Effect of mild carboxy-hemoglobin on exercising skeletal muscle: intravascular and intracellular evidence

We studied muscle blood flow, muscle oxygen uptake (VO(2)), net muscle CO uptake, Mb saturation, and intracellular bioenergetics during incremental single leg knee-extensor exercise in five healthy young subjects in conditions of normoxia, hypoxia (H; 11% O(2)), normoxia + CO (CO(norm)), and 100% O(2) + CO (CO(hyper)). Maximum work rates and maximal oxygen uptake (VO(2 max)) were equally reduced by approximately 14% in H, CO(norm), and CO(hyper). The reduction in arterial oxygen content (Ca(O(2))) (approximately 20%) resulted in an elevated blood flow (Q) in the CO and H trials. Net muscle CO uptake was attenuated in the CO trials. Suprasystolic cuff measurements of the deoxy-Mb signal were not different in terms of the rate of signal rise or maximum signal attained with and without CO. At maximal exercise, calculated mean capillary PO(2) was most reduced in H and resulted in the lowest Mb-associated PO(2). Reductions in ATP, PCr, and pH during H, CO(norm), and CO(hyper) occurred earlier during progressive exercise than in normoxia. Thus the effects of reduced Ca(O(2)) due to mild CO poisoning are similar to H.     

Evolving techniques for the investigation of muscle bioenergetics and oxygenation

Magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) are both powerful, non-invasive methodologies and, as such, offer great potential to investigate both human biochemistry and human physiology, and ultimately to contribute significantly to the field of medicine. Consequently there has been much effort devoted to fostering the evolution of these methodologies into distinct and applicable techniques. Here we will highlight several MRI and MRS techniques for the assessment of human biochemistry and physiology that ultimately may provide useful clinical assessments and diagnoses of various muscular and cardiovascular pathologies. Specifically, the evolving techniques that will be discussed are: (1) (1)H MRS of myoglobin to assess the intracellular partial pressure of O(2), (2) (31)P MRS to assess metabolic capacity, and (3) the combination of (31)P chemical shift imaging to assess local metabolic demand (oxygen uptake; .VO(2)) with arterial spin labelling to assess local perfusion (blood flow; .Q), in an effort to characterize the elusive spatial matching of skeletal muscle (.Q/.VO(2)).     

Accelerated evolution of the mitochondrial genome in an alloplasmic line of durum wheat

Background

Wheat is an excellent plant species for nuclear mitochondrial interaction studies due to availability of large collection of alloplasmic lines. These lines exhibit different vegetative and physiological properties than their parents. To investigate the level of sequence changes introduced into the mitochondrial genome under the alloplasmic condition, three mitochondrial genomes of the Triticum-Aegilops species were sequenced: 1) durum alloplasmic line with the Ae. longissima cytoplasm that carries the T. turgidum nucleus designated as (lo) durum, 2) the cytoplasmic donor line, and 3) the nuclear donor line.

Results

The mitochondrial genome of the T. turgidum was 451,678 bp in length with high structural and nucleotide identity to the previously characterized T. aestivum genome. The assembled mitochondrial genome of the (lo) durum and the Ae. longissima were 431,959 bp and 399,005 bp in size, respectively. The high sequence coverage for all three genomes allowed analysis of heteroplasmy within each genome. The mitochondrial genome structure in the alloplasmic line was genetically distant from both maternal and paternal genomes. The alloplasmic durum and the Ae. longissima carry the same versions of atp6, nad6, rps19-p, cob and cox2 exon 2 which are different from the T. turgidum parent. Evidence of paternal leakage was also observed by analyzing nad9 and orf359 among all three lines. Nucleotide search identified a number of open reading frames, of which 27 were specific to the (lo) durum line.

Conclusions

Several heteroplasmic regions were observed within genes and intergenic regions of the mitochondrial genomes of all three lines. The number of rearrangements and nucleotide changes in the mitochondrial genome of the alloplasmic line that have occurred in less than half a century was significant considering the high sequence conservation between the T. turgidum and the T. aestivum that diverged from each other 10,000 years ago. We showed that the changes in genes were not limited to paternal leakage but were sufficiently significant to suggest that other mechanisms, such as recombination and mutation, were responsible. The newly formed ORFs, differences in gene sequences and copy numbers, heteroplasmy, and substoichiometric changes show the potential of the alloplasmic condition to accelerate evolution towards forming new mitochondrial genomes.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-67) contains supplementary material, which is available to authorized users.     

Developmental expression of the cucumber <Emphasis Type="Italic">Cs-XTH1</Emphasis> and <Emphasis Type="Italic">Cs-XTH3</Emphasis> genes,encoding xyloglucan endotransglucosylase/hydrolases,can be influenced by mechanical stimuli

The expression of two genes encoding xyloglucan endotransglucosylase/hydrolases (XTHs), Cs-XTH1 and Cs-XTH3, was upregulated during the onset of cucumber somatic embryogenesis. As a means of characterising the developmental regulation of these genes, the activity of the respective upstream regulatory regions was investigated in seedlings and somatic embryos of Arabidopsis thaliana and Cucumis sativus. GUS assays revealed that both genes are under developmental control. In addition, elevated promoter activity was found in the tension-bearing regions of the plant and in response to touch and wounding, which is consistent with the existence of numerous stress-related cis elements in the 5′-regulatory regions. In vivo xyloglucan endotransglucosylase (XET) action assays were performed to gain an overview on the role of XTHs during somatic embryogenesis. The highest XET action was observed in the external cell layers of somatic embryos in the cotyledonary region and in the presumptive region of peg formation. Based on the results, we propose a dual mechanism (one developmental and the second adaptive) for the regulation of Cs-XTH1 and Cs-XTH3 activity wherein the developmental pattern can be modified by mechanical stimuli.     

Cellular PO2 as a determinant of maximal mitochondrial O(2) consumption in trained human skeletal muscle.

Previously, by measuring myoglobin-associated PO(2) (P(Mb)O(2)) during maximal exercise, we have demonstrated that 1) intracellular PO(2) is 10-fold less than calculated mean capillary PO(2) and 2) intracellular PO(2) and maximum O(2) uptake (VO(2 max)) fall proportionately in hypoxia. To further elucidate this relationship, five trained subjects performed maximum knee-extensor exercise under conditions of normoxia (21% O(2)), hypoxia (12% O(2)), and hyperoxia (100% O(2)) in balanced order. Quadriceps O(2) uptake (VO(2)) was calculated from arterial and venous blood O(2) concentrations and thermodilution blood flow measurements. Magnetic resonance spectroscopy was used to determine myoglobin desaturation, and an O(2) half-saturation pressure of 3.2 Torr was used to calculate P(Mb)O(2) from saturation. Skeletal muscle VO(2 max) at 12, 21, and 100% O(2) was 0.86 +/- 0.1, 1.08 +/- 0.2, and 1.28 +/- 0.2 ml. min(-1). ml(-1), respectively. The 100% O(2) values approached twice that previously reported in human skeletal muscle. P(Mb)O(2) values were 2.3 +/- 0.5, 3.0 +/- 0.7, and 4.1 +/- 0.7 Torr while the subjects breathed 12, 21, and 100% O(2), respectively. From 12 to 21% O(2), VO(2) and P(Mb)O(2) were again proportionately related. However, 100% O(2) increased VO(2 max) relatively less than P(Mb)O(2), suggesting an approach to maximal mitochondrial capacity with 100% O(2). These data 1) again demonstrate very low cytoplasmic PO(2) at VO(2 max), 2) are consistent with supply limitation of VO(2 max) of trained skeletal muscle, even in hyperoxia, and 3) reveal a disproportionate increase in intracellular PO(2) in hyperoxia, which may be interpreted as evidence that, in trained skeletal muscle, very high mitochondrial metabolic limits to muscle VO(2) are being approached.     

Riparian populations of minnesota reed canarygrass (Phalaris arundinacea) are most likely native,based on SNPs (DArTseqLD)

The native vs. exotic status of reed canarygrass (RCG), a major invasive species of Minnesota wetlands, is unknown. The aim of this study was to investigate this native vs. exotic status to enhance its management. Genetic comparison of wild RCG populations from six Minnesota and six Czech Republic rivers was performed. A total of 2521 polymorphic SNP markers (single nucleotide polymorphisms) were used to evaluate 478 RCG samples across all collections. In the PCoA, all (n = 256) tested extant wild, riparian RCG genotypes from six Minnesota Rivers and six Czech Republic Rivers were genetically distinct, although some SNPs were common in both populations since they are the same species. DAPC analysis also resulted in the formation of two primary clusters separating the Minnesota Rivers and Czech Republic Rivers riparian samples, with little overlap; STRUCTURE analysis also supported this clustering with k = 4 groups as it separated the Czech Republic Rivers populations into three groups, along with Minnesota Rivers. The uniformity of PCoA, DAPC, STRUCTURE, and Evanno results indicates the distinct separation of Minnesota Rivers and Czech Republic Rivers populations. Portions of the genome (specific SNPs) are preserved or in common across continents, as indicated by STRUCTURE similarities. Nonetheless, overall significant SNP differences between the continents indicate that the Minnesota riparian populations are distinct enough from the European (Czech) collections to be delineated as native N. American RCG. PCoA of all the Minnesota RCG collections clustered Minnesota Rivers, Herbarium, Extant Herbarium, Research Field and Native Field collections together. STRUCTURE analysis (k = 2; Evanno) divided these Minnesota collections from the Commercial Field and Cultivars collections. There are two genetically distinct groups of RCG in Minnesota and since the Minnesota Rivers, the Research Field, the Native Field and pre-1930 herbaria collections clustered together, they are most likely native N. American types. Analysis of molecular variance (AMOVA) indicated that the genetic variation was more significant within, rather than among, the RCG populations. Native, historic herbaria types cluster together with all wild RCG river populations in Minnesota, all of which were distinct from those in Central Europe, suggesting native RCG type persistence in N. America. Also, cultivated forage types of RCG are distinct from wild RCG Minnesota river populations. The SNP genetic data shows that riparian Minnesota RCG populations are native. These data will facilitate future management strategies to control RCG as a native, but invasive, species.

     

Skeletal muscle intracellular PO(2) assessed by myoglobin desaturation: response to graded exercise.

The relationship between skeletal muscle intracellular PO(2) (iPO(2)) and progressive muscular work has important implications for the understanding of O(2) transport and utilization. Presently there is debate as to whether iPO(2) falls progressively with increasing O(2) demand or reaches a plateau from moderate to maximal metabolic demand. Thus, using (1)H magnetic resonance spectroscopy of myoglobin (Mb), we studied cellular oxygenation during progressive single-leg knee extensor exercise from unweighted to 100% of maximal work rate in six active human subjects. In all subjects, the Mb peak at 73 ppm was not visible at rest, whereas the peak was small or indistinguishable from the noise in the majority of subjects during progressive exercise from unweighted to 50-60% of maximum work rate. In contrast, beyond this exercise intensity, a Mb peak of consistent magnitude was discernible in all subjects. When a Mb half saturation of 3.2 Torr was used, the calculated skeletal muscle PO(2) was variable before 60% of maximum work rate but in general was relatively high (>18 Torr, the measurable PO(2) with the poorest signal-to-noise ratio, in the majority of cases), whereas beyond this exercise intensity iPO(2) fell to a relatively uniform and invariant level of 3.8 +/- 0.5 Torr across all subjects. These results do not support the concept of a progressive linear fall in iPO(2) across increasing work rates. Instead, this study documents variable but relatively high iPO(2) from rest to moderate exercise and again confirms that from 50-60% of maximum work rate iPO(2) reaches a plateau that is then invariant with increasing work rate.     

Impaired muscle oxygen transfer in patients with chronic renal failure

We hypothesized that impaired O2 transport plays a role in limiting exercise in patients with chronic renal failure (CRF). Six CRF patients (25 +/- 6 yr) and six controls (24 +/- 6 yr) were examined twice during incremental single-leg isolated quadriceps exercise. Leg O2 delivery (QO2(leg)) and leg O2 uptake (VO2(leg)) were obtained when subjects breathed gas of three inspired O2 fractions (FI(O2)) (0.13, 0.21, and 1.0). On a different day, myoglobin O2 saturation and muscle bioenergetics were measured by proton and phosphorus magnetic resonance spectroscopy. CRF patients, but not controls, showed O2 supply dependency of peak VO2 (VO2(peak)) by a proportional relationship between peak VO2(leg) at each inspired O2 fraction (0.59 +/- 0.20, 0.47 +/- 0.10, 0.43 +/- 0.10 l/min, respectively) and 1) work rate (933 +/- 372, 733 +/- 163, 667 +/- 207 g), 2) QO(2leg) (0.80 +/- 0.20, 0.64 +/- 0.10, 0.59 +/- 0.10 l/min), and 3) cell PO2 (6.3 +/- 5.4, 1.7 +/- 1.3, 1.2 +/- 0.7 mmHg). CRF patients breathing 100% O2 and controls breathing 21% O2 had similar peak QO2(leg) (0.80 +/- 0.20 vs. 0.79 +/- 0.10 l/min) and similar peak VO2(leg) (0.59 +/- 0.20 vs. 0.57 +/- 0.10 l/min). However, mean capillary PO2 (47.9 +/- 4.0 vs. 38.2 +/- 4.6 mmHg) and the capillary-to-myocite gradient (40.7 +/- 6.2 vs. 34.4 +/- 4.0 mmHg) were both higher in CRF patients than in controls (P < 0.03 each). We conclude that low muscle O2 conductance, but not limited mitochondrial oxidative capacity, plays a role in limiting exercise tolerance in these patients.     

Lactate efflux from exercising human skeletal muscle: role of intracellular PO2

It remainscontroversial whether lactate formation during progressive dynamicexercise from submaximal to maximal effort is due to muscle hypoxia. Tostudy this question, we used direct measures of arterial and femoralvenous lactate concentration, a thermodilution blood flow technique,phosphorus magnetic resonance spectroscopy (MRS), and myoglobin (Mb)saturation measured by ¹H nuclearMRS in six trained subjects performing single-leg quadriceps exercise.We calculated net lactate efflux from the muscle and intracellularPO₂ with subjects breathing room airand 12% O₂. Data were obtained at50, 75, 90, and 100% of quadriceps maximalO₂ consumption at each fraction ofinspired O₂. Mb saturation wassignificantly lower in hypoxia than in normoxia [40 ± 3 vs. 49 ± 3% (SE)] throughout incremental exercise to maximalwork rate. With the assumption of aPO₂ at which 50% of Mb-binding sitesare bound with O₂ of 3.2 Torr,Mb-associated PO₂ averaged 3.1 ± 0.3 and 2.3 ± 0.2 Torr in normoxia and hypoxia, respectively. Netblood lactate efflux was unrelated to intracellular PO₂ across the range of incrementalexercise to maximum (r = 0.03 and 0.07 in normoxia and hypoxia, respectively) but linearly related toO₂ consumption(r = 0.97 and 0.99 in normoxia andhypoxia, respectively) with a greater slope in 12%O₂. Net lactate efflux was alsolinearly related to intracellular pH(r = 0.94 and 0.98 in normoxia andhypoxia, respectively). These data suggest that with increasing workrate, at a given fraction of inspiredO₂, lactate efflux is unrelated tomuscle cytoplasmic PO₂, yet theefflux is higher in hypoxia. Catecholamine values from comparablestudies are included and indicate that lactate efflux in hypoxia may bedue to systemic rather than intracellular hypoxia.