry, Fourier transform infrared imaging, and three‐point bending of notched specimens. Cortical porosity decreased in all groups from baseline to day 28. Intracortical mineralizing surface (MS/BS) and mineral apposition rate (MAR), as well as the mineral‐to‐matrix ratio were unaffected by treatment, but intracortical crystallinity was increased in the ZA group at day 10 compared with vehicle. Cortical area increased in all groups over 28 days mainly because of an addition of bone at the endocortical surface. Endocortical MS/BS did not vary among the groups, but endocortical MAR was suppressed in the NaF group at day 2 and elevated in the Scl‐Ab group at day 4 compared with vehicle. Endocortical mineral‐to‐matrix ratio was increased at days 5 and 10 following NaF treatment and endocortical crystallinity was increased at day 5 following ZA treatment compared with vehicle. Fracture toughness did not differ among the groups. Thus, the treatments affected matrix maturation more strongly at the endocortical then intracortical envelope. In this model of induced remodeling, the bone formation phase is synchronized at multiple sites, facilitating study of the effects of drugs or other bone‐targeting agents on matrix maturation independent of their effects on the initiation of remodeling. © 2020 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

59 BASIC BIOLOGICAL SCIENCES↗

Adaptation of metabolism to multicellular aggregation, hypoxia and obese stromal cell incorporation as potential measure of survival of ovarian metastases

Ovarian metastases exfoliate from the primary tumor and it is thought that aggregation supports their survival in the peritoneal cavity during dissemination but the underlying mechanisms are not clearly identified. We have previously shown that ovarian cancer cells acquire an increasingly glycolytic and metabolic flexible phenotype during progression. In the present study, we investigated how hypoxia, aggregation, and the incorporation of the obese stromal vascular fraction (SVF) affect cellular metabolism and the response to common anti-cancer and anti-diabetic drugs. Our results show a reduction of glucose uptake, lactate secretion, cellular respiration and ATP synthesis in response to hypoxia and aggregation, suggesting that the observed reduced proliferation of cells aggregated into spheroids is the result of a down-regulation of respiration. Recruitment of SVF to spheroids increased the spheroids invasive capacity but reduced respiration only in the most aggressive cells. Further, aggregation and hypoxia reduced the response to the metabolic drugs AICAR and metformin, and the chemotherapeutic agents cisplatin and paclitaxel. Our results suggest that the adaptation of cellular metabolism may contribute to enhanced survival under non-permissive conditions, and that these metabolic alterations may provide targets for future interventions that aim to enhance the survival of women with metastatic ovarian cancer.

60 APPLIED LIFE SCIENCES↗

SMYD2 suppresses p53 activity to promote glucose metabolism in cervical cancer

Reprogrammed energy metabolism, especially the Warburg effect, is emerged as a hallmark of cancer. The protein lysine methyltransferase SMYD2 functions as an oncogene and is implicated in various malignant phenotypes of human cancers. However, the role of SMYD2 in tumor metabolism is still largely unknown. Here, we report that SMYD2 is highly expressed in human cervical cancer and its aberrant expression is linked to a poor prognosis. Bioinformatic analysis revealed a novel link between SMYD2 expression and aerobic glycolysis. Through loss-of-function experiments, we demonstrated that SMYD2 knockdown or inhibition induced a metabolic shift from aerobic glycolysis to oxidative phosphorylation, as evidenced by glucose uptake, lactate production, extracellular acidification, and the oxygen consumption rate. In contrast, SMYD2 overexpression promoted glycolytic metabolism in cervical cancer cells. Moreover, SMYD2 was required for tumor growth in cervical cancer and this oncogenic activity was largely glycolysis-dependent. Mechanistically, SMYD2 altered the methylation status of p53 and inhibited its transcriptional activity. Genetic silencing of p53 largely abrogated the effects of SMYD2 in promoting aerobic glycolysis. Taken together, our findings reveal a novel function of SMYD2 in regulating the Warburg effect in cervical cancer.

60 APPLIED LIFE SCIENCES↗

Organic Microbial Electrochemical Transistor Monitoring Extracellular Electron Transfer

Abstract Extracellular electron transfer (EET) denotes the process of microbial respiration with electron transfer to extracellular acceptors and has been exploited in a range of microbial electrochemical systems (MESs). To further understand EET and to optimize the performance of MESs, a better understanding of the dynamics at the microscale is needed. However, the real‐time monitoring of EET at high spatiotemporal resolution would require sophisticated signal amplification. To amplify local EET signals, a miniaturized bioelectronic device, the so‐called organic microbial electrochemical transistor (OMECT), is developed, which includes Shewanella oneidensis MR‐1 integrated onto organic electrochemical transistors comprising poly(3,4‐ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) combined with poly(vinyl alcohol) (PVA). Bacteria are attached to the gate of the transistor by a chronoamperometric method and the successful attachment is confirmed by fluorescence microscopy. Monitoring EET with the OMECT configuration is achieved due to the inherent amplification of the transistor, revealing fast time‐responses to lactate. The limits of detection when using microfabricated gates as charge collectors are also investigated. The work is a first step toward understanding and monitoring EET in highly confined spaces via microfabricated organic electronic devices, and it can be of importance to study exoelectrogens in microenvironments, such as those of the human microbiome.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Domesticating a food spoilage yeast into an organic acid–tolerant metabolic engineering host: Lactic acid production by engineered Zygosaccharomyces bailii

Lactic acid represents an important class of commodity chemicals, which can be produced by microbial cell factories. However, due to the toxicity of lactic acid at lower pH, microbial production requires the usage of neutralizing agents to maintain neutral pH. Zygosaccharomyces bailii, a food spoilage yeast, can grow under the presence of organic acids used as food preservatives. This unique trait of the yeast might be useful for producing lactic acid. With the goal of domesticating the organic acid!tolerant yeast as a metabolic engineering host, seven Z. bailii strains were screened in a minimal medium with 10 g/L of acetic, or 60 g/L of lactic acid at pH 3. The Z. bailii NRRL Y7239 strain was selected as the most robust strain to be en- gineered for lactic acid production. By applying a PAN!ARS!based CRISPR!Cas9 system consisting of a transfer RNA promoter and NAT selection, we demonstrated the targeted deletion of ADE2 and site!specific integration of Rhizopus oryzae ldhA coding for lactate dehydrogenase into the PDC1 locus. The resulting pdc1::ldhA strain produced 35 g/L of lactic acid without ethanol production. Here, this study de- monstrates the feasibility of the CRISPR!Cas9 system in Z. bailii, which can be applied for a fundamental study of the species.

59 BASIC BIOLOGICAL SCIENCES↗

Intrinsically Chiral Twist‐Bend Nematogens: Interplay of Molecular and Structural Chirality in the N TB Phase

Abstract Non‐symmetric lactate‐based chiral liquid crystal dimers containing an odd‐membered spacer are shown to exhibit a chiral twist‐bend nematic phase which is stable on cooling to room temperature. A comparison of racemic and optically pure materials reveals that the pitch length in the N* TB phase is not influenced by molecular chirality, whereas the nematic‐twist‐bend nematic transition temperature is increased.

Chemistry↗

TALSPEAK-based separation of the trivalent actinides from rare earth elements using LN resin

The separation of 241 Am, 244 Cm and 249 Cf from rare earth elements using LN resin and lactate buffer solutions with diethylenetriaminepentaacetic acid is described with numerous columns studies. The elutions are based on TALSPEAK liquid–liquid extraction chemistry, and provide high yield (> 90%), rapid, simple separations of the trivalent actinides from the lanthanides, which is often challenging due to similarities in chemical behavior among the trivalent f-block elements. Further, all three actinides can be separated from rare earth elements, including from massless fission product samples and samples with small amounts (~ 1–5 mg) of stable lanthanides. Separations with no detectable overlap between the actinide and lanthanide elutions is possible with massless samples, and in samples with mass high separation factors can be achieved (Am/Eu: 171; Am/La: 10 6 ).

38 RADIATION CHEMISTRY, RADIOCHEMISTRY, AND NUCLEA↗

Cation exchange separation of radium and actinium using lactic acid DTPA buffer

The separation of actinium ( 228 Ac) from radium ( 223,228 Ra) on cation exchange resin with a diethylenetriaminpentaacetic acid-lactate buffer solution is demonstrated with series of columns. High yield, high radiopurity (~ 100%) separations of Ac from Ra are feasible with small columns in biologically compatible conditions and pHs. As Ac is eluted before Ra on these columns, further studies were performed to determine if this separation system could be applied to Ra/Ac isotope generators, but these were not successful. In conclusion, the separations presented in this work may be relevant for radiopharmaceutical purifications of 225 Ac which is typically obtained from its 225 Ra parent isotope.

and nuclear chemistry↗

Rewiring metabolism of Clostridium thermocellum for consolidated bioprocessing of lignocellulosic biomass poplar to produce short-chain esters

Consolidated bioprocessing (CBP) of lignocellulosic biomass uses cellulolytic microorganisms to enable enzyme production, saccharification, and fermentation to produce biofuels, biochemicals, and biomaterials in a single step. However, understanding and redirecting metabolisms of these microorganisms compatible with CBP are limited. Here, a cellulolytic thermophile Clostridium thermocellum was engineered and demonstrated to be compatible with CBP integrated with a Co-solvent Enhanced Lignocellulosic Fractionation (CELF) pretreatment for conversion of hardwood poplar into short-chain esters with industrial use as solvents, flavors, fragrances, and biofuels. The recombinant C. thermocellum engineered with deletion of carbohydrate esterases and stable overexpression of alcohol acetyltransferases improved ester production without compromised deacetylation activities. These esterases were discovered to exhibit promiscuous thioesterase activities and their deletion enhanced ester production by rerouting the electron and carbon metabolism. Ester production was further improved up to 80-fold and ester composition could be modulated by deleting lactate biosynthesis and using poplar with different pretreatment severity.

59 BASIC BIOLOGICAL SCIENCES↗

Coupling gas purging with inorganic carbon supply to enhance biohydrogen production with Clostridium thermocellum

Clostridium thermocellum is a desirable biocatalyst for biohydrogen production, with a native ability to simultaneously saccharify cellulose and to metabolize released cellodextrins for hydrogen production. During fermentation with C. thermocellum , partial pressures of two gases - CO 2 and H 2 - are critical drivers of overall reaction kinetics. Biohydrogen production is enhanced by maintaining a low hydrogen partial pressure, while high concentrations of dissolved CO 2 promote microbial biomass synthesis. Our study evaluates the inherent trade-offs between hydrogen stripping and inorganic carbon supply for optimized biohydrogen synthesis. Here, we find that nitrogen sparging at low flow rates increases hydrogen production when compared with an equivalent nitrogen overlay, but that high rates of nitrogen sparging inhibit cell growth and hydrogen production. Decreasing dissolved hydrogen partial pressure via nitrogen sparging also lowers the production of reduced metabolites, including lactate and ethanol. To address potential stripping of inorganic carbon from the production medium, we supplemented CO 2 to the sparging gas and co-optimized for gas flow rate and for the CO 2 fraction of the sparging gas. Total hydrogen production increased from 50 mmol∙L -1 in the base condition, when the bioreactor was sparged with 0.1 LPM of pure nitrogen, to 181.3 mmol∙L -1 when sparged with 1.3 LPM of 33 % CO 2 , demonstrating that biohydrogen production is highly sensitive to both parameters. Fine sensitivity of biohydrogen production to sparging conditions highlights the critical importance of bioreactor design and operation to achieve maximum H 2 removal without compromising inorganic carbon supply to bacterial central metabolism.

09 BIOMASS FUELS↗

Impact of organic acids and sulfate on the biogeochemical properties of soil from urban subsurface environments

Urban subsurface environments are often different from undisturbed subsurface environments due to the impacts of human activities. For example, deterioration of underground infrastructure can introduce elevated levels of Ca, Fe, and heavy metals into subsurface soils and groundwater. Likewise, leakage from sewer systems can lead to contamination by organic C, N, S, and P. However, the impact of these organic and inorganic compounds on biogeochemical processes including microbial redox reactions, mineral transformations, and microbial community transitions in urban subsurface environments is poorly understood. Here we conducted a microcosm experiment with soil samples from an urban construction site to investigate the possible biotic and abiotic processes impacted when sulfate and acetate or lactate were introduced into an urban subsurface environment. In the top-layer soil (0-0.3 m) microcosms, which were highly alkaline (pH > 10), the major impact was on abiotic processes such as secondary mineral precipitation. In the mid-layer (2-3 m) soil microcosms, the rate of Fe(III)-reduction and the amount of Fe(II) produced were greatly impacted by the specific organic acid added, and sulfate-reduction was not observed until after Fe(III)-reduction was complete. Near the end of the incubation, some genera related to syntrophic acetate oxidation and methanogenesis were observed in the lactateamended microcosms. In the bottom-layer (7-8 m) soil microcosms, the rate of Fe(III)-reduction and the amount of Fe(II) produced were affected by the concentration of amended sulfate. Sulfate-reduction was concurrent with Fe(III)-reduction, suggesting that Fe(II) production was likely due to abiotic reduction of Fe(III) by sulfide produced by microbial sulfate reduction. The slightly acidic initial pH (~5.8) of the mid-soil system was a major factor controlling sequential microbial Fe(III) and sulfate reduction versus parallel Fe(III) and sulfate reduction in the bottom soil system, which had a neutral initial pH (~7.2). Finally, 16S rRNA gene-based community analysis revealed a variety of indigenous microbial groups including alkaliphiles, dissimilatory iron and sulfate reducers, syntrophes, and methanogens tightly coupled with, and impacted by, these complex abiotic and biogeochemical processes occurring in urban subsurface environments.

54 ENVIRONMENTAL SCIENCES↗

Facile hyperpolarization chemistry for molecular imaging and metabolic tracking of [1– 13 C]pyruvate in vivo

Hyperpolarization chemistry based on reversible exchange of parahydrogen, also known as Signal Amplification By Reversible Exchange (SABRE), is a particularly simple approach to attain high levels of nuclear spin hyperpolarization, which can enhance NMR and MRI signals by many orders of magnitude. SABRE has received significant attention in the scientific community since its inception because of its relative experimental simplicity and its broad applicability to a wide range of molecules, however, in vivo detection of molecular probes hyperpolarized by SABRE has remained elusive. Here we describe a first demonstration of SABRE-hyperpolarized contrast detected in vivo, specifically using hyperpolarized [1– 13 C]pyruvate. Biocompatible formulations of hyperpolarized [1– 13 C]pyruvate in, both, methanol-water, and ethanol-water mixtures followed by dilution with saline and catalyst filtration were prepared and injected into healthy Sprague Dawley and Wistar rats. Effective hyperpolarization-catalyst removal was performed with silica filters without major losses in hyperpolarization. Metabolic conversion of pyruvate to lactate, alanine, and bicarbonate was detected in vivo. Pyruvate-hydrate was also observed as a minor byproduct. Measurements were performed on the liver and kidney at 4.7 T via time-resolved spectroscopy and chemical-shift-resolved MRI. In addition, whole-body metabolic measurements were obtained using a cryogen-free 1.5 T MRI system, illustrating the utility of combining lower-cost MRI systems with simple, low-cost hyperpolarization chemistry to develop safe and scalable molecular imaging.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Impact of Fermentation-Derived Substrates on Hydrogen Production in Zero-Gap Microbial Electrolysis Cells

Zero-gap microbial electrolysis cells (MECs) represent a promising platform for hydrogen production from liquid waste streams due to reduced interelectrode spacing that lowers internal resistance and enhances mass transport. However, the performance and stability of zero-gap MECs treating chemically complex feedstocks remain insufficiently characterized. Here, we operated zero-gap MECs with real, unamended, corn stover dark fermentation effluent containing a wide range of organic substrates. The MECs fed fermentation effluent achieved a maximum current density of 24 A/m2 (15+-6 A/m2 over the cycle) and a hydrogen production rate of 75 L/L-d (42+-19 L/L-d over the cycle). The substrates were consumed at different rates, indicating substrate-selective utilization by the anodic microbial community. Acetate supported high and stable current generation, whereas ethanol, formate, lactate, and amino acids induced varying degrees of inhibition depending on their concentration. Residual sugars caused pronounced current fluctuations, consistent with ongoing fermentation and local pH changes. A diverse microbial community was crucial for efficiently utilizing complex organics and maximizing electrochemical performance. These results demonstrate how and to what extent substrate composition regulates zero-gap MEC performance and that microbial community and operational conditions can be leveraged to enhance performance. These novel findings provide practical guidance for achieving robust hydrogen recovery from chemically heterogeneous real liquid waste streams.

08 HYDROGEN↗

Cooling and dietary crude protein affected milk production on heat-stressed dairy cows

Environmental factors, such as temperature and humidity, exert dramatic effects on the production, health, and well-being of lactating dairy cows. Heat-stressed cows undergo a series of homeorhetic adaptations that alters cellular homeostasis and nutrient utilization. As such, the objective of this study was to determine the effects of cooling and dietary crude protein (CP) content on fermentability of rumen inoculum, milk production, and health of heat-stressed cows. It was hypothesized that cooled but not heat-stressed cows would support productivity in response to an increase in dietary CP content. Thirty-six multiparous Holstein cows were randomly assigned to 1 of 4 treatments in a 2 × 2 factorial arrangement (9 cows/treatment). Treatments consisted of cooling (CO) or moderate long-term heat stress (HS) and diets containing low CP content (LCP; 12.5% CP) or moderate CP content (MCP; 16.1% CP) of dry matter. The CO treatment provided heat abatement with sprinklers and fans that came on at > 20.0°C, whereas the HS treatment did not provide sprinklers and fans. Cows were housed in pens in a freestall barn, and treatments were imposed for 21 d during July and August. Compared with the HS treatment, CO decreased afternoon rectal and vaginal temperatures (0.60 and 0.70°C) and afternoon respiration rate (27.6 breaths/min). Treatment interactions showed that compared with the LCP treatment, MCP increased milk fat yield (23%) in CO but not in HS cows. Compared with the LCP treatment, MCP decreased milk protein percent (0.14 units) in CO but increased milk protein percent (0.09 units) in HS cows. Rumen inoculum collected on d 21 showed a ~3-fold increase in total gas production in vitro in CO compared with the HS treatment. Relative to the LCP treatment, MCP increased plasma glucose and insulin and decreased total free fatty acids concentrations. In summary, under the conditions of this study CO but not HS cows responded to a CP stimulus by increasing productivity.

59 BASIC BIOLOGICAL SCIENCES↗

Associations between SARS-CoV-2 Infection or COVID-19 Vaccination and Human Milk Composition: A Multi-Omics Approach

Background: The risk of contracting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) via human milk-feeding is virtually nonexistent. Adverse effects of coronavirus disease 2019 (COVID-19) vaccination for lactating individuals are not different from the general population, and no evidence has been found that their infants exhibit adverse effects. Yet, there remains substantial hesitation among this population globally regarding the safety of these vaccines. Objectives: Herein, we aimed to determine if compositional changes in milk occur following SARS-CoV-2 infection or COVID-19 vaccination, including any evidence of vaccine components. Methods: An extensive multiomics approach was taken using a subset of milk samples obtained as part of our broad studies examining the effects on milk of SARS-CoV-2 infection and COVID-19 vaccination. Results: We found that compared with unvaccinated individuals, SARS-CoV-2 infection was associated with significant compositional differences in 67 proteins, 385 lipids, and 13 metabolites. In contrast, COVID-19 vaccination was not associated with any changes in lipids or metabolites, although it was associated with changes in 13 or fewer proteins. Compositional changes in milk differed by vaccine. Changes following vaccination were greatest after 1–6 h for the mRNA-based Moderna vaccine (8 changed proteins), 3 d for the mRNA-based Pfizer (4 changed proteins), and adenovirus-based Johnson and Johnson (13 changed proteins) vaccines. Proteins that changed after both natural infection and Johnson and Johnson vaccine were associated mainly with systemic inflammatory responses. In addition, no vaccine components were detected in any milk sample. Conclusions: Together, our data provide evidence of only minimal changes in milk composition because of COVID-19 vaccination, with much greater changes after natural SARS-CoV-2 infection.

60 APPLIED LIFE SCIENCES↗

Sex-specific respiratory and systemic endocrine effects of acute acrolein and trichloroethylene inhalation

Acrolein and trichloroethylene (TCE) are priority hazardous air pollutants due to environmental prevalence and adverse health effects; however, neuroendocrine stress-related systemic effects are not characterized. Comparing acrolein, an airway irritant, and TCE with low irritancy, we hypothesized that airway injury would be linked to neuroendocrine-mediated systemic alterations. Male and female Wistar-Kyoto rats were exposed nose-only to air, acrolein or TCE in incremental concentrations over 30 min, followed by 3.5-hr exposure to the highest concentration (acrolein - 0.0, 0.1, 0.316, 1, 3.16 ppm; TCE - 0.0, 3.16, 10, 31.6, 100 ppm). Real-time head-out plethysmography revealed acrolein decreased minute volume and increased inspiratory-time (males>females), while TCE reduced tidal-volume. Acrolein, but not TCE, inhalation increased nasal-lavage-fluid protein, lactate-dehydrogenase activity, and inflammatory cell influx (males>females). Neither acrolein nor TCE increased bronchoalveolar-lavage-fluid injury markers, although macrophages and neutrophils increased in acrolein-exposed males and females. Systemic neuroendocrine stress response assessment indicated acrolein, but not TCE, increased circulating adrenocorticotrophic hormone, and consequently corticosterone, and caused lymphopenia, but only in males. Acrolein also reduced circulating thyroid-stimulating hormone, prolactin, and testosterone in males. Furthermore, acute acrolein inhalation resulted in sex-specific upper respiratory irritation/inflammation and systemic neuroendocrine alterations linked to hypothalamic-pituitary-adrenal axes activation, which is critical in mediating extra-respiratory effects.

60 APPLIED LIFE SCIENCES↗

Investigating overflow metabolism in heterotrophic cultures of the green alga Chromochloris zofingiensis

Chromochloris zofingiensis is of interest for its ability to perform a reversible trophic switch in the presence of glucose that is characterized by a shutdown of photosynthesis and an accumulation of energy storage metabolites. Previous work has shown that this trophic switch is accompanied by overflow metabolism and the production of lactate in aerobic conditions. This trophic switch is not observed in nutrient replete media. We utilized isotopically assisted metabolic flux analysis to characterize intracellular flux distributions that are associated with different metabolic phenotypes observed in this organism in different media formulations in light and dark conditions. The results of this analysis showed that low iron cultures have no flux through carbon fixation reactions, and that the carbon flux entering the TCA cycle in these cultures is approximately 40 % lower than that in iron replete cultures grown heterotrophically. This analysis was complemented with transcriptomics data collected for C. zofingiensis grown in iron limited conditions to provide further evidence towards the negative impact of iron limitation on both photosynthetic and respiratory activity. Overflow metabolism allows this alga to compensate for the lower energy production that results from iron limitation. This work highlights how nutrient availability can lead to changes in the metabolism of C. zofingiensis.

59 BASIC BIOLOGICAL SCIENCES↗