Presence of procoagulant peripheral blood mononuclear cells in severe COVID-19 patients relate to ventilation perfusion mismatch and precede pulmonary embolism.

PURPOSE: Pulmonary emboli (PE) contribute substantially to coronavirus disease 2019 (COVID-19) related mortality and morbidity. Immune cell-mediated hyperinflammation drives the procoagulant state in COVID-19 patients, resulting in immunothrombosis. To study the role of peripheral blood mononuclear cells (PBMC) in the procoagulant state of COVID-19 patients, we performed a functional bioassay and related outcomes to the occurrence of PE. Secondary aims were to relate this functional assay to plasma D-dimer levels, ventilation perfusion mismatch and TF expression on monocyte subsets. METHODS: PBMC from an ICU biobank were obtained from 20 patients with a computed tomography angiograph (CTA) proven PE and compared to 15 COVID-19 controls without a proven PE. Functional procoagulant properties of PBMC were measured using a modified fibrin generation time (MC-FGT) assay. Tissue factor (TF) expression on monocyte subsets were measured by flow cytometry. Additional clinical data were obtained from patient records including end-tidal to arterial carbon dioxide gradient. RESULTS: MC-FGT levels were highest in the samples taken closest to the PE detection, similar to the end-tidal to arterial carbon dioxide gradient (ETCO2 - PaCO2), a measurement to quantify ventilation-perfusion mismatch. In patients without proven PE, peak MC-FGT relates to an increase in end-tidal to arterial carbon dioxide gradient. We identified non-classical, CD16 positive monocytes as the subset with increased TF expression. CONCLUSION: We show that the procoagulant state of PBMC could aid in early detection of PE in COVID-19 ICU patients. Combined with end-tidal to ETCO2 - PaCO2 gradient, these tests could improve early detection of PE on the ICU.

Animal and translational models of SARS-CoV-2 infection and COVID-19.

COVID-19 is causing a major once-in-a-century global pandemic. The scientific and clinical community is in a race to define and develop effective preventions and treatments. The major features of disease are described but clinical trials have been hampered by competing interests, small scale, lack of defined patient cohorts and defined readouts. What is needed now is head-to-head comparison of existing drugs, testing of safety including in the background of predisposing chronic diseases, and the development of new and targeted preventions and treatments. This is most efficiently achieved using representative animal models of primary infection including in the background of chronic disease with validation of findings in primary human cells and tissues. We explore and discuss the diverse animal, cell and tissue models that are being used and developed and collectively recapitulate many critical aspects of disease manifestation in humans to develop and test new preventions and treatments.

Evaluation of DXA vs. MRI for body composition measures in 1-month olds.

BACKGROUND: Detailed measures of infant body composition are needed for understanding the impact of genes and environment on growth early in life. OBJECTIVE: The purpose of this study was to compare the accuracy and bias of body composition in infants. METHODS: Dual energy X-ray absorptiometry (DXA) and magnetic resonance imaging (MRI) were used to determine body composition and the trunk depot. The depots measured were total fat mass (FM), total fat-free mass (FFM) and trunk FM and FFM using DXA and MRI in 14 infants. RESULTS: None of the regression lines between DXA and MRI significantly deviate from the line of identity for any of the depots studied. However, Bland-Altman analyses revealed bias for trunk FM and trunk FFM. CONCLUSION: Our data showed DXA to be accurate (regression not significantly deviating from the line of identity), with high agreement (indicated by high R(2) ) and without bias (non-significant Bland-Altman) when estimating total FM and FFM. This could not be said for trunk estimates.

Diabetes in old age: an emerging epidemic.

Diabetes in the elderly is emerging as one of the most important public health problems of the 21st century. In developing countries, the majority of people with diabetes are in the age range of 45-64 years. A better understanding on the pathogenesis of diabetes in the aging population is required to successfully treat and prevent its devastating complications. Changes in body composition with accumulation of fat in the abdomen is a key factor in the causation of diabetes in the aging population. The size and strength of skeletal muscle, a major tissue involved in glucose metabolism, also declines leading to muscle weakness and a reduction in physical activity. These changes lead to marked reduction in energy expenditure and abdominal fat accumulation causing insulin resistance. Recent evidence suggests that four months of aerobic exercise can improve muscle oxidative capacity similarly in younger and older people, but that insulin sensitivity is less likely to improve in older people. It appears that older people need to exercise more frequently to improve their insulin sensitivity. Diagnosis and management of diabetes in the elderly requires special attention since age, genetics, body composition and lifestyle factors all interact. Increasing evidence suggests that postprandial hyperglycemia is more sensitive to diagnose diabetes in elderly people than in the young. Age related changes in body function and cognition demand special caution in the selection of hypoglycemic drugs in the elderly. Targets of diabetes therapy in the elderly have to be individualized, considering the age of the patient, remaining life-expectancy and severity of co-morbid conditions. Short acting insulin secretogogues are preferred to avoid prolonged and frequent hypoglycemia. Judicious choice of insulin sensitizers, timely introduction of insulin, meticulous control of hypertension and hyperlipidemia are critical to prevent complications.

Effects of caloric restriction on mitochondrial function and gene transcripts in rat muscle.

Rodent skeletal muscle mitochondrial DNA has been shown to be a potential site of oxidative damage during aging. Caloric restriction (CR) is reported to reduce oxidative stress and prolong life expectancy in rodents. Gene expression profiling and measurement of mitochondrial ATP production capacity were performed in skeletal muscle of male rats after feeding them either a control diet or calorie-restricted diet (60% of control diet) for 36 wk to determine the potential mechanism of the beneficial effects of CR. CR enhanced the transcripts of genes involved in reactive oxygen free radical scavenging function, tissue development, and energy metabolism while decreasing expression of those genes involved in signal transduction, stress response, and structural and contractile proteins. Real-time PCR measurements confirmed the changes in transcript levels of cytochrome-c oxidase III, superoxide dismutase (SOD)1, and SOD2 that were noted by the microarray approach. Mitochondrial ATP production and citrate synthase were unaltered by the dietary changes. We conclude that CR alters transcript levels of several genes in skeletal muscle and that mitochondrial function in skeletal muscle remains unaltered by the dietary intervention. Alterations in transcripts of many genes involved in reactive oxygen scavenging function may contribute to the increase in longevity reported with CR.

Impact of high-fat diet and antioxidant supplement on mitochondrial functions and gene transcripts in rat muscle.

High-fat diets are reported to increase oxidative stress in a variety of tissues, whereas antioxidant supplementation prevents many diseases attributed to high-fat diet. Rodent skeletal muscle mitochondrial DNA has been shown to be a potential site of oxidative damage. We hypothesized that the effects of a high-fat diet on skeletal muscle DNA functions would be attenuated or partially reversed by antioxidant supplementation. Gene expression profiling and measurement of mitochondrial ATP production capacity were performed in skeletal muscle from male rats after feeding one of three diets (control, high-fat diet with or without antioxidants) for 36 wk. The high-fat diet altered transcript levels of 18 genes of 800 surveyed compared with the control-fed rats. Alterations included reduced expression of genes involved in free-radical scavenging and tissue development and increased expression of stress response and signal transduction genes. The magnitude of these alterations due to high-fat diet was reduced by antioxidant supplementation. Real-time PCR measurements confirmed the changes in transcript levels of cytochrome c oxidase subunit III and superoxide dismutase-1 and -2 noted by microarray approach. Mitochondrial ATP production was unaltered by dietary changes or antioxidant supplementation. It is concluded that the high-fat diet increases the transcription of genes involved in stress response but reduces those of free-radical scavenger enzymes, resulting in reduced DNA repair/metabolism (increased DNA damage). Antioxidants partially prevent these changes. Mitochondrial functions in skeletal muscle remain unaltered by the dietary intervention due to many adaptive changes in gene transcription.

Tissue-specific regulation of mitochondrial and cytoplasmic protein synthesis rates by insulin.

In vivo studies have reported conflicting effects of insulin on mixed tissue protein synthesis rates. To test the hypothesis that insulin has differential effects on synthesis rates of various protein fractions in different organs, we infused miniature swine (n = 8 per group) with saline, insulin alone (at 0.7 mU/kg(-1). min(-1)), or insulin plus an amino acid mixture for 8 h. Fractional synthesis rate (FSR) of mitochondrial and cytoplasmic proteins in liver, heart, and skeletal muscle, as well as myosin heavy chain (MHC) in muscle, were measured using L-[1-(13)C]leucine as a tracer. The FSR of mitochondrial and cytoplasmic proteins were highest in liver, followed by heart and then muscle. Mitochondrial FSR in muscle was higher during insulin and insulin plus amino acid infusions than during saline. Insulin had no significant effect on FSR of MHC in muscle. In contrast, FSR of both mitochondrial and cytoplasmic proteins were not stimulated by insulin in liver. Insulin also did not increase FSR of mitochondrial in heart, whereas insulin and amino acid stimulated FSR of cytoplasmic protein. In conclusion, insulin stimulates the synthesis of muscle mitochondrial proteins, with no significant stimulatory effect on synthesis of sarcoplasmic and MHC. These results demonstrate that insulin has different effects on synthesis rates of specific protein fractions in the liver, heart, and skeletal muscle.

Dopaminergic and cholinergic antagonism in a novel-object detection task with rats.

In a free-choice test, rats display a tendency to interact more with a novel object than a familiar object. In the present report, we assessed the role of the dopaminergic and cholinergic systems in the expression of this novelty detection. Rats were injected with a dopaminergic antagonist (sulpiride, U-99194A, clozapine, or L-745,870) or a cholinergic antagonist (mecamylamine or scopolamine) prior to the free-choice novel-object test. The dopamine antagonists did not block novel-object detection. In contrast, scopolamine, but not mecamylamine, reliably blocked the expression of novelty detection, indicating a role for muscarinic receptors.

Does aging adversely affect muscle mitochondrial function?

Muscle oxidative function appears to decline with aging, and evidence suggests that this is related to reduced synthesis of mitochondrial and other muscle proteins. Causes for these events may include mtDNA damage or reduced mtDNA copy numbers, reduced oxidative enzyme activities and ATP production, and increased proton leak.

T(3) increases mitochondrial ATP production in oxidative muscle despite increased expression of UCP2 and -3.

Triiodothyronine (T(3)) increases O(2) and nutrient flux through mitochondria (Mito) of many tissues, but it is unclear whether ATP synthesis is increased, particularly in different types of skeletal muscle, because variable changes in uncoupling proteins (UCP) and enzymes have been reported. Thus Mito ATP production was measured in oxidative and glycolytic muscles, as well as in liver and heart, in rats administered T(3) for 14 days. Relative to saline-treated controls, T(3) rats had 80, 168, and 62% higher ATP production in soleus muscle, liver, and heart, respectively, as well as higher activities of citrate synthase (CS; 63, 90, 25%) and cytochrome c oxidase (COX; 119, 225, 52%) in the same tissues (all P < 0.01). In plantaris muscle of T(3) rats, CS was only slightly higher (17%, P < 0.05) than in controls, and ATP production and COX were unaffected. mRNA levels of COX I and III were 33 and 47% higher in soleus of T(3) rats (P < 0.01), but there were no differences in plantaris. In contrast, UCP2 and -3 mRNAs were 2.5- to 14-fold higher, and protein levels were 3- to 10-fold higher in both plantaris and soleus of the T(3) group. We conclude that T(3) increases oxidative enzymes and Mito ATP production and Mito-encoded transcripts in oxidative but not glycolytic rodent tissues. Despite large increases in UCP expression, ATP production was enhanced in oxidative tissues and maintained in glycolytic muscle of hyperthyroid rats.

Muscle protein metabolism and the sarcopenia of aging.

Loss of muscle mass, strength, and oxidative capacity accompanies normal aging in humans. The mechanisms responsible for these changes remain to be clearly defined. Muscle protein mass and function depend on protein turnover. Synthesis rate of the major muscle contractile protein, myosin heavy chain (MHC), and transcript levels of fast MHC isoforms decrease in association with strength reductions, while mitochondrial protein synthesis rate declines in parallel with activities of mitochondrial enzymes and maximal oxidative capacity (VO2max). Resistance exercise training increases the synthesis rate of MHC and transcript levels of the slow MHC isoform in older humans, along with increasing muscle strength. The relationship between the synthesis of muscle proteins, and muscle size and function, with aging and exercise training are discussed in this review.

Effects of aging on mitochondrial DNA copy number and cytochrome c oxidase gene expression in rat skeletal muscle, liver, and heart.

Mitochondrial DNA (mtDNA) deletions and mutations have been reported to occur with aging in various tissues. To determine the functional impact of these changes, we measured mtDNA copy number, mitochondria-encoded cytochrome c oxidase (COX) subunit I and III transcript levels, and COX enzyme activity in skeletal muscles (medial and lateral gastrocnemius and soleus), liver, and heart in 6- and 27-month-old rats. Substantial age-related reductions of mtDNA copy number occurred in skeletal muscle groups (-23-40%, p < 0.03) and liver (-50%, p < 0.01) but not in the heart. The decline in mtDNA was not associated with reduced COX transcript levels in tissues with high oxidative capacities such as red soleus muscle or liver, while transcript levels were reduced with aging in the less oxidative mixed fiber gastrocnemius muscle (-17-22%, p < 0.05). Consistent with transcript levels, COX activity also remained unchanged in aging liver and heart but declined with age in the lateral gastrocnemius (-32%, p < 0.05). Thus, the effects of aging on mitochondrial gene expression are tissue-specific. A substantial age-related decline in mtDNA copy number proportional to tissue oxidative capacities is demonstrated in skeletal muscle and liver. mtDNA levels are in contrast preserved in the aging heart muscle, presumably due to its incessant aerobic activity. Reduced mtDNA copy number has no major effects on mitochondrial encoded transcript levels and enzyme activities in various tissues under these base-line study conditions. In contrast, maintenance of mitochondrial transcript levels that may be linked to oxidative metabolism and energy demand appears to be the main determinant of mitochondrial oxidative capacity in aging tissues.

The effect of age on protein metabolism.

The mechanisms of senescence remain to be fully defined. This review focuses on recent advances in our understanding of body protein turnover, which is essential for the remodeling of tissues and production of specific proteins in time of need. Recent advances in technology make it possible to measure the synthesis rate of muscle myosin heavy chain, mitochondrial proteins and sarcoplasmic proteins, providing insight into the mechanisms of the sarcopenia of aging. A reduced synthesis rate of myosin heavy chain and mitochondrial protein may explain muscle weakness and fatiguability that occurs with aging. Aging also seems to affect selected liver proteins such as fibrinogen. The potential roles of exercise and hormone replacement in slowing the age-related decline in protein turnover is discussed.

Mechanisms of sarcopenia of aging.

Age-related sarcopenia is characterized by decreased muscle mass and muscle strength, and increased muscle fatigability. A decrease in synthesis rates of mixed muscle proteins (average of all muscle proteins), myosin heavy chain (responsible for adenosine triphosphatase action) and mitochondrial proteins (site of adenosine triphosphate production) have been described with aging. Most of these changes start by middle age, thus contributing to the progressive decline in muscle size and function. How closely these changes are related to lifestyle and the decline in several hormones, particularly growth hormone, insulin-like growth factor-I, testosterone and dehydroepiandrosterone, remains to be clearly defined. The ability to measure the specific effects of different types of exercise training on muscle protein metabolism has only recently become available. Thus, future investigations will continue to improve our understanding of protein metabolism in aging skeletal muscles. The development and assessment of successful countermeasures to age-related sarcopenia will hopefully follow these discoveries.

Whole body protein kinetics using Phe and Tyr tracers: an evaluation of the accuracy of approximated flux values.

Phenylalanine (Phe) kinetics are increasingly used in studies of amino acid kinetics, because the metabolic fate of Phe is limited to incorporation into protein (protein synthesis, Sp) and catabolism via hydroxylation (Qpt) to tyrosine (Tyr). Besides an infusion of labeled Phe to measure Phe flux (Qp), a priming dose of Tyr and an independent Tyr tracer are used to measure Tyr flux (Qt) and Qpt. Alternatively, Qt, Qpt, and Sp can be approximated by using equations, based on Phe and Tyr concentrations in body proteins, that eliminate the need for a Tyr tracer. To evaluate the accuracy of this approach, data were obtained from 12 type I diabetic patients and 24 nondiabetic control subjects who were studied with the full complement of tracers both with and without insulin infusion. Sp approximations closely matched measured values in both groups (mean difference <2%, all values <5%), but the agreement was poor for Qpt (error range = -8 to +43%) and Qt (error range -22 to +41%). Insulin status had no effect on these comparisons. The lower approximation error for Sp vs. Qpt is due to the small contribution ( approximately 10%) of Qpt to Qp. Approximation error for Qpt (r > 0.99) can be explained by variability in the ratio of Tyr to Phe coming from protein breakdown, (Qt - Qpt)/Qp. Ideally, all fluxes should be directly measured, but these data suggest that whole body Sp can be approximated with an acceptably small margin of error. However, the same equations do not yield reliably accurate values for Qpt or Qt.

Effects of precooling on thermoregulation during subsequent exercise.

PURPOSE: The purpose of this study was to examine the effect of a decreased body core temperature before a simulated portion of a triathlon (swim,15 min; bike, 45 min) and examine whether precooling could attenuate thermal strain and increase subjective exercise tolerance in a warm environment (26.6 degrees C/60% relative humidity (rh)). METHODS: Six endurance trained triathletes (28+/-2 yr, 8.2+/-1.7% body fat) completed two randomly assigned trials 1 wk apart. The precooling trial (PC) involved lowering body core temperature (-0.5 degrees C rectal temperature, Tre) in water before swimming. The control trial (CON) was identical except no precooling was performed. Water temperature and environmental conditions were maintained at 25.6 degrees C and 26.6 degrees C/60% rh, respectively, throughout all testing. RESULTS: Mean time to precool was 31+/-8 min and average time to reach baseline Tre during cycling was 9+/-7 min. Oxygen uptake (VO2), HR, skin temperature (Tsk), Tre, RPE, and thermal sensation (TS) were recorded following the swim segment and throughout cycling. No significant differences in mean body (Tb) or Tsk were noted between PC and CON, but a significant difference (P < 0.05) in Tre between treatments was noted through the early phases of cycling. No significant differences were reported in HR, VO2, RPE, TS, or sweat rate (SR) between treatments. Body heat storage (S) was negative following swimming in both PC (-92+/-6 W x m2) and CON (-66+/-9 W x m2). A greater S occurred in PC (109+/-6 W x m2) vs CON (79+/-4 W x m2) during cycling (P < 0.05). CONCLUSIONS: Precooling attenuated the rise in Tre, but this effect was transient. Therefore, precooling is not recommended before a triathlon under similar environmental conditions.

Excess postexercise oxygen consumption and recovery rate in trained and untrained subjects.

The purpose of this study was to determine whether aerobic fitness level would influence measurements of excess postexercise oxygen consumption (EPOC) and initial rate of recovery. Twelve trained [Tr; peak oxygen consumption (VO2 peak) = 53.3 +/- 6.4 ml . kg-1 . min-1] and ten untrained (UT; VO2 peak = 37.4 +/- 3.2 ml . kg-1 . min-1) subjects completed two 30-min cycle ergometer tests on separate days in the morning, after a 12-h fast and an abstinence from vigorous activity of 24 h. Baseline metabolic rate was established during the last 10 min of a 30-min seated preexercise rest period. Exercise workloads were manipulated so that they elicited the same relative, 70% VO2 peak (W70%), or the same absolute, 1.5 l/min oxygen uptake (VO2) (W1.5), intensity for all subjects, respectively. Recovery VO2, heart rate (HR), and respiratory exchange ratio (RER) were monitored in a seated position until baseline VO2 was reestablished. Under both exercise conditions, Tr had shorter EPOC duration (W70% = 40 +/- 15 min, W1.5 = 21 +/- 9 min) than UT (W70% = 50 +/- 14 min; W1.5 = 39 +/- 14 min), but EPOC magnitude (Tr: W70% = 3.2 +/- 1.0 liters O2, W1.5 = 1.5 +/- 0.6 liters O2; UT: W70% = 3.5 +/- 0.9 liters O2, W1.5 = 2.4 +/- 0.6 liters O2) was not different between groups. The similarity of Tr and UT EPOC accumulation in the W70% trial is attributed to the parallel decline in absolute VO2 during most of the initial recovery period. Tr subjects had faster relative decline during the fast-recovery phase, however, when a correction for their higher exercise VO2 was taken. Postexercise VO2 was lower for Tr group for nearly all of the W1.5 trial and particularly during the fast phase. Recovery HR kinetics were remarkably similar for both groups in W70%, but recovery was faster for Tr during W1.5. RER values were at or below baseline throughout much of the recovery period in both groups, with UT experiencing larger changes than Tr in both trials. These findings indicate that Tr individuals have faster regulation of postexercise metabolism when exercising at either the same relative or same absolute work rate.

Glycemic and insulinemic responses to multiple preexercise carbohydrate feedings.

This investigation was undertaken to determine whether consuming several small feedings of preexercise carbohydrate (CHO), rather than a single bolus, would affect blood glucose and insulin responses during rest and exercise. Eight trained cyclists ingested 22.5, 45, or 75 total g maltodextrin and dextrose dissolved in 473 ml of water or an equal volume of placebo (PL). Drinks were divided into four portions and consumed at 15-min intervals in the hour before a 120-min ride at 66% VO2max. Serum glucose values were elevated by the CHO feedings at rest and fell significantly below baseline and PL at 15 min of exercise. However, glucose concentrations were similar in each of the CHO trials. Insulin concentrations also increased rapidly during rest, then fell sharply at the onset of exercise. The findings demonstrate that CHO consumed within an hour before exercise, even when taken in several small feedings, can produce transient hypoglycemia near the onset of exercise. Additionally, the magnitude of the response appears to be unrelated to either the amount of CHO ingested or the insulin response.

Thermoregulatory responses to cycling with and without a helmet.

This study examined the effects of wearing a helmet on selected body temperatures and perceived heat sensation of the head and body while cycling in a hot-dry (D) (35 degrees C, 20% relative humidity (RH) and hot-humid (H) (35 degrees C, 70% RH) environment. Ten male and four female cyclists (mean +/- SD: males = age 27 +/- 7 yr, peak O2 uptake (VO2) 4.10 +/- 0.54 L.min-1; females = age 26 +/- 3 yr, peak O2 uptake (VO2) 3.08 +/- 0.49 L.min-1) performed four randomized 90-min cycling trials at 60% of peak VO2 both with (HE) and without (NH) a commercially available cycling helmet in both D and H environments. VO2, core (Te), skin (Tsk), and head skin temperatures, heart rate (HR), rating of perceived exertion (RPE), and perceived thermal sensation of head (TSH) and body (TSB) were measured throughout exercise. For all measured variables, no significant difference was evident between HE and NH. However, Tc, Tsk, and mean head skin temperatures were higher (P < 0.001) in H than D. Likewise, RPE, TSH, TSB (P < 0.001), and sweat rates (H = 1.33 +/- 0.32, D = 1.14 +/- 0.23 L.h-1) (P < 0.01) were higher in H versus D. Results indicate that use of a commercially available cycling helmet while riding in a hot-dry or hot-humid environment does not cause the subjects to become more hyperthermic or increase perceived heat sensation of the head or body.

The effect of upper body exercise intensity and duration on post-exercise oxygen consumption.

This study assessed excess post-exercise oxygen consumption (EPOC) following upper body exercise (UBE) of different intensity and duration. Ten subjects, 5 male and 5 female (age: 26.7 +/- 4.9 yr; peak UBE oxygen uptake [VO2peak]: 1.78 +/- 0.57 l. min-1, 25.6 +/- 5.8 ml.kg-1. min-1) performed three randomized tests on an arm crank ergometer: 1) low intensity, short duration (LS) = 35% VO2peak for 15 min; 2) low intensity, long duration (LL) = 35% VO2peak for 30 min; 3) high intensity, short duration (HS) = 70% VO2peak for 15 min. Subjects reported for all tests in the morning and in a fasted and rested state. Exercise was preceded by a 30 min seated baseline. Recovery VO2 was continuously monitored until baseline was re-established. EPOC duration (p < 0.01) and magnitude (p < 0.01) were significantly greater following HS, while LL and LS did not differ in response (duration and magnitude: HS = 14.0 +/- 6.5 min, 32.5 +/- 24.6 kJ; LL = 5.5 +/- 4.4 min, 12.3 +/- 8.6 kJ; LS = 5.7 +/- 4.9 min, 10.3 +/- 5.3 kJ). HS also had higher HR (73 +/- 10 b.min-1, p < 0.01) at end-EPOC compared to LL (64 +/- 8 b.min-1) and LS (66 +/- 8 b.min-1), and baseline HS values (63 +/- 8 b. min-1). Results from this study indicate that UBE intensity has a greater effect on EPOC than exercise duration under these conditions. UBE appears to have similar EPOC patterns as lower body exercise.