I. Mark Olfert

Vascular endothelial growth factor (VEGF) is required for vasculogenesis and angiogenesis during embryonic and early postnatal life. However the organ‐specific functional role of VEGF in adult life, particularly in skeletal muscle, is less clear. To explore this issue, we engineered skeletal muscle‐targeted VEGF deficient mice (mVEGF−/−) by crossbreeding mice that selectively express Cre recombinase in skeletal muscle under the control of the muscle creatine kinase promoter (MCKcre mice) with mice having a floxed VEGF gene (VEGFLoxP mice). We hypothesized that VEGF is necessary for regulating both cardiac and skeletal muscle capillarity, and that a reduced number of VEGF‐dependent muscle capillaries would limit aerobic exercise capacity. In adult mVEGF−/− mice, VEGF protein levels were reduced by 90 and 80% in skeletal muscle (gastrocnemius) and cardiac muscle, respectively, compared to control mice (P < 0.01). This was accompanied by a 48% (P < 0.05) and 39% (P < 0.05) decreases in the capillary‐to‐fibre ratio and capillary density, respectively, in the gastrocnemius and a 61% decrease in cardiac muscle capillary density (P < 0.05). Hindlimb muscle oxidative (citrate synthase, 21%; β‐HAD, 32%) and glycolytic (PFK, 18%) regulatory enzymes were also increased in mVEGF−/− mice. However, this limited adaptation to reduced muscle VEGF was insufficient to maintain aerobic exercise capacity, and maximal running speed and endurance running capacity were reduced by 34% and 81%, respectively, in mVEGF−/− mice compared to control mice (P < 0.05). Moreover, basal and dobutamine‐stimulated cardiac function, measured by transthoracic echocardiography and left ventricular micromanomtery, showed only a minimal reduction of contractility (peak +dP/dt) and relaxation (peak –dP/dt, τ_E). Collectively these data suggests adequate locomotor muscle capillary number is important for achieving full exercise capacity. Furthermore, VEGF is essential in regulating postnatal muscle capillarity, and that adult mice, deficient in cardiac and skeletal muscle VEGF, exhibit a major intolerance to aerobic exercise.

Decreased skeletal muscle mass and proportion of fast-twitch glycolytic fibers are well-documented correlates of aging; however, data on concomitant changes on capillary-to-fiber ratio (C:F) are inconsistent. We simultaneously examined fiber-type composition and arteriolar and venular portions of capillaries in the distal hind-limb muscles of 12-, 24-, and 35-month old F1 hybrid F344 Brown Norway rats. Aging significantly increased C:F of venular capillaries in muscles, which also presented significant age-related increase in slow-(type I) and fast-(type IIa) oxidative fibers (plantaris, tibialis anterior, medial gastrocnemius; p <.05). In contrast, arteriolar and venular capillary proportions did not change in the soleus, extensor digitorum longus, or lateral gastrocnemius. These data suggest that age-associated increases in skeletal muscle capillarity may be due to the venular portion of capillaries and that the increase occurs primarily in muscles that demonstrate increased oxidative potential with age.

Thrombospondin-1 (TSP-1) is believed to be an endogenous angiogenic inhibitor. In this study, we report that a single 1 h bout of treadmill running increases TSP-1 mRNA 3–4-fold (p < 0.001). Interestingly, with short-term training (up to 5 days, 1 h/day) the acute response of TSP-1 mRNA to exercise was ablated after 3 days. Following long-term training (8 weeks, 1 h/day, 5 d/wk), in either normoxia or chronic hypoxia, the TSP-1 mRNA response to an acute bout of exercise was restored and increased 3–4-fold (p < 0.01). However, chronic exposure to hypoxia (8-weeks) decreases both the basal and acute exercise-induced TSP-1 mRNA levels by 44 and 48%, respectively (p < 0.05). Based on the robust TSP-1 gene response to a single acute exercise bout, its temporal response to repetitive exercise bouts, and the putative role of TSP-1 in the angiogenic process, we speculate that TSP-1 may play a role in regulating the onset of skeletal muscle angiogenesis in response to exercise.

The phase III slope of sulfur hexafluoride (SF₆) in a single-breath washout (SBW) is greater than that of helium (He) under normal gravity (i.e., 1G), thus resulting in a positive SF₆-He slope difference. In microgravity (μG), SF₆-He slope difference is smaller because of a greater fall in the phase III slope of SF₆than He. We sought to determine whether increasing thoracic fluid volume using 60° head-down tilt (HDT) in 1G would produce a similar effect to μG on phase III slopes of SF₆and He. Single-breath vital capacity (SBW) and multiple-breath washout (MBW) tests were performed before, during, and 60 min after 1 h of HDT. Compared with baseline (SF₆1.050 ± 0.182%/l, He 0.670 ± 0.172%/l), the SBW phase III slopes for both SF₆and He tended to decrease during HDT, reaching nadir at 30 min (SF₆0.609 ± 0.211%/l, He 0.248 ± 0.138%/l; P = 0.08 and P = 0.06, respectively). In contrast to μG, the magnitude of the phase III slope decrease was similar for both SF₆and He; therefore, no change in SF₆-He slope difference was observed. MBW analysis revealed a decrease in normalized phase III slopes at all time points during HDT, for both SF₆( P < 0.01) and He ( P < 0.01). This decrease was due to changes in the acinar, and not the conductive, component of the normalized phase III slope. These findings support the notion that changes in thoracic fluid volume alter ventilation distribution in the lung periphery but also demonstrate that the effect during HDT does not wholly mimic that observed in μG.

We characterized O₂ consumption (V̇_{O2) during treadmill exercise in 12-, 24-, and 35-month-old Fischer 344 × Brown Norway F1 hybrid (F344BNF1) rats. When accounting for differences in body mass (Mb), V̇O2peak decreased by 10% and 33% in 24- and 35-month-old rats, respectively, compared with rats at 12 months (analysis of covariance, p <.01). O2 cost per unit work at V̇O2peak (i.e., V̇O2peak/work) was greater in 35-month-old rats compared with 12- and 24-month-old rats (p <.001). During submaximal exercise, the O2 cost was greater in 24- and 35-month-old than 12-month-old rats (p <.01). Analysis of covariance revealed similar patterns irrespective of differences in Mb or lean Mb as covariates. The underlying mechanism responsible for increasing O2 consumption in aged F344BNF1 rats during exercise, although partly explained by mechanical inefficiencies of locomotion, still remains to be determined.}

Women may experience greater pulmonary gas exchange impairment during exercise than men. To test this we used the multiple inert gas elimination technique to study eight women and seven men matched for age, height and V̇_{O2 max} (∼48 ml kg⁻¹ min⁻¹) during normoxic and hypoxic (inspired P_O2= 95 Torr) cycle exercise. Resting lung function was similar between the sexes, except for a lower carbon monoxide diffusing capacity (DL_CO) in women (P < 0.05). Arterial P_O2,P_CO2 and alveolar–arterial O₂ difference (A−aD_O2) were not significantly different in men and women. Despite a lower diffusing capacity for O₂ (DL_O2) in women, the ratio DL_O2/βQ̇ (which estimates pulmonary end‐capillary diffusion equilibrium) was similar between men and women and estimates of diffusion limitation during hypoxic exercise were not different between the sexes. Ventilation–perfusion inequality (described by the second moment of the perfusion distribution, logSD) increased during both normoxic and hypoxic exercise. Surprisingly, logSD values were slightly lower for women under all conditions (P < 0.05), but this did not significantly affect gas exchange. These data indicate that these active women, despite a lower DL_CO and DL_O2, do not experience greater exercise‐induced abnormalities in gas exchange than men matched for age, height, aerobic capacity and lung size. Possibly fitness level and lung size are more important in determining whether or not pulmonary gas exchange impairment occurs during exercise than sex per se.

Gene expression of vascular endothelial growth factor (VEGF), and to a lesser extent of transforming growth factor-β₁ (TGF-β₁) and basic fibroblast growth factor (bFGF), has been found to increase in rat skeletal muscle after a single exercise bout. In addition, acute hypoxia augments the VEGF mRNA response to exercise, which suggests that, if VEGF is important in muscle angiogenesis, hypoxic training might produce greater capillary growth than normoxic training. Therefore, we examined the effects of exercise training (treadmill running at the same absolute intensity) in normoxia and hypoxia (inspired O₂ fraction = 0.12) on rat skeletal muscle capillarity and on resting and postexercise gene expression of VEGF, its major receptors (flt-1 and flk-1), TGF-β₁, and bFGF. Normoxic training did not alter basal or exercise-induced VEGF mRNA levels but produced a modest twofold increase in bFGF mRNA ( P < 0.05). Rats trained in hypoxia exhibited an attenuated VEGF mRNA response to exercise (1.8-fold compared 3.4-fold with normoxic training; P< 0.05), absent TGF-β₁ and flt-1 mRNA responses to exercise, and an approximately threefold ( P < 0.05) decrease in bFGF mRNA levels. flk-1 mRNA levels were not significantly altered by either normoxic or hypoxic training. An increase in skeletal muscle capillarity was observed only in hypoxically trained rats. These data show that, whereas training in hypoxia potentiates the adaptive angiogenic response of skeletal muscle to a given absolute intensity of exercise, this was not evident in the gene expression of VEGF or its receptors when assessed at the end of training.