Maximum Respiration Rate Research Articles

Respiration adaptation of activated sludge under dissolved oxygen and hypochlorite stressed conditions

In this work, the relationship between environmental disturbance and the recovery of activated sludge at a low dissolved oxygen of 0.5mg/l or in the presence of sodium hypochlorite of 1–15mg Cl2/gSS/d was examined. When microorganisms entered their physiological adaptation period, a sharp increase in endogenous respiration rate was observed. The activity recovery potential of sludge depended on the ratio of the endogenous respiration rate to the maximum respiration rate. A subsequent decrease in this ratio after a sharp increase indicates that the disturbance was recoverable. An increase in this ratio to a certain value, e.g., 0.35, suggests that the sludge system could not adapt to the new environments and thus was unrecoverable. In addition, the recoveries of sludge respiration and effluent quality were asynchronous, which was impacted by both sludge activity and operating conditions. These results provide a useful approach for the operation of activated sludge systems.

Abstract 3548: Hypoxia-induced Cystathionine-β-synthase (CBS) expression: impact on colonic epithelial cells metabolism, proliferation and migration

Abstract Introduction: We have reported aberrant overexpression of CBS in human colorectal cancers, where its activity (H2S production) promotes tumor progression and metastasis. New data show that CBS expression increases in adenomatous polyps. The aims of this study were to identify possible mechanisms for CBS upregulation and determine the biological impact of its upregulation using a premalignant colonic epithelial cell line NCM356. Methods: Induction of CBS in response to LPS, TLR2&4 agonists, and hypoxia was assessed by immunoblotting. CBS overexpression was achieved using lentiviral expression vector (pReceiver-Lv103, GeneCopoeia™) in NCM356 cells. Global metabolic profiling (Metabolon, Durham) was performed on cellular extracts from NCM365 overexpressing cells (CBS-hi) and vector-transduced controls. High-resolution respirometry (Orobors Oxygraph-2k) assessed bioenergetics. H2S levels were measured using the fluorogenic probe 7-Azido-4-methylcoumarin. A Coulter counter was used to assess cell proliferation rates. Transwell assays were used to assess cell migration. CBS activity was inhibited with aminooxyacetic acid (AOAA). Results: CBS protein expression was increased by ~3-fold in pNCM after 2 and 24 hr of hypoxia (1% O2). Treatment with LPS but not the TLR agonists increased CBS expression 2-fold. Metabolic profiling of CBS-hi cells and controls identified 85 metabolites that were differentially expressed (65 increased and 20 decreased; p≤0.05 Welch’s 2-Sample t-Test). The metabolite lanthionine, which is produced by the CBS-dependent condensation of cysteine to produce H2S, showed the largest increase (12.3-fold increase, p=1.5E-06). A 4-fold increase in basal H2S production was measured in CBS-hi cells compared to control cultures. High-resolution respirometry showed that CBS-hi cells exhibited a significant increase in maximum respiration rate, reserve respiratory capacity and increased citrate synthase activity, suggesting increased cellular mitochondria mass. Treatment of CBS-hi cells either with pentose phosphate pathway inhibitor (oxythiamine) or lactate dehydrogenase inhibitor (XF-11) reducing growth by 90% and ~25% at day 5, respectively; the inhibitors had no effect on the vector control cells. Exposure of NCM356 cells to hypoxic conditions resulted in increased migration to conditioned media (p<0.001, NCM356 normoxia vs. hypoxia); inhibition of CBS using AOAA attenuated the hypoxia-induced migration. Conclusions: Hypoxia and LPS can induce CBS expression in a premalignant colonic epithelial cell line. Upregulation of CBS has a board impact on cellular metabolism including enhanced flux through the transsulfuration, glycolytic and pentose phosphate pathways, resulting in enhanced cellular bioenergetics, growth and migration. These data suggest that enhance CBS activity may be involved the adenoma to carcinoma sequence. Citation Format: Ches'Nique M. Phillips, John R. Zatarain, Michael E. Nicholls, Craig Porter, Steven Widen, Ketan Thanki, James W. Randall, Judith L. Hellmich, Manjit Maskey, Suimin Qiu, Thomas G. Wood, Nadiya Druzhyna, Bartosz Szczesny, Katalin Modis, Csaba Szabo, Celia Chao, Mark R. Hellmich. Hypoxia-induced Cystathionine-β-synthase (CBS) expression: impact on colonic epithelial cells metabolism, proliferation and migration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3548. doi:10.1158/1538-7445.AM2017-3548

Evolution of Cytochrome c Oxidase in Hypoxia Tolerant Sculpins (Cottidae, Actinopterygii).

Vertebrate hypoxia tolerance can emerge from modifications to the oxygen (O2) transport cascade, but whether there is adaptive variation to O2 binding at the terminus of this cascade, mitochondrial cytochrome c oxidase (COX), is not known. In order to address the hypothesis that hypoxia tolerance is associated with enhanced O2 binding by mitochondria we undertook a comparative analysis of COX O2 kinetics across species of intertidal sculpins (Cottidae, Actinopterygii) that vary in hypoxia tolerance. Our analysis revealed a significant relationship between hypoxia tolerance (critical O2 tension of O2 consumption rate; Pcrit), mitochondrial O2 binding affinity (O2 tension at which mitochondrial respiration was half maximal; P50), and COX O2-binding affinity (apparent Michaelis-Menten constant for O2 binding to COX; Km,app O2). The more hypoxia tolerant species had both a lower mitochondrial P50 and lower COX Km,app O2, facilitating the maintenance of mitochondrial function to a lower O2 tension than in hypoxia intolerant species. Additionally, hypoxia tolerant species had a lower overall COX Vmax but higher mitochondrial COX respiration rate when expressed relative to maximal electron transport system respiration rate. In silico analyses of the COX3 subunit postulated as the entry point for O2 into the COX protein catalytic core, points to variation in COX3 protein stability (estimated as free energy of unfolding) contributing to the variation in COX Km,app O2. We propose that interactions between COX3 and cardiolipin at four amino acid positions along the same alpha-helix forming the COX3 v-cleft represent likely determinants of interspecific differences in COX Km,app O2.

Estrogen regulates spatially distinct cardiac mitochondrial subpopulations

Increased susceptibility to permeability transition pore (mPTP) is a significant concern to decreased cardiac performance in postmenopausal females. The goal of this study was to assess the effects of estrogen deficiency on the two spatially distinct mitochondrial subpopulations from left ventricle: subsarcolemmal mitochondria (SSM) and intermyofibrillar mitochondria (IFM) based on: morphology, membrane potential, oxidative phosphorylation, mPTP and reactive oxygen species production. Female rats (8weeks old) that underwent bilateral ovariectomy were randomly assigned to receive daily treatment with placebo (OVX), estrogen replacement (OVX+E2) and Sham for 60days. The yield for IFM was found higher in the OVX group and lower in the SSM. SSM internal complexity and size were higher in the OVX group, although membrane potential was not different. The maximal rate of mitochondrial respiration, states 3 and 4, using glutamate+malate as substrate, were higher in IFM and SSM from the OVX group. The respiratory control ratio (RCR - state3/state 4), was not different in both SSM and IFM with glutamate+malate. The ADP:O ratio was found lower in IFM and SSM from OVX compared to Sham. When pyruvate was used, state 3 was found unchanged in both IFM and SSM, state 4 was greater in IFM from OVX rats compared to Sham and the ADP/O ratio was decreased. The RCR was unchanged in both subpopulations. The IFM from OVX rats presented a lower Ca2+retention capacity compared to Sham, however, the SSM remained unchanged. Hydrogen peroxide formation was found increased in the IFM from OVX animals with glutamate+malate and rotenone+succinate as substrates. The SSM showed increased ROS production only with rotenone+succinate. Western blot analyzes showed decreased levels of PGC-1α and NRF-1 in the OVX group. Estrogen replacement was able to restore most of the alterations induced by ovariectomy. In conclusion, our data shows that estrogen deficiency has distinct effects on the two spatially distinct mitochondrial subpopulations in oxidative phosphorylation, morphology, calcium retention capacity and ROS production.

Noninvasive positive pressure ventilation enhances the effects of aerobic training on cardiopulmonary function.

PurposeThe purpose of this study was to determine the effect of aerobic training under noninvasive positive pressure ventilation (NPPV) on maximal oxygen uptake ().MethodsTen healthy young male volunteers participated in the study. Before the training, stroke volume (SV) and cardiac output (CO) were measured in all subjects under 0, 4, 8, and 12 cmH2O NPPV at rest. Then, the subjects exercised on a cycle ergometer at 60% of pre-training for 30 min daily for 5 consecutive days with/without NPPV. The 5-day exercise protocol was repeated after a three-week washout period without/with NPPV. The primary endpoint was changes in . The secondary endpoints were changes in SV, CO, maximum heart rate (HRmax), maximum respiratory rate (RRmax), maximum expiratory minute volume (VEmax) and the percent change in plasma volume (PV).ResultsNPPV at 12 cmH2O significantly reduced SV and CO at rest. significantly increased after 5 days training with and without NPPV, but the magnitude of increase in after training under 12 cmH2O NPPV was significantly higher than after training without NPPV. VEmax significantly increased after training under NPPV, but not after training without NPPV. HRmax and RRmax did not change during training irrespective of NPPV. The percent change in PV was similar between training with and without NPPV. The 5-day training program with NPPV resulted in greater improvement in than without NPPV.ConclusionsAerobic training under NPPV has add-on effects on and exercise-related health benefits in healthy young men.

Risk factors and prediction model for inpatient surgical site infection after major abdominal surgery

BackgroundSurgical site infections (SSIs) are a common source of postoperative morbidity and a marker of surgical quality. The ability to predict the incidence of SSIs is limited and most models have poor predictive value. We sought to identify risk factors associated with SSIs and develop a prediction model for SSIs after major abdominal surgery. MethodsA total of 1744 patients undergoing pancreatic, hepatobiliary, and colorectal resections between January 1, 2010 and August 31, 2013 at Johns Hopkins Hospital were identified. Risk factors for any inpatient SSI (superficial and deep) were evaluated using multivariable logistic regression. ResultsMedian patient age was 58 y (interquartile range 47, 68); surgical procedures included colorectal (59.0%), liver (26.2%), and pancreas (14.8%) resections. SSI occurred in 7.6% (n = 132) of patients. Factors associated with SSI included preoperative weight loss >4.5 kg (odds ratio [OR], 2.12; 95% confidence interval [CI], 1.06-4.25), emergency operations (OR, 2.05; 95% CI, 1.32-3.17), and colorectal resections (OR, 1.65; 95% CI, 1.13-2.43) (all P ≤ 0.003). Intraoperative and postoperative risk factors included estimated blood loss (EBL) >600 mL (OR, 2.23; 95% CI, 1.54-3.25), maximum respiratory rate (tachypnea) >20 breaths/min (OR, 1.74; 95% CI, 1.19-2.54), and perioperative transfusion (OR, 2.01; 95% CI, 1.33-3.04) (all P = 0.001). Intraoperative hypothermia, hyperthermia, bradycardia, tachycardia, hypotension, and hypertension were not associated with SSIs (all P > 0.05). After controlling competing risk factors, transfusion, EBL >600 mL, tachypnea, and colorectal resection were independently associated with SSIs (all P < 0.003). On the basis of the beta-coefficients in the multivariable model, an SSI scoring system was created by assigning 2 points for EBL >600 mL, 2 points for a colorectal resection, 3 points for tachypnea, and 3 points for a transfusion. The model showed good discriminatory ability to predict SSI (c-statistic = 0.7232; Akaike information criterion 875.37). ConclusionsA novel, simple 10-point SSI scoring system that incorporated perioperative risk factors such as blood transfusion, EBL, tachypnea, and the type of surgical procedure accurately stratifies patients according to SSI risk.

Markers of Disease Severity in Patients with Spanish Influenza in the Japanese Armed Forces, 1919-1920.

We examined preserved medical charts of 470 Spanish influenza patients (8 with fatal cases) hospitalized at former army hospitals in Japan during 1919–1920. The following factors were associated with longer periods of hospitalization: adventitious discontinuous lung sounds, maximum respiration rate, continuation of high fever after hospital admission, and diphasic fever.

Model of the MitoNEET [2Fe-2S] Cluster Shows Proton Coupled Electron Transfer.

MitoNEET is an outer membrane protein whose exact function remains unclear, though a role of this protein in redox and iron sensing as well as in controlling maximum mitochondrial respiratory rates has been discussed. It was shown to contain a redox active and acid labile [2Fe-2S] cluster which is ligated by one histidine and three cysteine residues. Herein we present the first synthetic analogue with biomimetic {SN/S2} ligation which could be structurally characterized in its diferric form, 52-. In addition to being a high fidelity structural model for the biological cofactor, the complex is shown to mediate proton coupled electron transfer (PCET) at the {SN} ligated site, pointing at a potential functional role of the enzyme's unique His ligand. Full PCET thermodynamic square schemes for the mitoNEET model 52- and a related homoleptic {SN/SN} capped [2Fe-2S] cluster 42- are established, and kinetics of PCET reactivity are investigated by double-mixing stopped-flow experiments for both complexes. While the N-H bond dissociation free energy (BDFE) of 5H2- (230 ± 4 kJ mol-1) and the free energy ΔG°PCET for the reaction with TEMPO (-48.4 kJ mol-1) are very similar to values for the homoleptic cluster 4H2- (232 ± 4 kJ mol-1, -46.3 kJ mol-1) the latter is found to react significantly faster than the mitoNEET model (data for 5H2-: k = 135 ± 27 M-1 s-1, ΔH‡ = 17.6 ± 3.0 kJ mol-1, ΔS‡ = -143 ± 11 J mol-1 K-1, and ΔG‡ = 59.8 kJ mol-1 at 293 K). Comparison of the PCET efficiency of these clusters emphasizes the relevance of reorganization energy in this process.

Alterations to mitochondrial fatty-acid use in skeletal muscle after chronic exposure to hypoxia depend on metabolic phenotype.

We investigated the effects of chronic hypoxia on the maximal use of and sensitivity of mitochondria to different substrates in rat slow-oxidative (soleus, SOL) and fast-glycolytic (extensor digitorum longus, EDL) muscles. We studied mitochondrial respiration in situ in permeabilized myofibers, using pyruvate, octanoate, palmitoyl-carnitine (PC), or palmitoyl-coenzyme A (PCoA). The hypophagia induced by hypoxia may also alter metabolism. Therefore, we used a group of pair-fed rats (reproducing the same caloric restriction, as observed in hypoxic animals), in addition to the normoxic control fed ad libitum. The resting respiratory exchange ratio decreased after 21 days of exposure to hypobaric hypoxia (simulated elevation of 5,500 m). The respiration supported by pyruvate and octanoate were unaffected. In contrast, the maximal oxidative respiratory rate for PCoA, the transport of which depends on carnitine palmitoyltransferase 1 (CPT-1), decreased in the rapid-glycolytic EDL and increased in the slow-oxidative SOL, although hypoxia improved affinity for this substrate in both muscle types. PC and PCoA were oxidized similarly in normoxic EDL, whereas chronic hypoxia limited transport at the CPT-1 step in this muscle. The effects of hypoxia were mediated by caloric restriction in the SOL and by hypoxia itself in the EDL. We conclude that improvements in mitochondrial affinity for PCoA, a physiological long-chain fatty acid, would facilitate fatty-acid use at rest after chronic hypoxia independently of quantitative alterations of mitochondria. Conversely, decreasing the maximal oxidation of PCoA in fast-glycolytic muscles would limit fatty-acid use during exercise.NEW & NOTEWORTHY Affinity for low concentrations of long-chain fatty acids (LCFA) in mitochondria skeletal muscles increases after chronic hypoxia. Combined with a lower respiratory exchange ratio, this suggests facility for fatty acid utilization at rest. This fuel preference is related to caloric restriction in oxidative muscle and to hypoxia in glycolytic one. In contrast, maximal oxidation for LCFA is decreased by chronic hypoxia in glycolytic muscle and can explain glucose dependence at exercise.

Pore-scale investigation on the response of heterotrophic respiration to moisture conditions in heterogeneous soils

The relationship between microbial respiration rate and soil moisture content is an important property for understanding and predicting soil organic carbon degradation, CO2 production and emission, and their subsequent effects on climate change. This paper reports a pore-scale modeling study to investigate the response of heterotrophic respiration to moisture conditions in soils and to evaluate various factors that affect this response. X-ray computed tomography was used to derive soil pore structures, which were then used for pore-scale model investigation. The pore-scale results were then averaged to calculate the effective respiration rates as a function of water content in soils. The calculated effective respiration rate first increases and then decreases with increasing soil water content, showing a maximum respiration rate at water saturation degree of 0.75, which is consistent with field and laboratory observations. The relationship between the respiration rate and moisture content is affected by various factors, including pore-scale organic carbon bioavailability, the rate of oxygen delivery, soil pore structure and physical heterogeneity, soil clay content, and microbial drought resistivity. Overall, this study provides mechanistic insights into the soil respiration response to the change in moisture conditions, and reveals a complex relationship between heterotrophic microbial respiration rate and moisture content in soils that is affected by various hydrological, geophysical, and biochemical factors.

Contrasting response of summer soil respiration and enzyme activities to long-term warming and drought in a wet shrubland (NE Wales, UK)

Evaluating the response of soil organic matter decomposition to warming and changes in rainfall is critical to assess the likelihood of proposed positive feedbacks from the terrestrial to the atmospheric system. The response of soil respiration and extracellular activities (EEAs) to long-term warming and recurrent summer drought was studied in a wet shrubland ecosystem in Wales (UK), after 13 years of climate change simulation in a whole-ecosystem experiment. Over a year soil respiration, temperature and moisture was monitored in the field. During the summer season, coinciding with maximum soil respiration rates, soil inorganic N and P, microbial biomass and the extracellular activities (EEAs) of a selection of enzymes involved in C, N and P cycling were analysed. Based on previous field measurements of C and N mineralization, we expected a stronger response of C-cycling EEAs, in comparison to N-cycling EEAs, to drought and warming, and a greater sensitivity of C-cycling EEAs to drought than to warming. Drought had a clear impact on soil respiration during the summer season. However, the availability of inorganic N or P was not significantly affected by the treatments. Microbial biomass and C:N ratio also remained unchanged. In contrast to one of our hypothesis, C-cycling EEAs measured under non-optimal conditions that simulated soil environment in the field (pH of 4.1 and with a temperature incubation of 10°C) showed no significant differences due to long-term warming and recurring drought treatments. Possibly, this assay approach may have obscured treatment effects on the soil enzyme pool. Our results highlight the need for developing methods for the in-situ analysis of EEAs to determine rates of reactions.

Abstract 19455: A Novel Caveolin-1 Frameshift Mutation Shows Critical Regulation of Mitochondrial Structure, Function and Extracellular Transport

Introduction: Caveolins act as molecular scaffolds for signal transduction. A number of caveolin-1 (Cav-1) mutations are associated with pulmonary hypertension with one recent frameshift mutation in exon 3 (c.478_479delTT) resulting in progeroid-like phenotype, lipodystrophy, and pulmonary hypertension. The cellular etiology of these diverse phenotypes is unknown. We have recently shown a critical relationship between caveolins in regulation of mitochondrial function. Hypothesis: As mitochondria are a key to cellular homeostasis, we hypothesize that a Cav-1 mutation (CM) can alter mitochondrial structure and function. Methods and Results: We obtained fibroblasts from a patient containing the CM as well as controls and utilized a variety of microscopy, cell biology, and biochemical techniques to study the role of Cav-1 in mitochondrial regulation. Transmission electron microscopy revealed dysmorphic nuclei, enlarged and increased number of endoplasmic reticulum, and increased number of smaller, swollen, and rounded mitochondria compared to normal cells. Mitochondrial DNA content was increased in CM cells with immunoblots showing decreased mitochondrial fission protein DRP1. Mitotracker staining showed large pools of aggregated mitochondria and DCFDA assay showed 3-fold increase in production of reactive oxygen species in CM cells compared to normal cells (p&lt;0.0001, n=8). Seahorse metabolic flux assay showed lower maximal respiration rates in CM cells compared to normal cells (p=0.0035, n=24). Mitochondrial complex II activity was impaired as addition of succinate failed to evoke any response to ADP in CM cells compared to normal cells. Live 3D microscopy with mitotracker staining showed extracellular mitochondria being transported along structured nanotubes between normal cell; however, impaired extracellular mitochondrial cargo arrangement and movement was observed in cells with the CM. Rescue experiments with Cav-1 overexpression resulted in improved mitochondrial respiration in CM cells. Conclusion: Together the results indicate that Cav-1 may be a critical regulator of mitochondrial structure, function, and transport and mutations in Cav-1 may explain the mitochondrial etiology of a variety of diseases.

APOL1 Renal-Risk Variants Induce Mitochondrial Dysfunction.

APOL1 G1 and G2 variants facilitate kidney disease in blacks. To elucidate the pathways whereby these variants contribute to disease pathogenesis, we established HEK293 cell lines stably expressing doxycycline-inducible (Tet-on) reference APOL1 G0 or the G1 and G2 renal-risk variants, and used Illumina human HT-12 v4 arrays and Affymetrix HTA 2.0 arrays to generate global gene expression data with doxycycline induction. Significantly altered pathways identified through bioinformatics analyses involved mitochondrial function; results from immunoblotting, immunofluorescence, and functional assays validated these findings. Overexpression of APOL1 by doxycycline induction in HEK293 Tet-on G1 and G2 cells led to impaired mitochondrial function, with markedly reduced maximum respiration rate, reserve respiration capacity, and mitochondrial membrane potential. Impaired mitochondrial function occurred before intracellular potassium depletion or reduced cell viability occurred. Analysis of global gene expression profiles in nondiseased primary proximal tubule cells from black patients revealed that the nicotinate phosphoribosyltransferase gene, responsible for NAD biosynthesis, was among the top downregulated transcripts in cells with two APOL1 renal-risk variants compared with those without renal-risk variants; nicotinate phosphoribosyltransferase also displayed gene expression patterns linked to mitochondrial dysfunction in HEK293 Tet-on APOL1 cell pathway analyses. These results suggest a pivotal role for mitochondrial dysfunction in APOL1-associated kidney disease.

Seasonal variation of metabolism in lizard Phrynocephalus vlangalii at high altitude

Seasonal acclimatization is important for animals to live optimally in the varying environment. Phrynocephalus vlangalii, a species of lizard endemic in China, distributes on Qinghai-Tibet Plateau ranging from 2000 to 4600m above sea level. To dissect how this lizard mediate metabolism to adapt various season, the preferred body temperature (Tb), standard metabolic rate (SMR), mitochondrial respiration rates and activities of four metabolic enzymes in this species were tested in different seasons (spring, summer, and autumn). The results showed that the preferred Tb was the lowest in spring and the highest in summer. SMR, maximal mitochondrial respiration rates in liver and skeletal muscle were the highest in spring. Similarly, higher activities of lactate dehydrogenase (LDH), citrate synthase (CS) and cytochrome c oxidase (CCO) activities of liver and skeletal muscle were observed in spring. However, β-hydroxyacyl coenzyme A dehydrogenase (HOAD) activities of liver and skeletal muscle were higher in autumn. On the whole, seasonal variation of metabolism is the highest in spring and the lowest in summer. Seasonal variation of metabolism is the opposite of preferred body temperature, this may be one of the mechanisms to adapt to the environment in P. vlangalii. Our results suggested that P. vlangalii at high altitude has certain adaptive characteristics on metabolism in different seasons.

Hyperpalatable Diet and Physical Exercise Modulate the Expression of the Glial Monocarboxylate Transporters MCT1 and 4.

Hyperpalatable diets (HP) impair brain metabolism, and regular physical exercise has an apparent opposite effect. Here, we combined a prior long-term exposure to HP diet followed by physical exercise and evaluated the impact on some neuroenergetic components and on cognitive performance. We assessed the extracellular lactate concentration, expression of monocarboxylate transporters (MCTs), pyruvate dehydrogenase (PDH), and mitochondrial function in the hippocampus. Male C57BL/6J mice were fed 4months with HP or a control diet. Subsequently, they were divided in the following groups: control diet sedentary (CDS), control diet exercise (CDE), HP diet sedentary (HPS), and HP diet exercise (HPE) (n=15 per group) and were engaged for an additional 30-day period of voluntary exercise and HP diet. Relative to the control situation, exercise increased MCT1, MCT4, and PDH protein levels, while the HP diet increased MCT1 and MCT4 protein levels. The production of hydrogen peroxide (H2O2) and the mitochondrial membrane potential (∆Ѱm) stimulated by succinate in hippocampal homogenates were not significantly different between groups. ADP phosphorylation and the maximal respiratory rate induced by FCCP showed similar responses between groups, implying a normal mitochondrial function. Also, extracellular brain lactate levels were increased in the HPE group compared to other groups soon after performing the Y-maze task. However, such enhanced lactate levels were not associated with improved memory performance. In summary, hippocampal protein expression levels of MCT1 and 4 were increased by physical exercise and HP diet, whereas PDH was only increased by exercise. These observations indicate that a hippocampal metabolic reprogramming takes place in response to these environmental factors.

Hyperglycemia induced damage to mitochondrial respiration in renal mesangial and tubular cells: Implications for diabetic nephropathy

Damage to renal tubular and mesangial cells is central to the development of diabetic nephropathy (DN), a complication of diabetes which can lead to renal failure. Mitochondria are the site of cellular respiration and produce energy in the form of ATP via oxidative phosphorylation, and mitochondrial dysfunction has been implicated in DN. Since the kidney is an organ with high bioenergetic needs, we postulated that hyperglycemia causes damage to renal mitochondria resulting in bioenergetic deficit. The bioenergetic profiles and the effect of hyperglycemia on cellular respiration of human primary mesangial (HMCs) and proximal tubular cells (HK-2) were compared in normoglycemic and hyperglycemic conditions using the seahorse bio-analyzer. In normoglycemia, HK-2 had significantly lower basal, ATP-linked and maximal respiration rates, and lower reserve capacity compared to HMCs. Hyperglycemia caused a down-regulation of all respiratory parameters within 4 days in HK-2 but not in HMCs. After 8 days of hyperglycemia, down-regulation of respiratory parameters persisted in tubular cells with compensatory up-regulated glycolysis. HMCs had reduced maximal respiration and reserve capacity at 8 days, and by 12 days had compromised mitochondrial respiration despite which they did not enhance glycolysis. These data suggest that diabetes is likely to lead to a cellular deficit in ATP production in both cell types, although with different sensitivities, and this mechanism could significantly contribute to the cellular damage seen in the diabetic kidney. Prevention of diabetes induced damage to renal mitochondrial respiration may be a novel therapeutic approach for the prevention/treatment of DN.

Abstract 031: Placental Ischemia Causes Mitochondrial Dysfunction in Reduced Uterine Perfusion Pressure Rats

Introduction: Placental ischemia is believed to be the initial event in the development of preeclampsia (PE). PE is characterized by new onset hypertension and is associated with reduced fetal weight and placental oxidative stress. Mitochondrial dysfunction is the major cause of reactive oxygen species (ROS) generation. We hypothesize that the placental ischemia causes mitochondrial dysfunction in the reduced uterine perfusion pressure (RUPP) rat model of PE. Thus, the purpose of this study is to examine the effect of placental ischemia on mitochondrial function. Methods: Female Sprague Dawley rats were dived into two groups; normal pregnant (NP) and RUPP rats. On gestational day (GD) 14, RUPP surgery was performed, GD18 carotid catheters were inserted, and GD19 conscious blood pressure (MAP) was measured. GD 19 placentas were collected and mitochondria were isolated for respiration measurements. Respiration measurements included: basal state (isolated mitochondria with no substrates), state 2 (glutamate and malate as complex I substrates), state 3 (ADP stimulated), leak state (oligomycin induced ATP synthase inhibition), and maximal state (FCCP stimulated uncoupled). Cytochrome C oxidase (complex IV) activity was also measured in isolated mitochondria. Results: MAP was elevated in RUPP (n=16) compared to NP rats (n=14) (119±2 vs. 100±2 mmHg, p&lt;0.05). State 3 (149±8 vs 220±12 pmol/sec/mg, p&lt;0.05) and maximal (123±8 vs 159±5 pmol/sec/mg, p&lt;0.05) respiration rates were significantly reduced in RUPP (n=5) vs NP (n=7) rats. However, basal, state 2, and state 4 respiratory rates were not different between RUPP vs NP. Respiratory control ratio (state 3/ state 4) was significantly reduced in RUPP (n=5) vs NP (n=7) (7±1 vs 10±1, p&lt;0.05). Complex IV activity was reduced in RUPP (n=5) vs. NP (n=6) rats (508±14 vs 644±17 nmol e-/min/mg, P&lt;0.05). Conclusion: Reduced mitochondrial respiration and cytochrome c oxidase activity, indicators of mitochondrial dysfunction, occur in response to placental ischemia during pregnancy. Thus, suggesting that mitochondrial dysfunction may contribute to the pathophysiology of oxidative stress and hypertension during PE.

Left Ventricular Transmural Gradient in Mitochondrial Respiration Is Associated with Increased Sub-Endocardium Nitric Oxide and Reactive Oxygen Species Productions.

Objective: Left ventricle (LV) transmural gradient in mitochondrial respiration has been recently reported. However, to date, the physiological mechanisms involved in the lower endocardium mitochondrial respiration chain capacity still remain to be determined. Since, nitric oxide (NO) synthase expression in the heart has spatial heterogeneity and might impair mitochondrial function, we investigated a potential association between LV transmural NO and mitochondrial function gradient.Methods: Maximal oxidative capacity (VMax) and relative contributions of the respiratory chain complexes II, III, IV (VSucc) and IV (VTMPD), mitochondrial content (citrate synthase activity), coupling, NO (electron paramagnetic resonance), and reactive oxygen species (ROS) production (H2O2 and dihydroethidium (DHE) staining) were determined in rat sub-endocardium (Endo) and sub-epicardium (Epi). Further, the effect of a direct NO donor (MAHMA NONOate) on maximal mitochondrial respiratory rates (Vmax) was determined.Results: Mitochondrial respiratory chain activities were reduced in the Endo compared with the Epi (−16.92%; P = 0.04 for Vmax and –18.73%; P = 0.02, for Vsucc, respectively). NO production was two-fold higher in the Endo compared with the Epi (P = 0.002) and interestingly, increasing NO concentration reduced Vmax. Mitochondrial H2O2 and LV ROS productions were significantly increased in Endo compared to Epi, citrate synthase activity and mitochondrial coupling being similar in the two layers.Conclusions: LV mitochondrial respiration transmural gradient is likely related to NO and possibly ROS increased production in the sub-endocardium.

The Effects of Fentanyl on Hepatic Mitochondrial Function.

Remifentanil interferes with hepatic mitochondrial function. The aim of the present study was to evaluate whether hepatic mitochondrial function is affected by fentanyl, a more widely used opioid than remifentanil. Human hepatoma HepG2 cells were exposed to fentanyl or pretreated with naloxone (an opioid receptor antagonist) or 5-hydroxydecanoate (5-HD, an inhibitor of mitochondrial adenosine triphosphate (ATP)-sensitive potassium [mitoKATP] channels), followed by incubation with fentanyl. Mitochondrial function and metabolism were then analyzed. Fentanyl marginally reduced maximal mitochondrial complex-specific respiration rates using exogenous substrates (decrease in medians: 11%-18%; P = 0.003-0.001) but did not affect basal cellular respiration rates (P = 0.834). The effect on stimulated respiration was prevented by preincubation with naloxone or 5-HD. Fentanyl reduced cellular ATP content in a dose-dependent manner (P < 0.001), an effect that was not significantly prevented by 5-HD and not explained by increased total ATPase concentration. However, in vitro ATPase activity of recombinant human permeability glycoprotein (an ATP-dependent drug efflux transporter) was significantly stimulated by fentanyl (P = 0.004). Our data suggest that fentanyl reduces stimulated mitochondrial respiration of cultured human hepatocytes by a mechanism that is blocked by a mitoKATP channel antagonist. Increased energy requirements for fentanyl efflux transport may offer an explanation for the substantial decrease in cellular ATP concentration.

Oxygen utilization rate (OUR) underestimates ocean respiration: A model study

AbstractWe use a simple 1‐D model representing an isolated density surface in the ocean and 3‐D global ocean biogeochemical models to evaluate the concept of computing the subsurface oceanic oxygen utilization rate (OUR) from the changes of apparent oxygen utilization (AOU) and water age. The distribution of AOU in the ocean is not only the imprint of respiration in the ocean's interior but is strongly influenced by transport processes and eventually loss at the ocean surface. Since AOU and water age are subject to advection and diffusive mixing, it is only when they are affected both in the same way that OUR represents the correct rate of oxygen consumption. This is the case only when advection prevails or with uniform respiration rates, when the proportions of AOU and age are not changed by transport. In experiments with the 1‐D tube model, OUR underestimates respiration when maximum respiration rates occur near the outcrops of isopycnals and overestimates when maxima occur far from the outcrops. Given the distribution of respiration in the ocean, i.e., elevated rates near high‐latitude outcrops of isopycnals and low rates below the oligotrophic gyres, underestimates are the rule. Integrating these effects globally in three coupled ocean biogeochemical and circulation models, we find that AOU‐over‐age based calculations underestimate true model respiration by a factor of 3. Most of this difference is observed in the upper 1000 m of the ocean with the discrepancies increasing toward the surface where OUR underestimates respiration by as much as factor of 4.