Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a 5% (MI-CT) overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01), as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01) and lipoprotein lipase (LPL; P < 0.05). Lactate dehydrogenase also presented a higher maximal activity (nmol·min-1·mg protein-1) in HI-IT (around 83%). We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a "time-efficient" strategy inducing metabolic adaptation.

Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats.

We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a 5% (MI-CT) overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01), as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01) and lipoprotein lipase (LPL; P < 0.05). Lactate dehydrogenase also presented a higher maximal activity (nmol·min(-1)·mg protein(-1)) in HI-IT (around 83%). We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a "time-efficient" strategy inducing metabolic adaptation.

Exercise training decreases adipose tissue inflammation in cachectic rats.

Bearing in mind that cancer cachexia is associated with chronic systemic inflammation and that endurance training has been adopted as a nonpharmacological anti-inflammatory strategy, we examined the effect of 8 weeks of moderate intensity exercise upon the balance of anti- and pro-inflammatory cytokines in 2 different depots of white adipose tissue in cachectic tumour-bearing (Walker-256 carcinosarcoma) rats. Animals were assigned to a sedentary control (SC), sedentary tumour-bearing (ST), sedentary pair-fed (SPF) or exercise control (EC), exercise tumour-bearing (ET), and exercise pair-fed (EPF) group. Trained rats ran on a treadmill (60% VO(2)max) 60 min/day, 5 days/week, for 8 weeks. The retroperitoneal (RPAT) and mesenteric (MEAT) adipose pads were excised and the mRNA (RT-PCR) and protein (ELISA) expression of IL-1ß, IL-6, TNF-α, and IL-10 were evaluated. The number of infiltrating monocytes in the adipose tissue was increased in cachectic rats. TNF-α mRNA in MEAT was increased in the cachectic animals (p<0.05) in relation to SC. RPAT protein expression of all studied cytokines was increased in cachectic animals in relation to SC and SPF (p<0.05). In this pad, IL-10/TNF-α ratio was reduced in the cachectic animals in comparison with SC (p<0.05) indicating inflammation. Exercise training improved IL-10/TNF-α ratio and induced a reduction of the infiltrating monocytes both in MEAT and RPAT (p<0.05), when compared with ST. We conclude that cachexia is associated with inflammation of white adipose tissue and that exercise training prevents this effect in the MEAT, and partially in RPAT.

Endurance training restores peritoneal macrophage function in post-MI congestive heart failure rats.

Congestive heart failure (CHF) induces a state of immune activation, and peritoneal macrophages (M phi s) may play an important role in the development and progression of one such condition. Moderate endurance training modulates peritoneal M phi function. We evaluated the effect of endurance training on different stages of the phagocytic process and in the production of interleukin-6 (IL-6), interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) after LPS stimulation. Either ligation of the left coronary artery or Sham operations were performed in adult Wistar rats. After 4 wk, control (Sham operated) and MI (ligation of the left coronary artery) animals were randomly assigned to either a sedentary (Sham-operated sedentary, n = 7 and MI sedentary, n = 10) or a trained group (Sham-operated trained, n = 8 and MI trained, n = 8). Trained rats ran on a treadmill (0% grade at 13-20 m/min) for 60 min/day, 5 days/wk, for 8-10 wk, whereas sedentary rats had only limited activity. Training increased maximal oxygen uptake normalized for body weight (ml.kg(-1).min(-1)), as well as skeletal muscle citrate synthase maximal activity, when compared with sedentary groups. The resident and total cell number, the chemotaxis index, and the production of TNF-alpha stimulated by LPS were significantly higher in the MI sedentary group when compared with the Sham sedentary group. Moderate endurance training reversed these alterations promoted by post-MI. These results demonstrate that moderate intensity exercise training modulates peritoneal M phi function and induces beneficial metabolic effects in rats with post-MI CHF.

Changes in the pro-inflammatory cytokine production and peritoneal macrophage function in rats with chronic heart failure.

Chronic heart failure (CHF) is a state of immune activation, and pro-inflammatory cytokines play an important role in its development and progression. Macrophages (Mphis), when activated, are the main source of pro-inflammatory cytokines. We measured interleukin-6 (IL-6), interleukin (IL-1beta) and tumor necrosis factor - alpha (TNF-alpha) production after lipopolysaccharide (LPS)-stimulation, as well as peritoneal Mphis migration, phagocytic capacity, chemotaxis index, and hydrogen peroxide production, in an attempt to clarify the role of this cell in an animal model of CHF. Ligature of the left coronary artery or sham operation was performed in adult Wistar rats. After 12 weeks, resident and total cell number, phagocytic capacity, chemotaxis index, and hydrogen peroxide production in Mphis were significantly higher in CHF than in control rats. The production of IL-6 and TNF- alpha was similarly significantly enhanced in CHF as compared with controls. Mphis obtained from CHF rats were more responsive to LPS, suggesting the existence, in vivo, of possible factor(s) modulating the production of pro-inflammatory cytokines. The results demonstrated that there is modification of peritoneal Mphis function along CHF development, possibly contributing to the pathophysiological process in the establishment of CHF.

Heterogeneous response of adipose tissue to cancer cachexia

Cancer cachexia causes disruption of lipid metabolism. Since it has been well established that the various adipose tissue depots demonstrate different responses to stimuli, we assessed the effect of cachexia on some biochemical and morphological parameters of adipocytes obtained from the mesenteric (MES), retroperitoneal (RPAT), and epididymal (EAT) adipose tissues of rats bearing Walker 256 carcinosarcoma, compared with controls. Relative weight and total fat content of tissues did not differ between tumor-bearing rats and controls, but fatty acid composition was modified by cachexia. Adipocyte dimensions were increased in MES and RPAT from tumor-bearing rats, but not in EAT, in relation to control. Ultrastructural alterations were observed in the adipocytes of tumor-bearing rat RPAT (membrane projections) and EAT (nuclear bodies)

Heterogeneous response of adipose tissue to cancer cachexia.

Cancer cachexia causes disruption of lipid metabolism. Since it has been well established that the various adipose tissue depots demonstrate different responses to stimuli, we assessed the effect of cachexia on some biochemical and morphological parameters of adipocytes obtained from the mesenteric (MES), retroperitoneal (RPAT), and epididymal (EAT) adipose tissues of rats bearing Walker 256 carcinosarcoma, compared with controls. Relative weight and total fat content of tissues did not differ between tumor-bearing rats and controls, but fatty acid composition was modified by cachexia. Adipocyte dimensions were increased in MES and RPAT from tumor-bearing rats, but not in EAT, in relation to control. Ultrastructural alterations were observed in the adipocytes of tumor-bearing rat RPAT (membrane projections) and EAT (nuclear bodies).

Effect of a moderate intensity exercise training protocol on the metabolism of macrophages and lymphocytes of tumour-bearing rats.

It is commonly accepted that moderate intensity exercise is beneficial to the immune system. We tested the influence of a moderate intensity training protocol (8 weeks) upon immune system function in Wistar tumour-bearing (TB) rats. The metabolism of glucose and glutamine in lymphocytes and macrophages was assessed, together with some functional parameters (hydrogen peroxide production and lymphocyte proliferative response). These substrates were chosen since they represent the most important energetic and synthetic metabolites for these cellular types. The training protocol caused a decrease of 17.4 per cent in the production of H(2)O(2) by macrophages, as well as a decrease in glucose consumption (25 per cent) and lactate production (47.1 per cent), and an increase in the production of labelled CO(2) from the oxidation of [U-(14)C]-glucose, in TB rats. The training protocol was also able to induce changes in the maximal activity of some key enzymes in the metabolism of glucose and glutamine, a reduction of hexokinase (68.8 per cent) activity and an increase in the activity of citrate synthase (10.1 per cent) in TB rats. The training protocol increased the proliferative response of lymphocytes cultivated in the absence of mitogens (75 per cent), of those cultivated in the presence of ConA (38.2 per cent) and in the presence of LPS (45.0 per cent). These cells also showed an increase in the maximal activity of some key enzymes of the glycolytic and glutaminolytic pathways. Our data demonstrated that the training protocol was able to induce an increase in aerobic utilisation of both substrates in lymphocytes and macrophages. The training protocol was also able to prevent several changes in glucose and glutamine metabolism that are normally present in sedentary TB rats. These changes in immune cell metabolism induced by the training protocol were able to increase TB rat survival.

The effect of adrenaline and Walker-256 tumour-induced cachexia upon Kupffer cell metabolism.

Kupffer cells (KC), the liver macrophages, are able to produce PGE(2), which is involved in immune suppression and in the aggravation of cancer cachexia due to interference with lipid metabolism in the liver. Since tumour-bearing (TB) rats present high plasma epinephrine levels, and this hormone is able to affect macrophage metabolism and function, we have assessed the effect of epinephrine (5 nM) upon Kupffer cell PGE(2) production. Epinephrine induced increased production of PGE(2) both by control (3.5-fold) and TB rats (27 per cent) KC, an effect blocked by propranolol. Enhancement of cAMP content in the cells by addition of isoproterenol (0.1 microM) to the incubations, however, failed to induce the same response in the cells. Nevertheless, when phenylephrine (1 microM) was added to the incubation, a similar pattern of PGE(2) production to that observed for epinephrine was found for control and TB rat KC. We propose that the effect of epinephrine upon KC PGE(2) production is mediated by alpha-adrenergic receptors and that Ca(2+) is involved in the response, since increasing concentrations of the ion added to the incubation medium (0.25, 0.5 and 1.0 mM) enhanced the eicosanoid production, while EDTA abolished the response.

The effect of Walker-256 tumour development upon Kupffer cell metabolism.

The liver plays a central role in the establishment and maintenance of the cachectic state in rats bearing extra-hepatic tumours. Kupffer cells, which as macrophages, show a strong relationship between metabolism and function could be involved in the alterations observed in the disruption of many functions of the organ as a whole. To assess whether the metabolic/functional pattern of Kupffer cells was altered by cachexia we have investigated the utilization of glucose, glutamine and palmitate by the cells from tumour-bearing and control rats. We have found an enhanced utilization of the three substrates by the cells from tumour-bearing rats as compared with controls, which was related to greater energy production through the Krebs cycle and enhanced production of precursors for the synthesis of the many substances the cells secrete when activated. The use of palmitate as substrate was also augmented in these cells, in the opposition to the observation in stimulated peritoneal macrophages. The availability of palmitate however, was not associated with a reduction of glucose or glutamine consumption. The cycle of interconversion, free fatty acids/triacyglycerol in Kupffer cells from tumour-bearing rats was also found to be increased, as was hydrogen peroxide production. Taken together the results suggest an increased utilization of substrates for both energy production and for synthetic processes (e.g. NADPH for hydrogen peroxide production).

Carnitine palmitoyltransferase II activity is decreased in liver mitochondria of cachectic rats bearing the Walker 256 carcinosarcoma: effect of indomethacin treatment.

The syndrome of cancer cachexia is accompanied by several alterations of lipid metabolism, especially that in the liver. In this study we have investigated a possible mechanism whereby the presence of the Walker 256 carcinosarcoma affects hepatic fatty acid oxidative capacity in tumour-bearing rats. Hepatic mitochondrial outer membrane carnitine palmitoyltransferase I (CPT I), generally accepted as the main site of regulation of fatty acid oxidation, was unaffected by the presence of the extra-hepatic tumour. However, mitochondrial inner-membrane carnitine palmitoyltransferase II (CPT II) activity was markedly decreased in mitochondria isolated from the liver of tumour-bearing rats. Immuno-detection by Western blotting using a CPT II-specific antibody identified two bands (corresponding to M(r) 69,000 and 54,000) in tumour-bearing rats whereas only the normal-sized CPT II was present (at the expected M(r) 69,000) in mitochondria from control rats. It is suggested that the emergence of the second, smaller protein may represent a catalytically less active protein that arises in vivo, since its appearance was not affected by the inclusion of proteolysis inhibitors in the mitochondrial preparation buffers. Treatment of the tumour-bearing rats with indomethacin, a prostaglandin (including PGE2) synthesis inhibitor, increased CPT II activity to levels even higher than those found in the control animals. It is suggested that PGE2 may play a role in the control of CPT II expression in the liver of tumour-bearing rats. Indomethacin treatment did not affect either of the two CPT activities of the mitochondria isolated from tumour tissue.

Decreased secretion of very-low-density lipoprotein triacylglycerol and apolipoprotein B is associated with decreased intracellular triacylglycerol lipolysis in hepatocytes derived from rats fed orotic acid or n-3 fatty acids.

Hepatocytes from rats fed a chow (control) diet or from rats fed a chow diet supplemented with either orotic acid (OA; 1%, w/w) or fish oil (FO; 20%, v/w) were maintained in culture for periods up to 48 h. during the first 24 h period, the low rates of output of very-low-density lipoprotein (VLDL)-associated triacylglycerol (TAG) and apolipoprotein B (apoB) in hepatocytes from the FO- and OA-fed animals were associated with significantly lower rates of intracellular TAG lipolysis and re-esterification. Most of the VLDL TAG secreted was mobilized via lipolysis of the intracellular TAG pool, but the proportion of VLDL TAG secreted via this route in cells from the FO-fed and OA-fed animals was decreased compared with that in the control-fed animals' cells. In the presence of exogenous oleate the inhibitory effect of OA feeding on VLDL apoB and TAG secretion persisted in the derived hepatocytes for up to 48 h following isolation. However, when oleate was absent no inhibitory effect on the secretion of TAG and apoB was observed between 24 and 48 h. Under these conditions the rate of intracellular TAG turnover returned to normal. The initial inhibitory effect of FO feeding on VLDL TAG and apoB secretion did not persist in the derived hepatocytes between 24 h and 48 h of culture in the presence of exogenous oleate. Although intracellular TAG lipolysis and VLDL TAG and apoB secretion rates appear to be positively correlated, a causal relationship has not been conclusively established.

Studies on the lipid metabolism of Walker 256 tumour-bearing rats during the development of cancer cachexia.

Aspects of the lipid metabolism of Walker 256 carcinosarcoma-bearing cachectic rats (TB) were investigated during a 14 day interval of tumour growth. Food intake and body weight of the TB rats were reduced by 18% and 13%, respectively, on day 14, as compared with non-tumour-bearing animals. The tumour burden then, corresponded to 19% of total body weight. The total fat content was not different in the liver, heart, carcass, epididymal (EAT) and retroperitoneal (RPAT) adipose tissues of the two groups. The brown adipose tissue (BAT) and skeletal muscle (gastrocnemius-SM) of the TB rats had increased levels of fat (23% and 200%, respectively). Enteral absorption of 14C-triolein was decreased in the TB rats, but the liver, heart, and SM of these animals incorporated more radiolabelled lipid than the control animals, while the adipose tissues exhibited a decreased incorporation of radioactivity in relation to controls. More lipid was incorporated into the VLDL fraction secreted by the liver of TB rats, which exhibited a different distribution of the incorporated 14C-oleate in the various lipid subfractions. Ultrastructural studies showed that the hepatocytes of the TB rats had a greater incidence of lipid droplets in the cytoplasm.

Fatty acid oxidation in lymphocytes from Walker 256 tumor-bearing rats.

The oxidation of fatty acids in lymphocytes from the mesenteric lymph nodes of Walker 256 carcinosarcoma-bearing rats (TB) was studied, as well as the activity of the mitochondrial long-chain fatty acid transport system. Two-month old Wistar rats were subcutaneously implanted with 10(7) cells and after 2 weeks the tumor mass was 15-20% of the carcass weight. The activity of carnitine palmitoyltransferase (CPT) II was demonstrable in the lymphocytes of the TB group (8.2 +/- 5.6 nmol/min per mg mitochondria protein for 15 rats) and was not detected in the control, while that of CPT I was only slightly increased in the former. Similar rates of [1-14C]-palmitate decarboxylation were found for TB and control rat lymphocytes. However, when the rate of decarboxylation of [1-14C]-palmitate present in the intracellular pool of lipids was investigated in cultured lymphocytes, the cells of TB rats exhibited rates 17-fold higher than those of control animals in the presence of fetal calf serum (FCS). Decarboxylation in the presence of TB rat serum was 178-fold higher than obtained with normal rat serum, and 1.4-fold compared to FCS. These results suggest that, during cachexia, lymphocytes preferably oxidize intracellular lipids, and that this capacity is greatly enhanced by factors circulating in the serum of tumor-bearing rats.

Fatty acid oxidation in lymphocytes from Walker 256 tumor-bearing rats

The oxidation of fatty acids in lymphocytes from the mesenteric lymph nodes of Walker 256 carcinosarcoma-bearing rats (TB) was studied, as well as the activity of the mitochondrial long-chain fatty acid transport system. Two-month old Wistar rats were subcutaneously implanted with 10(7) cells and after 2 weeks the tumor mass was 15-20 per cent of the carcass weight. The activity of camitine palmitoyltransferase (CPT) II was demonstrable in the lymphocytes of the TB group (8.2 ñ 5.6 nmol/min per mg mitochondria protein for 15 rats) and was not detected in the control, while that of CPT I was only slightly increased in the former. Similar rates of [1-14C]-palmitate decarboxylation were found for TB and control rat lymphocytes. However, when the rate of decarboxylation of [1-14C]-paimitate present in the intracellular pool of lipids was investigated in cultured lymphocytes, the cells of TB rats exhibited rates 17-fold higher than those of control animals in the presence of fetal calf serum (FCS). Decarboxylation in the presence of TB rat serum was 178-fold higher than obtained with normal rat serum, and 1.4-fold compared to FCS. These results suggest that, during cachexia, lymphocytes preferably oxidize intracellular lipids, and that this capacity is greatly enhanced by factors circulating in the serum of tumor-bearing rats.

Insulin and prostaglandins as signals between tumor and the host immune system.

The development of malignant tissue in vivo is partially favored by the immunosuppression that occurs in cancer patients. However, the signals between tumor and immune tissues remain to be identified. We present evidence that prostaglandins may act as one of these signals by a direct action on cells of the immune system, or by inhibition of insulin secretion which in turn suppresses immune function, or both.

Insulin and prostaglandins as signals between tumor and the host immune system

The development of malignant tissue in vivo is partially favored by the immunosuppression that occurs in cancer patients. However, the signals between tumor and immune tissues remain to be identified. We present evidence that prostaglandins may act as one of these signals by a direct action on cells of the immune system, or by inhibition of insulin secretion which in turn suppresses immune function, or both

Lipid balance in HepG2 cells: active synthesis and impaired mobilization.

The human hepatoma cell line HepG2 in culture medium synthesized fatty acids de novo (144 +/- 9 nmol fatty acid/mg protein per 24 h) at a rate similar to that observed in freshly prepared rat hepatocytes (192 +/- 8 nmol/mg per 24 h) and in primary cultures of rat hepatocytes (165.4 +/- 29.3 nmol/mg per 24 h). In HepG2 cells, fatty acid synthesis was inhibited by extracellular oleate (0.75 mM), and, to a lesser extent, by glucagon (10(-7) M). Insulin (7.8 x 10(-8) M) had a mild stimulatory effect. Fatty acid synthesis was not influenced by lipogenic precursors (lactate plus pyruvate), substances which, in rat hepatocytes, had pronounced stimulatory effects. Fatty acid synthesis rates were also unchanged in the presence of prostaglandin E2 (PGE2). In general, compared to rat hepatocytes, fatty acid synthesis in HepG2 cells was less sensitive to manipulation of the culture medium in vitro. HepG2 cells had a high capacity for triacylglycerol synthesis from extracellular oleate (469 +/- 43 nmol triacylglycerol/mg protein per 24 h) but phospholipid synthesis was relatively low (15.8 +/- 0.4% of total glycerolipids). Very little of the above newly synthesized triacylglycerol was secreted as lipoprotein (4.62 +/- 0.88 nmol triacylglycerol/mg protein per 24 h) resulting in a large intracellular accumulation of triacylglycerol. This was exacerbated by the absence of any detectable ketogenesis. The secretion of triacylglycerol produced from de novo synthesized fatty acids was also very low in HepG2 compared to that observed in primary cultures of rat hapatocytes.(ABSTRACT TRUNCATED AT 250 WORDS)