Intracellular peptides in SARS-CoV-2-infected patients.

Intracellular peptides (InPeps) generated by the orchestrated action of the proteasome and intracellular peptidases have biological and pharmacological significance. Here, human plasma relative concentration of specific InPeps was compared between 175 patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and 45 SARS-CoV-2 non-infected patients; 2,466 unique peptides were identified, of which 67% were InPeps. The results revealed differences of a specific group of peptides in human plasma comparing non-infected individuals to patients infected by SARS-CoV-2, following the results of the semi-quantitative analyses by isotope-labeled electrospray mass spectrometry. The protein-protein interactions networks enriched pathways, drawn by genes encoding the proteins from which the peptides originated, revealed the presence of the coronavirus disease/COVID-19 network solely in the group of patients fatally infected by SARS-CoV-2. Thus, modulation of the relative plasma levels of specific InPeps could be employed as a predictive tool for disease outcome.

The Relevance of Thimet Oligopeptidase in the Regulation of Energy Metabolism and Diet-Induced Obesity.

Thimet oligopeptidase (EC 3.4.24.15; EP24.15; THOP1) is a potential therapeutic target, as it plays key biological functions in processing biologically functional peptides. The structural conformation of THOP1 provides a unique restriction regarding substrate size, in that it only hydrolyzes peptides (optimally, those ranging from eight to 12 amino acids) and not proteins. The proteasome activity of hydrolyzing proteins releases a large number of intracellular peptides, providing THOP1 substrates within cells. The present study aimed to investigate the possible function of THOP1 in the development of diet-induced obesity (DIO) and insulin resistance by utilizing a murine model of hyperlipidic DIO with both C57BL6 wild-type (WT) and THOP1 null (THOP1-/-) mice. After 24 weeks of being fed a hyperlipidic diet (HD), THOP1-/- and WT mice ingested similar chow and calories; however, the THOP1-/- mice gained 75% less body weight and showed neither insulin resistance nor non-alcoholic fatty liver steatosis when compared to WT mice. THOP1-/- mice had increased adrenergic-stimulated adipose tissue lipolysis as well as a balanced level of expression of genes and microRNAs associated with energy metabolism, adipogenesis, or inflammation. Altogether, these differences converge to a healthy phenotype of THOP1-/- fed a HD. The molecular mechanism that links THOP1 to energy metabolism is suggested herein to involve intracellular peptides, of which the relative levels were identified to change in the adipose tissue of WT and THOP1-/- mice. Intracellular peptides were observed by molecular modeling to interact with both pre-miR-143 and pre-miR-222, suggesting a possible novel regulatory mechanism for gene expression. Therefore, we successfully demonstrated the previously unanticipated relevance of THOP1 in energy metabolism regulation. It was suggested that intracellular peptides were responsible for mediating the phenotypic differences that are described herein by a yet unknown mechanism of action.

Pequi (Caryocar brasiliense Camb.) almond oil attenuates carbon tetrachloride-induced acute hepatic injury in rats: Antioxidant and anti-inflammatory effects.

Carbon tetrachloride (CCl4) is a potent hepatotoxin, capable of generating free radicals that lead to oxidative stress and the inflammation process. Pequi almond oil (PAO) has been reported to possess unsaturated fatty acid and antioxidant compounds related to beneficial effects on oxidation and inflammatory conditions. The present study was undertaken to evaluate the hepatoprotective effects of handmade and coldpressed PAO on CCl4-induced acute liver injury. The possible mechanisms underlying the effect on liver injury enzymes, histopathological parameters, lipid profile, lipid peroxidation, and antioxidant and detoxification defense systems, as well as inflammatory parameters, were determined. Rats treated with PAO (3 or 6 mL/kg) for 21 days before CCl4 induction (3 mL/kg, 70%) showed significantly decreased levels of alanine aminotransferase and aspartate aminotransferase, milder hepatic lesions and higher levels of serum high-density lipoprotein compared to CCl4 group. Moreover, PAO enhanced antioxidant capacity by increasing hepatic glutathione peroxidase and glutathione reductase enzyme activities, as well as reducing circulating concentrations of leptin and inflammatory mediators such as interleukin-6, leukotrienes -4 and -5 and the tumor necrosis factor receptor. In summary, PAO, especially cold-pressed oil, attenuated the CCl4-induced alterations in serum and hepatic tissue in rats due to its antioxidant and anti-inflammatory properties.

Adipose tissue inflammation and cancer cachexia: the role of steroid hormones.

Adipose tissue inflammation plays a role in the etiology of many chronic diseases, and has been the focus of much attention in the context of obesity and metabolic syndrome. Similarly, during cancer cachexia, a syndrome that markedly increases cancer-associated morbidity and mortality, local adipose inflammation is reported in animal models and in patients, potentially contributing to the chronic systemic inflammation that constitutes the hallmark of this condition. We discuss, on the basis of information generated by obesity-related studies, the possible relation between adipose tissue inflammation and compromised steroid hormone secretion and action in cachexia.

Cessation of physical exercise changes metabolism and modifies the adipocyte cellularity of the periepididymal white adipose tissue in rats.

All of the adaptations acquired through physical training are reversible with inactivity. Although significant reductions in maximal oxygen uptake (Vo2max) can be observed within 2 to 4 wk of detraining, the consequences of detraining on the physiology of adipose tissue are poorly known. Our aim was therefore to investigate the effects of discontinuing training (physical detraining) on the metabolism and adipocyte cellularity of rat periepididymal (PE) adipose tissue. Male Wistar rats, aged 6 wk, were divided into three groups and studied for 12 wk under the following conditions: 1) trained (T) throughout the period; 2) detrained (D), trained during the first 8 wk and detrained during the remaining 4 wk; and 3) age-matched sedentary (S). Training consisted of treadmill running sessions (1 h/day, 5 days/wk, 50-60% Vo2max). The PE adipocyte size analysis revealed significant differences between the groups. The adipocyte cross-sectional area (in µm(2)) was significantly larger in D than in the T and S groups (3,474 ± 68.8; 1,945.7 ± 45.6; 2,492.4 ± 49.08, respectively, P < 0.05). Compared with T, the isolated adipose cells (of the D rats) showed a 48% increase in the ability to perform lipogenesis (both basal and maximally insulin-stimulated) and isoproterenol-stimulated lipolysis. No changes were observed with respect to unstimulated lipolysis. A 15% reduction in the proportion of apoptotic adipocytes was observed in groups T and D compared with group S. The gene expression levels of adiponectin and PPAR-gamma were upregulated by factors of 3 and 2 in D vs. S, respectively. PREF-1 gene expression was 3-fold higher in T vs. S. From these results, we hypothesize that adipogenesis was stimulated in group D and accompanied by significant adipocyte hypertrophy and an increase in the lipogenic capacity of the adipocytes. The occurrence of apoptotic nuclei in PE fat cells was reduced in the D and T rats; these results raise the possibility that the adipose tissue changes after detraining are obesogenic.

Changes in the production of IL-10 and TNF-alpha in skeletal muscle of rats with heart failure secondary to acute myocardial infarction.

BACKGROUND: Recent studies show that the expression of inflammatory mediators, such as cytokines, is an important factor for the development and progression of heart failure (HF), especially in the presence of left ventricular dysfunction. These changes have been demonstrated both in the plasma and heart muscle and, more recently, in skeletal muscle of rats and in patients with HF. OBJECTIVE: To investigate the production and expression of tumor necrosis factor-alpha (TNF) and interleukin-10 (IL-10) in the soleus and the extensor digitorum longus (EDL) muscles of animals with left ventricular dysfunction after myocardial infarction (MI). METHODS: We used male Wistar rats that underwent ligation of the left coronary artery without reperfusion. Four weeks after this procedure, the animals underwent echocardiography and were divided into the following experimental groups: sham operated (sham) and IM. They remained under observation for a further period of 8 weeks. RESULTS: The level of the cytokine TNF-alpha increased by 26.5% (p <0.05), and its gene expression increased 3 times (p <0.01). The level of IL-10 decreased by 38.2% (p <0.05). Both changes occurred only in the soleus muscle, with no change in the EDL. The decrease (36.5%, p <0.05) in the IL-10/TNF-alpha ratio was due to both increased tissue levels of TNF-alpha and decreased tissue levels of IL-10. CONCLUSION: Our results showed significant changes in the IL-10/TNF-alpha ratio, which may have an additive role in the assessment of deterioration and progression of left ventricular dysfunction post-MI. Furthermore, our study suggests that these changes seem to be related to the muscle fiber type.

Alterações na produção de IL-10 e TNF-α no músculo esquelético em ratos com insuficiência cardíaca secundária ao infarto do miocárdio / Changes in the production of IL-10 and TNF-α in skeletal muscle of rats with heart failure secondary to acute myocardial infarction

FUNDAMENTO: Estudos recentes demonstram que a expressão de mediadores inflamatórios, como as citocinas, é um importante fator de desenvolvimento e progressão da insuficiência cardíaca (IC), principalmente na presença de disfunção ventricular esquerda. Essas alterações têm sido demonstradas tanto no plasma como no músculo cardíaco e, mais recentemente, no músculo esquelético de ratos e pacientes com IC. OBJETIVO: Investigar a produção e expressão do fator de necrose tumoral-α (TNF-α) e interleucina-10 (IL-10) no músculo sóleo e extensor digital longo (EDL) em animais com disfunção ventricular pós-infarto do miocárdio (IM). MÉTODOS: Utilizaram-se ratos Wistar machos que foram submetidos à ligadura da artéria coronária esquerda sem posterior reperfusão. Quatro semanas após esse procedimento, os animais foram submetidos à análise ecocardiográfica e divididos nos seguintes grupos experimentais: falso operado (Sham) e IM. Mantiveram-se em observação por um período adicional de 8 semanas. RESULTADOS: O nível da citocina TNF-α aumentou 26,5 por cento (p < 0,05), e sua expressão gênica, 3 vezes (p < 0,01). O nível de IL-10 apresentou diminuição de 38,2 por cento (p < 0,05). Ambas as alterações ocorreram apenas no músculo sóleo, sem alterações no EDL. A diminuição (36,5 por cento, p < 0,05) na razão IL-10/ TNF-α deveu-se tanto ao aumento dos níveis teciduais do TNF-α quanto à diminuição da IL-10 dos níveis teciduais. CONCLUSÃO: Nossos resultados demonstraram alterações relevantes na razão IL-10/ TNF-α, o que pode ter um papel aditivo na avaliação da deterioração e progressão do quadro da disfunção ventricular esquerda pós-IM. Além disso, nosso estudo sugere que essas alterações parecem estar relacionadas ao tipo de fibra muscular.

BACKGROUND: Recent studies show that the expression of inflammatory mediators, such as cytokines, is an important factor for the development and progression of heart failure (HF), especially in the presence of left ventricular dysfunction. These changes have been demonstrated both in the plasma and heart muscle and, more recently, in skeletal muscle of rats and in patients with HF. OBJECTIVE: To investigate the production and expression of tumor necrosis factor-α (TNF) and interleukin-10 (IL-10) in the soleus and the extensor digitorum longus (EDL) muscles of animals with left ventricular dysfunction after myocardial infarction (MI). METHODS: We used male Wistar rats that underwent ligation of the left coronary artery without reperfusion. Four weeks after this procedure, the animals underwent echocardiography and were divided into the following experimental groups: sham operated (sham) and IM. They remained under observation for a further period of 8 weeks. RESULTS: The level of the cytokine TNF-α increased by 26.5 percent (p <0.05), and its gene expression increased 3 times (p <0.01). The level of IL-10 decreased by 38.2 percent (p <0.05). Both changes occurred only in the soleus muscle, with no change in the EDL. The decrease (36.5 percent, p <0.05) in the IL-10/TNF-α ratio was due to both increased tissue levels of TNF-α and decreased tissue levels of IL-10. CONCLUSION: Our results showed significant changes in the IL-10/TNF-α ratio, which may have an additive role in the assessment of deterioration and progression of left ventricular dysfunction post-MI. Furthermore, our study suggests that these changes seem to be related to the muscle fiber type. (Arq Bras Cardiol 2010; 94(3):293-300)

Endurance training modulates lymphocyte function in rats with post-MI CHF.

PURPOSE: Exercise training restores innate immune system cell function in post-myocardial infarction (post-MI) rats. However, studies of the involvement of lymphocyte (Ly) in the setting of the congestive heart failure (CHF) are few. To address this issue, we investigated the function of Ly obtained from cervical lymph nodes from post-MI CHF rats submitted to treadmill running training. METHODS: Twenty-five male Wistar rats were randomly assigned to the following groups: rats submitted to ligation of the left coronary artery, which were sedentary (MI-S, N = 7, only limited activity) or trained (MI-T, N = 6, on a treadmill (0% grade at 13-20 m.m) for 60 min.d, 5 d.wk, for 8-10 wk); or sham-operated rats, which were sedentary (sham-S, N = 6) or trained (sham-T, N = 6). The incorporation of [2-C]-thymidine by Ly cultivated in the presence of concanavalin A (Con A) and lipopolysaccharide (LPS), cytokine production by Ly cultivated in the presence of phytohemagglutinin (PHA), and plasma concentration of glutamine were assessed in all groups, 48 h after the last exercise session. RESULTS: Proliferative capacity was increased, following incubation with Con-A in the MI groups, when compared with the sham counterparts. When incubated in the presence of PHA, MI-S produced more IL-4 (96%) than sham-S (P < 0.001). The training protocol induced a 2.2-fold increase in the production of interleukin-2 (P < 0.001) of the cells obtained from the cervical lymph nodes of MI-T, compared with MI-S. CONCLUSION: The moderate endurance training protocol caused an increase in IL-2 production, and a trend toward the reversion of the Th1/Th2 imbalance associated with IL-4 production increased in the post-MI CHF animal model.

Lipid metabolism in trained rats: effect of guarana (Paullinia cupana Mart.) supplementation.

BACKGROUND AND AIMS: Guarana is widely consumed by athletes, either in supplements or in soft drinks, under the belief that it presents ergogenic and "fat burning" effects. We examined the effect of guarana supplementation (14 days) upon aspects of lipid metabolism in sedentary (C) and trained rats (T). METHODS: To isolate the effect of caffeine from that of other components of guarana, we adopted two different doses of whole extract (G1-0.130 g/kg; G2-0.325 g/kg) or decaffeinated extract (DG1, DG2). Body weight, food and water intake; muscle fat content, oleate incorporation, glycogen content, and carnitine palmitoyltransferase I (CPT I) activity and mRNA expression; along with plasma lactate concentration, were assessed. RESULTS: Muscle oleate incorporation was decreased in rats receiving decaffeinated guarana in relation to G1 and G2; as was CPT I mRNA expression in the gastrocnemius. Whole extract supplementation, but not DG induced reduced plasma lactate concentration in trained rats. G1 showed higher muscle glycogen content compared with all other groups. The results show an effect of guarana on aspects of lipid metabolism, which is abolished by decaffeination. CONCLUSION: The changes in lipid metabolism of supplemented rats herein reported are associated with the methylxanthine content of guarana.

Cancer cachexia modifies the zonal distribution of lipid metabolism-related proteins in rat liver.

Cancer cachexia is a syndrome that causes profound metabolic disruption. Lipid metabolism in the liver is markedly affected. We investigated the effect of cachexia upon liver-acinus lipid-metabolism zonation in Walker 245 carcinosarcoma-bearing rats (TB). The expression of protein (by Western blotting) and mRNA (by semi-quantitative polymerase chain reaction) of the enzymes of the carnitine palmitoyltransferase system (CPT I and CPT II) and of liver fatty-acid-binding protein (L-FABP) was studied. Although no changes were found for these parameters, the maximal activities (by radioassay) of CPT I and II were reduced (P<0.05) in TB compared with controls. CPT II activity in the perivenous (PV) region was higher in TB compared with controls. The distribution of CPT II and L-FABP (by immunohistochemistry) within the acinus was modified by cachexia: whereas CPT II positivity was restricted to the PV zone, L-FABP labelling shifted from periportal (control) to perivenous (TB) zone. These changes in metabolic zonation, together with decreased CPT II activity, may contribute to the aggravation of cachexia.

Adipose tissue in Walker 256 tumour-induced cachexia: possible association between decreased leptin concentration and mononuclear cell infiltration.

The adipose tissue (AT) is severely affected by cachexia, a paraneoplastic syndrome, which increases the morbidity and mortality of cancer. There is, however, a heterogeneous response to the condition, according to the AT depot. As plasma leptin concentration has been often reported to vary in cachexia, we have measured (species specific radioimmunoassay) the local concentration of leptin in three AT depots: retroperitoneal (RPAT), epididymal (EAT) and mesenteric (MES) of Walker 256 tumour-bearing rats. A reduced concentration of leptin ( P<0.0001) was found in all the depots and in the plasma of the cachectic rats, compared with controls already from day 4 after tumour cell injection. The presence of a cell infiltrate was observed in all AT obtained from the tumour-bearing animals. Ultrastructural analysis, along with immunocytochemistry for RT1B (indicating the presence of MHCII) and using antibody against mononuclear phagocytes, showed the cells to be macrophages. The profile of TNFalpha and PGE2 secretion by the infiltrate was investigated (commercial kits). There was increased production of both factors by the cells of all AT ( P<0.05) compared with peritoneal macrophages obtained from the cachectic rats, while the cells isolated from MES showed the highest synthesis of TNFalpha. The results suggest a possible modulation of the chronic locally produced TNFalpha and PGE2 upon leptin synthesis by the AT of the cachectic rats.

Rat myocellular and perimysial intramuscular triacylglycerol: a histological approach.

PURPOSE: There is controversy as to the use of intramuscular triacylglycerol (IMTAG) during exercise and to whether endurance training increases its utilization, despite the various methodologies used to address these questions. We used a histological-morphometrical approach to study the relative contribution of the two compartments of IMTAG storage, intramyocellular, and perimysial adipocytes, during exercise in sedentary and endurance-trained rats. METHODS: After osmium impregnation, the soleus (SOL) and gastrocnemius (GAS) were studied under light and electron microscopy. IMTAG content (after Triton WR1339 treatment or not) and 14C-oleate incorporation into the muscles were studied. RESULTS: In GAS, training, but not exercise alone, decreased extramyocellular lipid (P < 0.001 vs sedentary), an effect not found for SOL. Both muscles presented reduced lipid inclusion number (P < 0.001) and area (P < 0.05), immediately after exercise in sedentary and trained rats. For SOL, a greater number (P < 0.001 vs sedentary) of inclusions was found 24 h after exercise in trained rats. Triton WR1339 treatment decreased IMTAG content 12 h after exercise in SOL (but not in GAS), in sedentary (33%), and trained rats (52%). CONCLUSIONS: The multi-analytical approach adopted allowed the discernment between the IMTAG compartments and provided evidence for an effect of training upon storage of lipid in perimysial adipocytes in rat gastrocnemius, as well as clearly showed that the IMTAG mobilized during submaximal exercise in sedentary and trained rats derives from intramyocellular lipid, both in SOL and GAS. Moreover, the reposition of these stores 12 h after exercise was shown to be different in GAS and SOL, as plasma triacylglycerol clearly contributed to the process only in the latter, possibly reflecting the differences in lipoprotein lipase activity in the muscles reported by others.

Liver denervation affects hepatocyte mitochondrial fatty acid transport capacity.

The effect of liver denervation on the activity of hepatic carnitine palmitoyltransferase (CPT) system, which catalyses the transfer of long-chain fatty acids into the mitochondria, was studied in rats. Noradrenaline content in phenol-denervated liver (D) was reduced by 87%. CPT I and II activities (measured by radioassay after detergent separation of the enzymes) were decreased (p < 0.001) in D (2.6 +/- 0.1 and 0.68 +/- 0.2 nmol min(-1) mg(-1) protein, respectively) as compared with controls (4.7 +/- 0.3 and 2.5 +/- 0.2 nmol min(-1) mg(-1) protein, for CPT I and II, respectively). A less intense immunoreactive band for denervated liver CPT II was obtained after Western blotting. Concomitantly, long-chain fatty acid incorporation (p < 0.001), evaluated after administration of [14C]-oleate and total fat content (p < 0.001) were increased in D in relation to controls, while incorporation of exogenous [14C]-oleate into secreted VLDL, was decreased (p < 0.01). The effect of sympathetic denervation on CPT activity was different from that evoked by adrenodemedullation, which caused an augmentation of CPT activity (p < 0.01), when compared with the liver of intact rats. The effects of denervation and adrenodemedullation on the other parameters of lipid metabolism studied, were similar. The results strongly suggest a role of liver sympathetic innervation in the regulation of liver lipid metabolism.

Metabolic aspects in cancer cachexia

The present review concerns metabolic alterations related to the establishment of cancer cachexia, a condition of extensive catabolism, responsible, in a great percentage of the cases, for the death of the patient. A brief review on the condition is presented and two major aspects of the host's altered metabolism are considered: the diminished plasma levels of insulin, triggering the catabolic mechanisms; and the reduced carnitine palmitoyltransferase II activity in hepatic mitochondria, rendering the liver unable of oxidizing fatty acids and of synthesizing ketone bodies. These modifications lead to the intensification of the wasting process and thus, from our point of view, might play an important role for the treatment of patients with neoplastic disease.