Liver and muscle proteolytic activity in male growing rats fed a field bean (Vicia faba L.) diet

Total liver cathepsin A and D and gastrocnemius acid proteolytic activities have been evaluated in male growing rats fed ad libitum over periods of 15 and 30 days on 20%-protein diets containing either casein or raw field bean (Vicia faba L.) as protein sources. It has been found that, compared to the control casein-fed rats, those fed the legume diet exhibited a marked reduction (p less than 0.05) in the rate of growth and protein efficiency rate; liver proteolytic activity increased with ageing in the two dietary treatments and was found to be significantly higher (p less than 0.05) in the legume-fed rats. However, muscle proteolytic activity decreased with ageing and proved significantly increased (p less than 0.05) in the legume-fed rats.     

Liver proteolytic activity in growing male chickens fed a raw field bean (Vicia faba L.) diet

As compared to well-nourished chickens, a significant reduction (p less than 0.05) in the rate of growth, along with a significant increase (p less than 0.05) in liver activities of both cathepsin A and D has been found in growing male chickens fed ad libitum over a period of 60 days on a diet containing raw field bean (Vicia faba L.) as the main source of protein. Moreover, addition of methionine (250 mg/100 g diet) to the legume diet had no significant effect in restoring either rate of growth or liver proteolytic activity. The possible nature of these findings are briefly discussed.     

Cathepsin B, D and H activities in muscles of chicks of fast and slow growing strains: effect of age and diet

1. Activities of cathepsins B, D and H were measured in leg and breast muscles of fast growing (broiler) and slow growing (layer) chicks at eight time intervals between 1 and 29 days of age. 2. These enzyme activities were also measured in muscles from fast and slow growing chicks given a low protein (125 g/kg crude protein) diet between the ages of 17 and 24 days. 3. Activities of none of these cathepsins differed greatly between muscle type or strain of chick. However in both strains of chick cathepsin D and H in muscles significantly decreased with increasing age (muscle size) of the chick. Cathepsin D activity also increased when muscle proteolytic rates were increased by feeding a low protein diet. This latter effect was significant only in the muscles of fast growing chicks. 4. The results suggest that lysosomal proteases are not responsible for the differences in muscle protein degradation and growth between fast and slow growing strains of chicks, or between muscle types in the chick.     

Effect of corticosterone and protein malnutrition on muscle protein breakdown in vivo in rats as measured by the urinary excretion of 3-methylhistidine

The role of corticosterone in regulating the rate of muscle protein breakdown was evaluated by measuring the urinary excretion of 3-methylhistidine (3-Mehis) during the administration of 0.0 (vehicle), 0.8 (physiological dose) and 10 (pharmacological dose) mg of the glucocorticoid/100 g body weight/day to adrenalectomized rats (AdX, AdX 0.8 and AdX 10 respectively). A fourth group of intact rats receiving only vehicle (In) was included as control. Rats were fed on either adequate protein and energy (Co) or low-protein (1-P) diets, for eight consecutive days. No differences were found between AdX and AdX 0.8 groups as compared to the In group in regard to body and liver weights. The AdX 10 group exhibited a significant reduction in body weight and a considerable increase in liver weight; these results were found in rats fed on the Co and 1-P diets, although rats on the 1-P diet showed a proportional decrease in those parameters as compared to the rats fed on the Co diet. Gastrocnemius, tibialis and E.D.L. muscle weights were significantly reduced in AdX 10 group, approximatley at the same extent in the two dietary groups. Soleus muscle weight increased in the AdX 10 group, at the same extent in the two dietary groups, as compared to the In group. Plasma corticosterone levels were significantly greater in the AdX 10 group in both dietary treatments, though restriction of protein in the diet induced a higher plasma hormone level than that of the Co group. Urea-N and creatinine outputs were significantly higher in the AdX 10 group. 3-Mehis excretion underwent an immediate and significant rise in the AdX 10 group, although rats fed on 1-P diet showed a more persistent rise than those fed on the Co diet. No differences were found among the other groups. It is concluded that high plasma corticosterone levels can accelerate muscle protein breakdown and that this action is not seriously affected by the protein content of the diet.     

Excessive Energy Intake Does Not Modify Fed‐state Tissue Protein Synthesis Rates in Adult Rats

The impact of chronic excessive energy intake on protein metabolism is still controversial. Male Wistar rats were fed ad libitum during 5 weeks with either a high‐fat high‐sucrose diet (HF: n = 9) containing 45% of total energy as lipids (protein 14%; carbohydrate 40% with 83.5% sucrose) or a standard diet (controls: n = 10). Energy intake and body weight were recorded. At the end of the experiment, we measured body composition, metabolic parameters (plasma amino acid, lipid, insulin, and glucose levels), inflammatory parameter (plasma α2‐macroglobulin), oxidative stress parameters (antioxidant enzyme activities, lipoperoxidation (LPO), protein carbonyl content in liver and muscle), and in vivo fed–state fractional protein synthesis rates (FSRs) in muscle and liver. Energy intake was significantly higher in HF compared with control rats (+28%). There were significant increases in body weight (+8%), body fat (+21%), renal (+41%), and epidydimal (+28%) fat pads in HF compared with control rats. No effect was observed in other tissue weights (liver, muscle, spleen, kidneys, intestine). Liver and muscle FSRs, plasma levels of lipids, glucose, insulin and α2‐macroglobulin, soleus and liver glutathione reductase and peroxidase acitivities, MnSOD activity, LPO, and protein carbonyl content were not altered by the HF diet. Only soleus muscle and liver Cu/ZnSOD activity and soleus muscle catalase activities were reduced in HF rats compared with control rats. Thus, chronic excessive energy intake and increased adiposity, in the absence of other metabolic alterations, do not stimulate fed‐state tissue protein synthesis rates.     

The isoelectric fractionation of hen''s-egg ovotransferrin

1. ATP sulphurylase was assayed in various organs from vitamin A-deficient and pair-fed control rats at different stages of deficiency. Activity decreased slightly in the liver and markedly in the adrenal gland. Striking differences in liver activity were observed between pair-fed control and ad libitum-fed animals. This observation suggested that diet (apart from vitamin A) strongly influenced the activity of ATP sulphurylase. 2. Total starvation caused a severe decrease in activity in liver within 48hr. This was due to a lack of protein intake. 3. By feeding groups of vitamin A-deficient and pair-fed control rats on a diet containing 80% protein, the specific activity of the liver ATP sulphurylase was maintained in the pair-fed control group at the normal level of an ad libitum-fed rat, whereas it decreased by 25% (statistically significant at P<0.01) in the deficient rat. On a 20%-protein diet, there were no significant differences between vitamin A-deficient and pair-fed control rats. These relationships held also for enzyme activity expressed per g. of liver, per total liver and per g. of DNA. There were no differences in liver protein or DNA concentration between vitamin A-deficient and control rats on either protein intake. 4. Control rats on a 20%-protein diet had liver specific enzyme activities about one-half of those in control rats on an 80%-protein diet, as well as lower liver protein concentrations. 5. It is concluded that, when the effect of protein deprivation on ATP sulphurylase is separated from the effect of vitamin A deficiency, a lowering of the enzyme activity caused by the vitamin deficiency is demonstrable.     

Rapid dephosphorylation of eIF4E by dietary protein in the skeletal muscle and liver of food-deprived rats

The effect of dietary protein on eIF4E phosphorylation was examined in rats starved for 18 h and then fed on either a 20% casein diet (20C) or a protein-free diet (0C). Refeeding with the 20C diet, but not the 0C diet, resulted in partial dephosphorylation of eIF4E in both the skeletal muscle and liver. The results suggest that the dephosphorylation of eIF4E in response to food intake was regulated by the increase in plasma amino acid concentration that occurred after feeding with the 20C diet.     

维生素E对青鱼幼鱼生长、免疫及抗氨氮胁迫能力的影响

以初始体质量(7.270.40) g的青鱼为研究对象, 采用维生素E(VE)有效含量分别为14.36(对照组)、25.14、37.66、62.97、113.92和210.45 mg/kg 6种等氮等能的实验饲料, 饲养青鱼幼鱼8周后, 根据生长情况选取对照组、62.97和210.45 mg/kg VE组进行24h氨氮胁迫(20 mg/L), 研究VE对青鱼幼鱼生长、免疫及抗氨氮胁迫能力的影响。结果表明: 以特定生长率为指标, 折线模型分析表明青鱼有效维生素E需要量为45.00 mg/kg。肌肉、肝脏和血清VE含量与饲料中VE含量呈明显正相关, 当饲料VE含量超过113.92 mg/kg时, 肌肉和肝脏VE含量均达到饱和。VE对鳃丝Na+/K+-ATP酶活性(NKA)和血清皮质醇(COR)无显著影响, 但随着饲料VE含量的升高, 过氧化氢酶(CAT)和总超氧化物歧化酶活性(T-SOD)呈上升趋势, 丙二醛含量(MDA)呈下降趋势。氨氮胁迫对各处理组肌肉VE含量和血清CAT活性无影响, 但肝脏VE含量均显著降低(P0.05), 且62.97和210.4 5 mg/kg VE组血清VE水平有所升高。在胁迫后, 对照组血清T-SOD、鳃丝NKA活性显著降低, 皮质醇含量显著增加(P0.05)。与对照组相比, 62.97和210.45 mg/kg VE组T-SOD、NKA活性和皮质醇含量在胁迫前后无显著变化。各处理组MDA含量在胁迫后虽均显著升高, 但210.45 mg/kg VE组在胁迫后MDA含量仍显著低于对照组(P0.05)。以上结果说明, 青鱼幼鱼获得最大生长的有效维生素E需求量为45.00 mg/kg, 且较高VE能有效提高青鱼机体免疫力, 缓解氨氮胁迫对青鱼机体的负面影响。       

Digestive enzymes and metabolic profile of Labeo rohita fingerlings fed diets with different crude protein levels

Labeo rohita, commonly called rohu is one of the most important fish species for aquaculture in India. Digestive enzyme response and metabolic profile of fingerling L. rohita to different dietary crude protein (CP) levels (viz. 25, 30, 35 and 40%) were studied in an attempt to optimize a practical diet formulation for this species. After 45 days of feeding, activity of digestive enzymes and metabolite concentrations were assayed. Amylase, lipase and alkaline phosphatase (ALP) activities were not influenced by the dietary protein, but proteolytic and acid phosphatase (ACP) activities varied (P<0.05) between the treatments. Proteolytic activity showed a second order polynomial relationship with dietary crude protein (CP) as Y = 0.0734X(2) + 4.937X - 68.37, r(2)=0.97. A positive correlation was observed between dietary CP and amylase (r(2)=0.78). All the metabolites except muscle glucose showed significant change corresponding to the dietary protein levels. Glucose and glycogen levels corresponded to the dietary carbohydrate levels. Muscle and plasma pyruvic acid increased as the crude protein in the diet increased, whereas liver pyruvic acid showed the opposite trend. Muscle protein content was not affected by dietary CP. Protein fractions in plasma (total protein, albumin and globulin) showed maximum values in 30% CP fed group. It is concluded that proteolytic activity and ACP are the major digestive enzymes responsive to dietary CP in L. rohita fingerlings. Considering the cost effectiveness of the diet, and based on liver and plasma free amino acid levels and plasma protein fractions, 30% crude protein is recommended as the optimal dietary protein for L. rohita fingerlings.     

Effect of dietary vitamin E level on growth, tissue lipid peroxidation, and erythrocyte fragility of transgenic coho salmon, Oncorhynchus kisutch

This study was conducted to investigate the effect of dietary vitamin E concentration on growth performance, iron-catalyzed lipid peroxidation in liver and muscle tissue, and erythrocyte fragility of transgenic growth hormone coho salmon (Oncorhynchus kisutch). Fish were fed one of four isoenergetic and isonitrogenous experimental diets that contained either 11, 29, 50, or 105 IU of vitamin E/kg. Following the 10-week feeding trial, no significant (P>0.05) diet-related differences were detected in growth, whole body proximate composition or erythrocyte fragility. The vitamin E contents of liver and muscle, however, were affected by the dietary treatment. Fish fed diets containing > or =50 IU of vitamin E/kg had significantly increased vitamin E concentrations in their tissues. Iron-catalyzed lipid peroxidation of liver and muscle tissue of fish fed elevated dietary vitamin E (> or =50 IU vitamin E/kg diet) was significantly lower (P<0.05) than that noted for fish fed the diet containing no supplemental vitamin E. The results indicated that changes in tissue lipid peroxidation measurements precede clinical signs of sub-optimal vitamin E intake.     

Digestive enzymes and metabolic profile of Pseudoplatystoma corruscans (Teleostei: Siluriformes) in response to diet composition

Digestive enzyme responsiveness to feeding and associated adjustments of metabolism can be used to derive nutritionally effective diet formulations. Juvenile pintado (Pseudoplatystoma corruscans) were fed different diets. After feeding, fish were killed and blood, liver and white muscle were collected to evaluate metabolites. Stomach along with anterior, middle and posterior intestine were sampled for enzyme analysis. Non-specific protease, trypsin, chymotrypsin, amylase and lipase were assayed. Crude protein (CP) did not induce proteolytic activity; highest protease activities were observed in the stomach. Amylase was higher in the stomach in fish feeding on diets containing 13-25% starch. Lipase activity was observed along the gastrointestinal tract, with the highest activities observed in the middle section. The metabolic profile of white muscle was not affected by CP. In contrast, some plasma and liver metabolites were altered concomitant with changes in the digestive enzymes. Amino acid catabolism was increased. Digestion in pintado was responsive to cornstarch, reflected in intermediary metabolism; proteolytic activities of the digestive tract seem to be sufficient to deal with large amounts of dietary protein. As a result, we are able to recommend a balance between protein and energetic compounds, such as lipids and carbohydrates, in the diet to optimize fish growth.     

Neonatal low-protein diet changes deiodinase activities and pituitary TSH response to TRH in adult rats

Protein malnutrition during neonatal programs for a lower body weight and hyperthyroidism in the adult offspring were analyzed. Liver deiodinase is increased in such animals, contributing to the high serum triiodothyronine (T3) levels. The level of deiodinase activities in other tissues is unknown. We analyzed the effect of maternal protein restriction during lactation on thyroid, skeletal muscle, and pituitary deiodinase activities in the adult offspring. For pituitary evaluation, we studied the in vitro, thyrotropin-releasing hormone (TRH)-stimulated thyroid-stimulating hormone (TSH) secretion. Lactating Wistar rats and their pups were divided into a control (C) group, fed a normal diet (23% protein), and a protein-restricted (PR) group, fed a diet containing 8% protein. At weaning, pups in both groups were fed a normal diet until 180 days old. The pituitary gland was incubated before and after TRH stimulation, and released TSH was measured by radioimmunoassay. Deiodinase activities (D1 and D2) were determined by release of (125)I from [(125)I]reverse triiodothyronine (rT3). Maternal protein malnutrition during lactation programs the adult offspring for lower muscle D2 (-43%, P<0.05) and higher muscle D1 (+83%, P<0.05) activities without changes in thyroidal deiodinase activities, higher pituitary D2 activity (1.5 times, P<0.05), and lower TSH response to in vitro TRH (-56%, P<0.05). The evaluations showed that the lower in vivo TSH detected in adult PR hyperthyroid offspring, programmed by neonatal undernutrition, may be caused by an increment of pituitary deiodination. As described for liver, higher skeletal muscle D1 activity suggests a hyperthyroid status. Our data broaden the knowledge about the adaptive changes to malnutrition during lactation and reinforce the concept of neonatal programming of the thyroid function.     

Increased muscle proteasome activities in rats fed a polyunsaturated fatty acid supplemented diet

Changes in the proteasome system, a dominant actor in protein degradation in eukaryotic cells, have been documented in a large number of physiological and pathological conditions. We investigated the influence of monounsaturated or polyunsaturated fatty acids (PUFAs) supplemented diets on the proteasome system, in rat skeletal muscles. Thirty rats were randomly assigned to three groups. The control group received only a standard diet. The monounsaturated fatty acid (MUFA) enriched diet group was fed with 3% sunflower oil in addition to standard food, and the polyunsaturated fatty acid supplemented diet group received 9% Maxepa) in addition to the standard diet. We analyzed muscle proteasome activities and content. Monounsaturated or PUFAs supplemented diets given for 8 weeks induced a significant increase in proteasome activities. With the polyunsaturated fatty acid enriched diet, the chymotrypsin-like and peptidylglutamylpeptide hydrolase activities increased by 45% in soleus and extensor digitorum longus (EDL), and by 90% in the gastrocnemius medialis (GM) muscle. Trypsin-like activity of the proteasome increased by 250% in soleus, EDL and GM. This increase in proteasome activities was associated with a concomitant enhancement in the muscle content of proteasome. Proteasome activities and level were less stimulated with a monounsaturated fatty acid supplemented diet. This study provides evidence that a monounsaturated or polyunsaturated fatty acid supplemented diet may regulate muscle proteasomes. Unsaturated fatty acids are particularly prone to free radical attack. Thus, we suggest that alterations in muscle proteasome may result from monounsaturated and polyunsaturated fatty acid-induced peroxidation, in order to eliminate damaged proteins.     

Leucine-rich diet supplementation modulates foetal muscle protein metabolism impaired by Walker-256 tumour

Background

Cancer-cachexia induces a variety of metabolic disorders of protein turnover and is more pronounced when associated with pregnancy. Tumour-bearing pregnant rats have impaired protein balance, which decreases protein synthesis and increases muscle breakdown. Because branched-chain amino acids, especially leucine, stimulate protein synthesis, we investigated the effect of a leucine-rich diet on protein metabolism in the foetal gastrocnemius muscles of tumour-bearing pregnant rats.

Methods

Foetuses of pregnant rats with or without Walker 256 tumours were divided into six groups. During the 20 days of the experiment, the pregnant groups were fed with either a control diet (C, control rats; W, tumour-bearing rats; Cp, rats pair-fed the same normoprotein-diet as the W group) or with a leucine-rich diet (L, leucine rats; LW, leucine tumour-bearing rats; and Lp, rats pair-fed the same leucine-rich diet as the LW group). After the mothers were sacrificed, the foetal gastrocnemius muscle samples were resected, and the protein synthesis and degradation and tissue chymotrypsin-like, cathepsin and calpain enzyme activities were assayed. The muscle oxidative enzymes (catalase, glutathione-S-transferase and superoxide dismutase), alkaline phosphatase enzyme activities and lipid peroxidation (malondialdehyde) were also measured.

Results

Tumour growth led to a reduction in foetal weight associated with decreased serum protein, albumin and glucose levels and low haematocrit in the foetuses of the W group, whereas in the LW foetuses, these changes were less pronounced. Muscle protein synthesis (measured by L-[3H]-phenylalanine incorporation) was reduced in the W foetuses but was restored in the LW group. Protein breakdown (as assessed by tyrosine release) was enhanced in the L and W groups, but chymotrypsin-like activity increased only in group W and tended toward an increase in the LW foetuses. The activity of cathepsin H was significantly higher in the W group foetuses, but the proteolytic calcium-dependent pathway showed similar enzyme activity. In parallel, an intense oxidative stress process was observed only in the group W foetuses.

Conclusions

These data suggested that the proteasomal and lysosomal proteolytic pathways and oxidative stress are likely to participate in the process of foetal muscle catabolism of Walker’s tumour-bearing pregnant rats. The present work shows that foetal muscle can be protected by supplementation with a leucine-rich diet.     

饲料中添加丝兰提取物对大菱鲆幼鱼生长和生理及水环境的影响

用丝兰提取物添加量为0.00% (D0)、0.05% (D1)、0.10% (D2)、0.20% (D3)和0.40% (D4)的实验饲料投喂大菱鲆幼鱼(42.200.06) g 60d, 研究其对大菱鲆幼鱼生长性能、体组成、血清免疫代谢指标及养殖水质指标的影响。结果显示, 在饲料中添加丝兰提取物对大菱鲆幼鱼增重率(WGR)、特定生长率(SGR)、饲料系数(FCR)、脏体比(VSI)、肝体比(HSI)、肥满度(CF)均无显著影响(P 0.05)。全鱼、肌肉及肝脏中水分、粗蛋白、粗脂肪含量变化无显著性差异(P 0.05)。D3组幼鱼血清溶菌酶显著高于其他各组(P 0.05), 而D4组各免疫指标明显低于对照组(P 0.05); D3和D4组幼鱼血清的谷丙转氨酶(ALT)、谷草转氨酶(AST)活力显著低于对照组。除A1-6h组外各实验组水体氨氮含量均显著低于对照组(P 0.05), 各实验组亚硝酸盐含量均显著高于对照组(P 0.05), 各实验组总氮、总磷、磷酸盐含量均与对照组无显著性差异(P 0.05)。研究证明, 在饲料中适量添加丝兰提取物能够显著增强大菱鲆幼鱼非特异性免疫能力, 并显著降低养殖水体氨氮含量, 对大菱鲆生长及体组成无显著影响。以非特异性免疫及水体氨氮含量为综合评定指标, 大菱鲆幼鱼饲料中丝兰提取物的适宜添加量为0.20%。       

Influence of salinity and ratio of lipid to protein in diets on certain enzyme activities in rainbow trout (Salmo gairdneri Richardson)

The connection between metabolic and sea water adaptation of the rainbow trout was investigated. The rainbow trout were kept in fresh water and diluted sea water of 8 and 20 0/00 S at 16 degrees C and fed on three different diets for 51 days. Hyperosmotic salinity (20 0/00) tends to inhibit growth in rainbow trout by reducing the food conversion efficiency. A higher protein concentration in the diet can partly compensate for this effect. The liver IDH, G6PDH and 6PGDH activities of the rainbow trout are influenced only by food quality, whereas the liver G1DH, AspT and A1T activities, like the muscle A1T, are also affected by salinity. The salinity had no significant effect on the activities of the kidney enzymes we investigated (Na/K-ATPase, G1DH, A1T, AspT) or of the muscle AspT in these experiments.     

Muscle and liver protein synthesis and degradation in growing rats fed a raw field bean (Vicia faba L.) diet

Body weight gain, food intake, gastrocnemius muscle and liver weight, protein and RNA content, as well as the fractional rates of muscle and liver protein synthesis (ks, according to the method of constant infusion of L-[14C]tyrosine), growth (kg) and degradation (kd), along with RNA activity (g of protein synthesized per day/g RNA) of both organs, were determined in growing male rats fed ad libitum over a period of 10 days on 18.7% protein diets containing either casein (5% of methionine added) (control) or the raw legume field bean (Vicia faba L.) as the sole sources of protein. It has been found that as compared to control rats, those fed the raw legume diet exhibited a significant reduction in the rate of growth, muscle RNA, ks, kg, kd and RNA activity, and a significant increase in liver ks, kd and RNA activity. All differences were statistically significant at least at the 5% level. The possible nature of these findings is discussed.