Vanesa Bol

Forced catch-up growth after fetal protein restriction alters the adipose tissue gene expression program leading to obesity in adult mice

A mismatch between fetal and postnatal environment can permanently alter the body structure and physiology and therefore contribute later to obesity and related disorders, as revealed by epidemiological studies. Early programming of adipose tissue might be central in this observation. Moreover, adipose tissue secretes adipokines that provide a molecular link between obesity and its related disorders. Therefore, our aim was to investigate whether a protein restriction during fetal life, followed by catch-up growth could lead to obesity in 9-mo-old male mice and could alter the adipose tissue gene expression profile. Dams were fed a low-protein (LP) or an isocaloric control (C) diet during gestation. Postnatal catch-up growth was induced in LP offspring by feeding dams with control diet and by culling LP litters to four pups instead of eight in the C group. At weaning, male mice were fed by lab chow alone (C) or supplemented with a hypercaloric diet (HC), to induce obesity (C-C, C-HC, LP-C, and LP-HC groups). At 9 mo, LP offspring featured increased relative fat mass, hyperglycemia, hypercholesterolemia, and hyperleptinemia. Using a microarray designed to study the expression of 89 genes involved in adipose tissue differentiation/function, we demonstrated that the expression profile of several genes were dependent upon the maternal diet. Among the diverse genes showing altered expression, we could identify genes encoding several enzymes involved in lipid metabolism. These results indicated that offspring submitted to early mismatched nutrition exhibited alterations in adipose tissue gene expression that probably increases their susceptibility to overweight when challenged after weaning with a HC diet.

Intrauterine programming of the endocrine pancreas.

Epidemiological studies have revealed strong relationships between poor foetal growth and subsequent development of the metabolic syndrome. Persisting effects of early malnutrition become translated into pathology, thereby determine chronic risk for developing glucose intolerance and diabetes. These epidemiological observations identify the phenomena of foetal programming without explaining the underlying mechanisms that establish the causal link. Animal models have been established and studies have demonstrated that reduction in the availability of nutrients during foetal development programs the endocrine pancreas and insulin‐sensitive tissues. Whatever the type of foetal malnutrition, whether there are not enough calories or protein in food or after placental deficiency, malnourished pups are born with a defect in their β‐cell population that will never completely recover, and insulin‐sensitive tissues will be definitively altered. Despite the similar endpoint, different cellular and physiological mechanisms are proposed. Hormones operative during foetal life like insulin itself, insulin‐like growth factors and glucocorticoids, as well as specific molecules like taurine, or islet vascularization were implicated as possible factors amplifying the defect. The molecular mechanisms responsible for intrauterine programming of the β cells are still elusive, but two hypotheses recently emerged: the first one implies programming of mitochondria and the second, epigenetic regulation.

Postnatal Catch-up Growth After Fetal Protein Restriction Programs Proliferation of Rat Preadipocytes.

We studied the in vitro proliferation and differentiation of rat preadipocytes to investigate whether catch‐up growth after prenatal protein restriction may program adipose precursor cells leading to development of increased adipose tissue mass. Pregnant rat dams were fed either an isocaloric low‐protein diet (LP‐8%) or control diet (C‐20%). During lactation, in order to induce catch‐up growth, dams from LP group were fed with the C diet and litter size was reduced to four pups instead of eight. Preadipocytes were isolated from weanling male pups (28 days of age). Differentiation and proliferation were assessed across time. At late stages of preadipocyte differentiation, no difference was observed in lipid accumulation of C or LP cultures but the mRNA expression of leptin was enhanced in LP cells. At early stages of culture, a higher DNA and protein content accompanied by a higher rate of proliferation was measured in adipocytes from LP cultures. Moreover, the mRNA expression of cyclin D1 was increased in these cells whereas the expression of peroxisome proliferators‐activated receptor γ (PPARγ) and steroyl regulatory element binding protein (SREBP‐1c) was significantly reduced during early stages. The results suggest that prenatal exposure to a LP followed by rapid catch‐up growth is associated with a higher rate for proliferation in preadipocytes.

Developmental programming of adult obesity and cardiovascular disease in rodents by maternal nutrition imbalance.

Studies on fetal undernutrition have generated the hypothesis that fetal programming corresponds to an attempt of the fetus to adapt to adverse conditions encountered in utero. These adaptations would be beneficial if these conditions prevail later in life, but they become detrimental in the case of normal or plentiful nutrition and favor the appearance of the metabolic syndrome. In this article, the discussion is limited to the developmental programming of obesity and cardiovascular disorders caused by an early mismatched nutrition, particularly intrauterine growth retardation followed by postnatal catch-up growth. Selected data in humans are reviewed before evoking some mechanisms revealed or suggested by experiments in rodents. A variety of physiologic mechanisms are implicated in obesity programming, 2 of which are detailed. In some, but not all observations, hyperphagia resulting namely from perturbed development of the hypothalamic circuitry devoted to appetite regulation may contribute to obesity. Another contribution may be the developmental changes in the population of fat cell precursors in adipose tissue. Even if the link between obesity and cardiovascular disease is well established, alteration of blood pressure regulation may appear independently of obesity. A loss of diurnal variation in heart rate and blood pressure in adulthood has resulted from maternal undernutrition followed by postnatal overnutrition. Further research should clarify the effect of mismatched early nutrition on the development of brain centers regulating energy intake, energy expenditure, and circadian rhythms.

Does early mismatched nutrition predispose to hypertension and atherosclerosis, in male mice?

Background A link between early mismatched nutritional environment and development of components of the metabolic syndrome later in life has been shown in epidemiological and animal data. The aim of this study was to investigate whether an early mismatched nutrition produced by catch-up growth after fetal protein restriction could induce the appearance of hypertension and/or atherosclerosis in adult male mice. Methodology/Principal Findings Wild-type C57BL6/J or LDLr−/− dams were fed a low protein (LP) or a control (C) diet during gestation. Catch-up growth was induced in LP offspring by feeding dams with a control diet and by culling the litter to 4 pups against 8 in controls. At weaning, male mice were fed either standard chow or an obesogenic diet (OB), leading to 4 experimental groups. Blood pressure (BP) and heart rate (HR) were assessed in conscious unrestrained wild-type mice by telemetry. Atherosclerosis plaque area was measured in aortic root sections of LDLr−/− mice. We found that: (1) postnatal OB diet increased significantly BP (P<0.0001) and HR (P<0.008) in 3-month old OB-C and OB-LP offspring, respectively; (2) that maternal LP diet induced a significant higher BP (P<0.009) and HR (P<0.004) and (3) an altered circadian rhythm in addition to higher plasma corticosterone concentration in 9 months-old LP offspring; (4) that, although LP offspring showed higher plasma total cholesterol than control offspring, atherosclerosis assessed in aortic roots of 6-mo old mice featured increased plaque area due to OB feeding but not due to early mismatched nutrition. Conclusions/Significance These results indicate a long-term effect of early mismatched nutrition on the appearance of hypertension independently of obesity, while no effect on atherosclerosis was noticed at this age.

Biological Basis for Increased Sensitivity to Radiation Therapy in HPV-Positive Head and Neck Cancers

Although development of head and neck squamous cell carcinomas (HNSCCs) is commonly linked to the consumption of tobacco and alcohol, a link between human papillomavirus (HPV) infection and a subgroup of head and neck cancers has been established. These HPV-positive tumors represent a distinct biological entity with overexpression of viral oncoproteins E6 and E7. It has been shown in several clinical studies that HPV-positive HNSCCs have a more favorable outcome and greater response to radiotherapy. The reason for improved prognosis of HPV-related HNSCC remains speculative, but it could be owned to multiple factors. One hypothesis is that HPV-positive cells are intrinsically more sensitive to standard therapies and thus respond better to treatment. Another possibility is that HPV-positive tumors uniquely express viral proteins that induce an immune response during therapy that helps clear tumors and prevents recurrence. Here, we will review current evidence for the biological basis of increased radiosensitivity in HPV-positive HNSCC.

Potential role of hypoxia imaging using (18)F-FAZA PET to guide hypoxia-driven interventions (carbogen breathing or dose escalation) in radiation therapy.

BACKGROUND AND PURPOSE Hypoxia-driven intervention (oxygen manipulation or dose escalation) could overcome radiation resistance linked to tumor hypoxia. Here, we evaluated the value of hypoxia imaging using (18)F-FAZA PET to predict the outcome and guide hypoxia-driven interventions. MATERIAL AND METHODS Two hypoxic rat tumor models were used: rhabdomyosarcoma and 9L-glioma. For the irradiated groups, the animals were divided into two subgroups: breathing either room air or carbogen. (18)F-FAZA PET images were obtained just before the irradiation to monitor the hypoxic level of each tumor. Absolute pO2 were also measured using EPR oximetry. Dose escalation was used in Rhabdomyosarcomas. RESULTS For 9L-gliomas, a significant correlation between (18)F-FAZA T/B ratio and tumor growth delay was found; additionally, carbogen breathing dramatically improved the tumor response to irradiation. On the contrary, Rhabdomyosarcomas were less responsive to hyperoxic challenge. For that model, an increase in growth delay was observed using dose escalation, but not when combining irradiation with carbogen. CONCLUSIONS (18)F-FAZA uptake may be prognostic of outcome following radiotherapy and could assess the response of tumor to carbogen breathing. (18)F-FAZA PET may help to guide the hypoxia-driven intervention with irradiation: carbogen breathing in responsive tumors or dose escalation in tumors non-responsive to carbogen.

Reprogramming of tumor metabolism by targeting mitochondria improves tumor response to irradiation

Abstract Background. The Warburg phenotype identified decades ago describes tumor cells with increased glycolysis and decreased mitochondrial respiration even in the presence of oxygen. This particular metabolism also termed ‘aerobic glycolysis’ reflects an adaptation of tumor cells to proliferation in a heterogeneous tumor microenvironment. Although metabolic alterations in cancer cells are common features, their impact on the response to radiotherapy is not yet fully elucidated. This study investigated the impact of cellular oxygen consumption inhibition on the tumor response to radiotherapy. Material and methods. Warburg-phenotype tumor cells with impaired mitochondrial respiration (MD) were produced and compared in respect to their metabolism to the genetically matched parental cells (WT). After characterization of their metabolism we compared the response of MD cells to irradiation in vivo and in vitro to the genetically matched parental cells (WT). Results. We first confirmed that MD cells were exclusively glycolytic while WT cells exhibited mitochondrial respiration. We then used these cells for assessing the response of WT and MD tumors to a single dose of radiation and showed that the in vivo tumor growth delay of the MD group was increased, indicating an increased radiosensitivity compared to WT while the in vitro ability of both cell lines to repair radiation-induced DNA damage was similar. Conclusion. Taken together, these results indicate that in addition to intrinsic radiosensitivity parameters the tumor response to radiation will also depend on their metabolic rate of oxygen consumption.

Reducing the serine availability complements the inhibition of the glutamine metabolism to block leukemia cell growth.

Leukemia cells are described as a prototype of glucose-consuming cells with a high turnover rate. The role of glutamine in fueling the tricarboxylic acid cycle of leukemia cells was however recently identified confirming its status of major anaplerotic precursor in solid tumors. Here we examined whether glutamine metabolism could represent a therapeutic target in leukemia cells and whether resistance to this strategy could arise. We found that glutamine deprivation inhibited leukemia cell growth but also led to a glucose-independent adaptation maintaining cell survival. A proteomic study revealed that glutamine withdrawal induced the upregulation of phosphoglycerate dehydrogenase (PHGDH) and phosphoserine aminotransferase (PSAT), two enzymes of the serine pathway. We further documented that both exogenous and endogenous serine were critical for leukemia cell growth and contributed to cell regrowth following glutamine deprivation. Increase in oxidative stress upon inhibition of glutamine metabolism was identified as the trigger of the upregulation of PHGDH. Finally, we showed that PHGDH silencing in vitro and the use of serine-free diet in vivo inhibited leukemia cell growth, an effect further increased when glutamine metabolism was blocked. In conclusion, this study identified serine as a key pro-survival actor that needs to be handled to sensitize leukemia cells to glutamine-targeting modalities.

Digital pathology: elementary, rapid and reliable automated image analysis.

Slide digitalization has brought pathology to a new era, including powerful image analysis possibilities. However, while being a powerful prognostic tool, immunostaining automated analysis on digital images is still not implemented worldwide in routine clinical practice.