Exercise Reduces the Resumption of Tumor Growth and Proteolytic Pathways in the Skeletal Muscle of Mice Following Chemotherapy

The pathogenesis of muscle atrophy plays a central role in cancer cachexia, and chemotherapy contributes to this condition. Therefore, the present study aimed to evaluate the effects of endurance exercise on time-dependent muscle atrophy caused by doxorubicin. For this, C57 BL/6 mice were subcutaneously inoculated with Lewis lung carcinoma cells (LLC group). One week after the tumor establishment, a group of these animals initiated the doxorubicin chemotherapy alone (LLC + DOX group) or combined with endurance exercise (LLC + DOX + EXER group). One group of animals was euthanized after the chemotherapy cycle, whereas the remaining animals were euthanized one week after the last administration of doxorubicin. The practice of exercise combined with chemotherapy showed beneficial effects such as a decrease in tumor growth rate after chemotherapy interruption and amelioration of premature death due to doxorubicin toxicity. Moreover, the protein degradation levels in mice undergoing exercise returned to basal levels after chemotherapy; in contrast, the mice treated with doxorubicin alone experienced an increase in the mRNA expression levels of the proteolytic pathways in gastrocnemius muscle (Trim63, Fbxo32, Myostatin, FoxO). Collectively, our results suggest that endurance exercise could be utilized during and after chemotherapy for mitigating muscle atrophy promoted by doxorubicin and avoid the resumption of tumor growth

The impact of doxorubicin treatment on the functions of white adipose tissue.

Introdução: A doxorrubicina (DOX) é um quimioterápico que gera efeitos tóxicos no tecido adiposo (T.A.) e reduz a qualidade de vida dos pacientes. Objetivos: Investigar os efeitos metabólicos do tratamento com DOX no T.A. branco e propor terapia adjuvante que atenue efeitos deletérios. Métodos: Procedimento experimental 1: ratos Wistar foram tratados com dose única de DOX (15mg/kg). Cultura de células: 3T3L1 foram incubadas por 24h, 96h e 12 dias com DOX. Procedimento experimental 2: animais C57/BL6 receberam doses fracionadas de DOX associado ao uso de metformina (MET) (300mg/kg, diário) ou não. Conclusão: A DOX gera um alto impacto sobre a homeostasia do T.A. branco tanto no tratamento agudo com dose única, como no tratamento crônico com doses mais baixas. Os processos fisiológicos do tecido adiposo sofreram profundas alterações, o que resultou em menor tamanho do adipócito, maior fibrose, diminuição das vias metabólicas e redução da adiponectina e leptina circulantes, e o tratamento com MET não reverteu esses efeitos, só prevenindo o processo de fibrose do TA.Introduction: Doxorubicin (DOX) is a chemotherapy that generates toxic effects on adipose tissue (AT) and reduces the quality of life of patients. Objectives: To investigate the metabolic effects of treatment with DOX on AT white and to propose adjuvant therapy to mitigate deleterious effects. Methods: Experimental Procedure 1: Wistar rats were treated with a single dose of doxorubicin (15mg/ kg). Cell Culture: 3T3-L1 were incubated for 24h, 96h and 12 days with doxorubicin. Experimental procedure 2: C57/BL6 mice received fractionated doses of DOX associated with the use of metformin (MET) (300 mg/kg daily) or not. Conclusion: DOX generates a high impact on the homeostasis of white AT in both acute single dose treatment, such as in chronic treatment with lower doses. The physiological processes of AT have undergone major changes, resulting in a smaller of adipocytes, increased fibrosis, reduction in metabolic pathways and decreased circulating adiponectin and leptin, and the treatment with MET did not reverse these effects, only prevent the fibrosis process on AT

Role of PPAR-γ on immunometabolic effects of adipose tissue and macrophages, in a model of induced colon cancer treated with high-fat diet.

Introdução: O câncer de colón é o segundo tipo de câncer que acomete maior número de casos no Brasil. Atualmente a obesidade é um fator de risco para o câncer de cólon, em contrapartida, alguns autores verificaram que o uso de dieta hiperlipídica associado ao modelo de carcinogênese de cólon aumentou o tempo de sobrevida, recuperação do peso corporal e reduziu o tamanho da região tumoral. Portanto, essa relação do câncer de cólon com a obesidade precisa ser melhor investigada. O PPARγ é um fator nuclear que possui propriedades metabólicas, anticarcinogênicas e anti-inflamatórias, que é expresso em adipócitos e macrófagos. Agonistas de PPARγ podem aumentar a sobrevida e qualidade de vida, apesar disso, ainda não se sabe se esse efeito resulta pela redução da perda de massa adiposa assim como suas funções metabólicas e endócrinas ou se esses efeitos são mais pronunciados pela resposta anti-inflamatória controlada pelo PPARγ, principalmente observada nos macrófagos. No entanto, o objetivo deste trabalho foi investigar o papel do PPARγ na carcinogênese de cólon, identificando como, de fato, a deleção desse receptor nuclear em macrófagos afeta a carcinogênese em animais obesos e em células CACO-2. Métodos: Camundongos PPARγ CreLox com deleção específica em células mieloides (KO) e animais controles da mesma ninhada (WT) foram divididos em: dieta padrão; dieta hiperlipídica; dieta hiperlipídica e indução do câncer de cólon; dieta hiperlipídica, câncer e pioglitazona por 12 semanas. Células humanas da linhagem CACO-2 foram tratadas com agonista e antagonista de PPARγ. Foram avaliados a expressão proteica de PPARγ, citocinas inflamatórias, adipocinas, citometria de fluxo, análise de ciclo celular e morte celular. Resultados: Animais KO possuem menor porcentagem de macrófagos CD80+Ly6Chigh no cólon; e a dieta hiperlipídica diminui a porcentagem de macrófagos Ly6Chigh no tecido adiposo. A deleção de PPARγ foi eficiente em atenuar a perda de peso promovida pelo câncer e reduzir a expressão gênica de IL-10, IL-6 e IL-1β no tecido adiposo subcutâneo. O câncer atenuou a porcentagem de macrófagos Ly6clow no tecido adiposo subcutâneo e pioglitazona recuperou essa porcentagem. O câncer levou ao aumento da porcentagem de macrófagos CD80+ Ly6Chigh e o total de macrófagos Ly6Clow no intestino grosso dos camundongos KO. Em células CACO-2, o uso de agonista e antagonista de PPARγ não modulou a permeabilidade intestinal, apoptose, ciclo celular, porém ambos reduziram a concentração de MCP-1. Conclusão: A modulação do PPARγ está alterando o perfil de macrófagos presentes no tecido adiposo subcutâneo e no cólon, além de alterar a resposta inflamatória resposta inflamatória do tecido adiposo subcutâneo. A ausência de PPARγ em células meloides reduziu marcadores inflamatórios no tecido adiposo de animais portadores de tumor, porém isso não foi efetivo para melhorar a sobrevida.Introduction: Colon cancer is the second type of cancer that affects the largest number of cases in Brazil. Obesity is currently a risk factor for colon cancer, on the other hand, some authors found that the use of high-fat diet associated with the colon carcinogenesis model increased survival time, body weight recovery and reduced the size of the tumor region. Therefore, this relationship between colon cancer and obesity needs to be further investigated. PPARγ is a nuclear factor that has metabolic, anti-carcinogenic and anti-inflammatory properties, which is expressed in adipocytes and macrophages. PPARγ agonists may increase survival and quality of life, although it is not known whether this effect results from reduced fat loss as well as its metabolic and endocrine functions or whether these effects are more pronounced by the controlled anti-inflammatory response by PPARγ, mainly seen in macrophages. However, the aim of this work was to investigate the role of PPARγ in colon carcinogenesis, identifying how, in fact, the deletion of this nuclear receptor in macrophages will affect the carcinogenesis process in obese animals and in CACO-2 cells. Methods: PPARγ CreLox mice with specific deletion in myeloid cells (KO) and control littermates (WT) were divided into: standard diet; high-fat diet; high-fat diet and colon cancer induction; high fat diet, cancer and pioglitazone during 12 weeks. Human cells of the CACO-2 lineage were treated with PPARγ agonist and antagonist. The protein content of PPARγ, inflammatory cytokines, adipokines, flow cytometry, cell cycle analysis and cell death were evaluated. Results: KO animals have a lower percentage of CD80+Ly6Chigh macrophages in the colon; and the high fat diet decreases the percentage of Ly6Chigh macrophages in the adipose tissue. The PPARγ deletion was effective in attenuating cancer-promoted weight loss and reducing the gene expression of IL-10, IL-6 and IL-1β in subcutaneous adipose tissue. Cancer attenuated the percentage of Ly6clow macrophages in the subcutaneous adipose tissue and pioglitazone regained this percentage. The cancer led to an increase in the percentage of CD80+ Ly6Chigh macrophages and the total of Ly6clow macrophages in the large intestine of KO mice. In CACO-2 cells the use of PPARγ agonist and antagonist did not modulate intestinal permeability, apoptosis, cell cycle, but both reduced MCP-1 concentration. Conclusion: PPARγ modulation is changing the profile of macrophages present in the subcutaneous adipose tissue and in the colon, besides to altering the inflammatory response to the inflammatory response of the subcutaneous adipose tissue. The absence of PPARγ in myeloid cells reduced inflammatory markers in the adipose tissue of tumor-bearing animals, however this was not effective in improving survival

Tributyrin in Inflammation: Does White Adipose Tissue Affect Colorectal Cancer?

Colorectal cancer affects the large intestine, leading to loss of white adipose tissue (WAT) and alterations in adipokine secretion. Lower incidence of colorectal cancer is associated with increased fibre intake. Fructooligosaccharides (FOS) are fibres that increase production of butyrate by the intestinal microbiota. Tributyrin, a prodrug of butyric acid, exerts beneficial anti-inflammatory effects on colorectal cancer. Our aim was to characterise the effects of diets rich in FOS and tributyrin within the context of a colon carcinogenesis model, and characterise possible support of tumorigenesis by WAT. C57/BL6 male mice were divided into four groups: a control group (CT) fed with chow diet and three colon carcinogenesis-induced groups fed either with chow diet (CA), tributyrin-supplemented diet (BUT), or with FOS-supplemented diet. Colon carcinogenesis decreased adipose mass in subcutaneous, epididymal, and retroperitoneal tissues, while also reducing serum glucose and leptin concentrations. However, it did not alter the concentrations of adiponectin, interleukin (IL)-6, IL-10, and tumour necrosis factor alpha (TNF)-α in WAT. Additionally, the supplements did not revert the colon cancer affected parameters. The BUT group exhibited even higher glucose tolerance and levels of IL-6, VEGF, and TNF-α in WAT. To conclude our study, FOS and butyrate supplements were not beneficial. In addition, butyrate worsened adipose tissue inflammation

Impact of Doxorubicin Treatment on the Physiological Functions of White Adipose Tissue

<div><p>White adipose tissue (WAT) plays a fundamental role in maintaining energy balance and important endocrine functions. The loss of WAT modifies adipokine secretion and disrupts homeostasis, potentially leading to severe metabolic effects and a reduced quality of life. Doxorubicin is a chemotherapeutic agent used clinically because of its good effectiveness against various types of cancer. However, doxorubicin has deleterious effects in many healthy tissues, including WAT, liver, and skeletal and cardiac muscles. Our objective was to investigate the effects of doxorubicin on white adipocytes through <i>in vivo</i> and <i>in vitro</i> experiments. Doxorubicin reduced the uptake of glucose by retroperitoneal adipocytes and 3T3-L1 cells via the inhibition of AMP-activated protein kinase Thr172 phosphorylation and glucose transporter 4 content. Doxorubicin also reduced the serum level of adiponectin and, to a greater extent, the expression of genes encoding lipogenic (<i>Fas</i> and <i>Acc</i>) and adipogenic factors (<i>Pparg</i>, <i>C/ebpa</i>, and <i>Srebp1</i>c) in retroperitoneal adipose tissue. In addition, doxorubicin inhibited both lipogenesis and lipolysis and reduced the hormone-sensitive lipase and adipose tissue triacylglycerol lipase protein levels. Therefore, our results demonstrate the impact of doxorubicin on WAT. These results are important to understand some side effects observed in patients receiving chemotherapy and should encourage new adjuvant treatments that aim to inhibit these side effects.</p></div

Palmitoleic acid reduces high fat diet-induced liver inflammation by promoting PPAR-γ-independent M2a polarization of myeloid cells

Palmitoleic acid (POA, 16:1n-7) is a lipokine that has potential nutraceutical use to treat non-alcoholic fatty liver disease. We tested the effects of POA supplementation (daily oral gavage, 300 mg/Kg, 15 days) on murine liver inflammation induced by a high fat diet (HFD, 59% fat, 12 weeks). In HFD-fed mice, POA supplementation reduced serum insulin and improved insulin tolerance compared with oleic acid (OA, 300 mg/Kg). The livers of POA-treated mice exhibited less steatosis and inflammation than those of OA-treated mice with lower inflammatory cytokine levels and reduced toll-like receptor 4 protein content. The anti-inflammatory effects of POA in the liver were accompanied by a reduction in liver macrophages (LM, CD11c+; F4/80+; CD86+), an effect that could be triggered by peroxisome proliferator activated receptor (PPAR)-γ, a lipogenic transcription factor upregulated in livers of POA-treated mice. We also used HFD-fed mice with selective deletion of PPAR-γ in myeloid cells (PPAR-γ KOLyzCre+) to test whether the beneficial anti-inflammatory effects of POA are dependent on macrophages PPAR-γ. POA-mediated improvement of insulin tolerance was tightly dependent on myeloid PPAR-γ, while POA anti-inflammatory actions including the reduction in liver inflammatory cytokines were preserved in mice bearing myeloid cells deficient in PPAR-γ. This overlapped with increased CD206+ (M2a) cells and downregulation of CD86+ and CD11c+ liver macrophages. Moreover, POA supplementation increased hepatic AMPK activity and decreased expression of the fatty acid binding scavenger receptor, CD36. We conclude that POA controls liver inflammation triggered by fat accumulation through induction of M2a macrophages independently of myeloid cell PPAR-γ.</p

Adipogenesis is compromised by doxorubicin in 3T3L1 cells.

<p>Gene expression of adipogenics factors <i>Ppparg</i> (A), <i>C/ebpa</i> (B) and <i>Srebp1c</i> (C) at 24 hours, 96 hours and 12 days after induction of cell differentiation in 3T3L1 cells treated with 1uM of doxorubicin. The gene expression was evaluated by real time PCR. Data are mean ± standard deviation of 4 to 6 experiments. The groups were compared using the Two way ANOVA followed by Bonferroni post test, *p<0.05, **p<0.01 and ***p<0.001.</p

Doxorubicin inhibits the lipolysis process.

<p>Glycerol released in assay of lipolysis the adipocytes isolated from retroperitoneal adipose tissue of rats after 72 hours of doxorubicin administration (15mg/kg of weighti.p), with or without isoproterenol stimulation (10uM) for 30 minutes (A). Concentration of glycerol (B) and lactate dehydrogenase LDH in incubation medium of 3T3L1 (C). Lipolysis was stimulated with isoproterenol (Iso) 10uM with doxorubicin (DX) at concentrations of 0.1 and 1uM for 24 hours, the parameters are normalized by the concentration of protein extracted from the cells. Values represent the mean ± standard deviation of 4 to 8 experiments. The groups were compared using the One-Way ANOVA followed by Bonferroni post test. *p<0.05, **p<0.01, ***p<0.001 (A, B and C). Protein expression of HSL phosphorylated at residue Ser-565, Ser-660, total HSL, ATGL and GAPDH (D). Proteins extracted from retroperitoneal adipose tissue of rats 72 hours after treatment with doxorubicin (15mg/kg). Figure represents 4 experiments per group (D).</p

Doxorubicin reduces lipogenic enzymes.

<p>Gene expression of lipogenic enzymes <i>Fas</i> (A) and <i>Acc</i> (B) at 24 hours, 96 hours and 12 days after induction of cell differentiation in 3T3L1 cells treated with 1uM of doxorubicin. The gene expression was evaluated by real time PCR. Data are mean ± standard deviation of 4 to 6 experiments. The groups were compared using the Two way ANOVA followed by Bonferroni post test, *p <0.05, **p<0.01 and ***p<0.001.</p