Normobaric hypoxia shows enhanced FOXO1 signaling in obese mouse gastrocnemius muscle linked to metabolism and muscle structure and neuromuscular innervation.

Skeletal muscle relies on mitochondria for sustainable ATP production, which may be impacted by reduced oxygen availability (hypoxia). Compared with long-term hypoxia, the mechanistic in vivo response to acute hypoxia remains elusive. Therefore, we aimed to provide an integrated description of the Musculus gastrocnemius response to acute hypoxia. Fasted male C57BL/6JOlaHsd mice, fed a 40en% fat diet for six weeks, were exposed to 12% O2 normobaric hypoxia or normoxia (20.9% O2) for six hours (n = 12 per group). Whole-body energy metabolism and the transcriptome response of the M. gastrocnemius were analyzed and confirmed by acylcarnitine determination and Q-PCR. At the whole-body level, six hours of hypoxia reduced energy expenditure, increased blood glucose and tended to decreased the respiratory exchange ratio (RER). Whole-genome transcriptome analysis revealed upregulation of forkhead box-O (FOXO) signalling, including an increased expression of tribbles pseudokinase 3 (Trib3). Trib3 positively correlated with blood glucose levels. Upregulated carnitine palmitoyltransferase 1A negatively correlated with the RER, but the significantly increased in tissue C14-1, C16-0 and C18-1 acylcarnitines supported that ß-oxidation was not regulated. The hypoxia-induced FOXO activation could also be connected to altered gene expression related to fiber-type switching, extracellular matrix remodeling, muscle differentiation and neuromuscular junction denervation. Our results suggest that a six-hour exposure of obese mice to 12% O2 normobaric hypoxia impacts M. gastrocnemius via FOXO1, initiating alterations that may contribute to muscle remodeling of which denervation is novel and warrants further investigation. The findings support an early role of hypoxia in tissue alterations in hypoxia-associated conditions such as aging and obesity.

Dietary Galactose Increases the Expression of Mitochondrial OXPHOS Genes and Modulates the Carbohydrate Oxidation Pathways in Mouse Intestinal Mucosa.

BACKGROUND: Prolonged lactation provides substantial health benefits, possibly because of galactose as part of milk sugar lactose. Isocaloric replacement of dietary glucose [16 energy%(en%)] with galactose within a normal diet (64en% carbohydrates) during a 3-wk postweaning period provided substantial benefits on short- and long-term physiologic and metabolic parameters at the whole-body level and liver in female mice, which might be attributable to intestinal function. OBJECTIVES: This study aimed to investigate if partial dietary replacement of glucose with galactose alters intestinal metabolism underlying hepatic health effects. METHODS: Proximal intestinal mucosa gene profiles in female mice were analyzed using RNAseq technology, validated, and correlated with hepatic health parameters. RESULTS: Transcriptome analysis revealed that the presence of galactose primarily affected the pathways involved in energy metabolism. A consistently higher expression was observed in the subset of mitochondrial transcripts (78 of 80, all P.adj < 0.1). Oxidative phosphorylation (OXPHOS) represented the most upregulated process (all top 10 pathways) independent of the total mitochondrial mass (P = 0.75). Moreover, galactose consistently upregulated carbohydrate metabolism pathways, specifically glycolysis till acetyl-CoA production and fructose metabolism. Also, the expression of transcripts involved in these pathways was negatively correlated with circulating serum amyloid A3 protein, a marker of hepatic inflammation [R (-0.61, -0.5), P (0.002, 0.01)]. Accordingly, CD163+ cells were decreased in the liver. Additionally, the expression of key fructolytic enzymes in the small intestinal mucosa was negatively correlated with triglyceride accumulation in the liver [R (-0.45, -0.4), P (0.03, 0.05)]. CONCLUSIONS: To our knowledge, our results show for the first time the role of galactose as an OXPHOS activator in vivo. Moreover, the concept of intestinal cells acting as the body's metabolic gatekeeper is strongly supported, as they alter substrate availability and thereby contribute to the maintenance of metabolic homeostasis, protecting other organs, as evidenced by their potential ability to shield the liver from the potentially detrimental effects of fructose.

Ndufs4 knockout mouse models of Leigh syndrome: pathophysiology and intervention.

Mitochondria are small cellular constituents that generate cellular energy (ATP) by oxidative phosphorylation (OXPHOS). Dysfunction of these organelles is linked to a heterogeneous group of multisystemic disorders, including diabetes, cancer, ageing-related pathologies and rare mitochondrial diseases. With respect to the latter, mutations in subunit-encoding genes and assembly factors of the first OXPHOS complex (complex I) induce isolated complex I deficiency and Leigh syndrome. This syndrome is an early-onset, often fatal, encephalopathy with a variable clinical presentation and poor prognosis due to the lack of effective intervention strategies. Mutations in the nuclear DNA-encoded NDUFS4 gene, encoding the NADH:ubiquinone oxidoreductase subunit S4 (NDUFS4) of complex I, induce 'mitochondrial complex I deficiency, nuclear type 1' (MC1DN1) and Leigh syndrome in paediatric patients. A variety of (tissue-specific) Ndufs4 knockout mouse models were developed to study the Leigh syndrome pathomechanism and intervention testing. Here, we review and discuss the role of complex I and NDUFS4 mutations in human mitochondrial disease, and review how the analysis of Ndufs4 knockout mouse models has generated new insights into the MC1ND1/Leigh syndrome pathomechanism and its therapeutic targeting.

Health relevance of lowering postprandial glycaemia in the paediatric population through diet': results from a multistakeholder workshop.

To summarize current knowledge and gaps regarding the role of postprandial glycaemic response in the paediatric population, a workshop was organized in June 2021 by the European branch of the International Life Science Institute (ILSI). This virtual event comprised of talks given by experts followed by in-depth discussions in breakout sessions with workshop participants. The main pre-specified topics addressed by the workshop organizing committee to the invited speakers and the workshop participants were: (1) the role of glycaemic responses for paediatric health, based on mechanistic insights from animal and human data, and long-term evidence from observational and intervention studies in paediatric populations, and (2) changes in metabolism and changes in dietary needs from infancy to adolescence. Each talk as well as the discussions were summarised, including the main identified research gaps. The workshop led to the consensus on the crucial role on health of postprandial glycaemic response in paediatric population. However, a lack of scientific data has been identified regarding detailed glucose and insulin profiles in response to foods commonly consumed by paediatric populations, as well as a lack of long-term evidence including the need for suitable predictors during childhood and adolescence to anticipate health effects during adulthood.

Galactose in the Post-Weaning Diet Programs Improved Circulating Adiponectin Concentrations and Skeletal Muscle Insulin Signaling.

Short-term post-weaning nutrition can result in long-lasting effects in later life. Partial replacement of glucose by galactose in the post-weaning diet showed direct effects on liver inflammation. Here, we examined this program on body weight, body composition, and insulin sensitivity at the adult age. Three-week-old female C57BL/6JRccHsd mice were fed a diet with glucose plus galactose (GAL; 16 energy% (en%) each) or a control diet with glucose (GLU; 32 en%) for three weeks, and afterward, both groups were given the same high-fat diet (HFD). After five weeks on a HFD, an oral glucose tolerance test was performed. After nine weeks on a HFD, energy metabolism was assessed by indirect calorimetry, and fasted mice were sacrificed fifteen minutes after a glucose bolus, followed by serum and tissue analyses. Body weight and body composition were not different between the post-weaning dietary groups, during the post-weaning period, or the HFD period. Glucose tolerance and energy metabolism in adulthood were not affected by the post-weaning diet. Serum adiponectin concentrations were significantly higher (p = 0.02) in GAL mice while insulin, leptin, and insulin-like growth factor 1 concentrations were not affected. Expression of Adipoq mRNA was significantly higher in gonadal white adipose tissue (gWAT; p = 0.03), while its receptors in the liver and skeletal muscles remained unaffected. Irs2 expression was significantly lower in skeletal muscles (p = 0.01), but not in gWAT or Irs1 expression (in both tissues). Gene expressions of inflammatory markers in gWAT and the liver were also not affected. Conclusively, galactose in the post-weaning diet significantly improved circulating adiponectin concentrations and reduced skeletal muscle Irs2 expression in adulthood without alterations in fat mass, glucose tolerance, and inflammation.

The macro-metastasis/organ parenchyma interface (MMPI) - A hitherto unnoticed area.

The macro-metastasis/organ parenchyma interface (MMPI) was previously considered an inert anatomical border which sharply separates the affected organ parenchyma from the macro-metastatic tissue. Recently, infiltrative growth of macro-metastases from various primary tumors was described in the brain, liver and lung, with significant impact on survival. Strikingly, the MMPI patterns differed between entities, so that at least nine different patterns were described. The MMPI patterns could be further classified into three major groups: displacing, epithelial and diffuse infiltrating. Additionally, macro-metastases are a source of further tumor cell dissemination in the affected organ; and these intra-organ metastatic dissemination tracks starting from the MMPI also vary depending on the anatomical structures of the colonized organ and influence disease outcome. In spite of their relevance, MMPIs and organ-specific dissemination tracks are still largely overlooked by many clinicians, pathologists and/or researchers. In this review, we aim to address this important issue and enhance our current understanding of the different MMPI patterns and dissemination tracks in the brain, liver and lung.

Exploring the associations between transcript levels and fluxes in constraint-based models of metabolism.

BACKGROUND: Several computational methods have been developed that integrate transcriptomics data with genome-scale metabolic reconstructions to increase accuracy of inferences of intracellular metabolic flux distributions. Even though existing methods use transcript abundances as a proxy for enzyme activity, each method uses a different hypothesis and assumptions. Most methods implicitly assume a proportionality between transcript levels and flux through the corresponding function, although these proportionality constant(s) are often not explicitly mentioned nor discussed in any of the published methods. E-Flux is one such method and, in this algorithm, flux bounds are related to expression data, so that reactions associated with highly expressed genes are allowed to carry higher flux values. RESULTS: Here, we extended E-Flux and systematically evaluated the impact of an assumed proportionality constant on model predictions. We used data from published experiments with Escherichia coli and Saccharomyces cerevisiae and we compared the predictions of the algorithm to measured extracellular and intracellular fluxes. CONCLUSION: We showed that detailed modelling using a proportionality constant can greatly impact the outcome of the analysis. This increases accuracy and allows for extraction of better physiological information.

Identification of blood cell transcriptome-based biomarkers in adulthood predictive of increased risk to develop metabolic disorders using early life intervention rat models.

Calorie restriction during gestation in rats has long-lasting adverse effects in the offspring. It induces metabolic syndrome-related alterations, which are partially reversed by leptin supplementation during lactation. We employed these conditions to identify transcript-based nutrient sensitive biomarkers in peripheral blood mononuclear cells (PBMCs) predictive of later adverse metabolic health. The best candidate was validated in humans. Transcriptome analysis of PBMCs from adult male Wistar rats of three experimental groups was performed: offspring of control dams (CON), and offspring of 20% calorie-restricted dams during gestation without (CR) and with leptin supplementation throughout lactation (CR-LEP). The expression of 401 genes was affected by gestational calorie restriction and reversed by leptin. The changes preceded metabolic syndrome-related phenotypic alterations. Of these genes, Npc1 mRNA levels were lower in CR vs CON, and normalized to CON in CR-LEP. In humans, NPC1 mRNA levels in peripheral blood cells (PBCs) were decreased in subjects with mildly impaired metabolic health compared to healthy subjects. Therefore, a set of potential transcript-based biomarkers indicative of a predisposition to metabolic syndrome-related alterations were identified, including NPC1, which was validated in humans. Low NPC1 transcript levels in PBCs are a candidate biomarker of increased risk for impaired metabolic health in humans.

Complexity of PEComas : Diagnostic approach, molecular background, clinical management.

Perivascular epithelioid cell neoplasms (PEComas) are a family of mesenchymal neoplasms with features of both melanotic and smooth muscle differentiation. PEComa morphology is highly variable and encompasses epithelioid to spindle cells often with clear cytoplasm and prominent nucleoli. Molecularly, most PEComas are defined by a loss of function of the TSC1/TSC2 complex. Additionally, a distinct small subset of PEComas harboring rearrangements of the TFE3 (Xp11) gene locus has been identified. By presenting a series of three case reports with distinct features, we demonstrate diagnostic pitfalls as well as the importance of molecular work-up of PEComas because of important therapeutic consequences.

Replacing Part of Glucose with Galactose in the Postweaning Diet Protects Female But Not Male Mice from High-Fat Diet-Induced Adiposity in Later Life.

BACKGROUND: Duration of breastfeeding is positively associated with decreased adiposity and increased metabolic health in later life, which might be related to galactose. OBJECTIVE: The aim of this study was to investigate if partial replacement of glucose with galactose in the postweaning diet had a metabolic programming effect. METHODS: Male and female mice (C57BL/6JRccHsd) received an isocaloric diet (16 energy% fat; 64 energy% carbohydrates; 20 energy% protein) with either glucose (32 energy%) (GLU) or glucose + galactose (GLU + GAL, 16 energy% each) for 3 wk postweaning. Afterwards, all mice were switched to the same 40 energy% high-fat diet (HFD) for 9 wk to evaluate potential programming effects in an obesogenic environment. Data were analyzed within sex. RESULTS: Female body weight (-14%) and fat mass (-47%) were significantly lower at the end of the HFD period (both P < 0.001) among those fed GLU + GAL than among those fed GLU; effects in males were in line with these findings but nonsignificant. Food intake was affected in GLU + GAL-fed females (+8% on postweaning diet, -9% on HFD) compared with GLU-fed females, but not for hypothalamic transcript levels at endpoint. Also, in GLU + GAL-fed females, serum insulin concentrations (-48%, P  < 0.05) and the associated homeostasis model assessment of insulin resistance (HOMA-IR) were significantly lower ( P < 0.05) at endpoint, but there were no changes in pancreas morphology. In GLU + GAL-fed females, expression of insulin receptor substrate 2 (Irs2) (-27%, P  < 0.01 ; -44%, P  < 0.001) and the adipocyte size markers leptin (Lep) (-40%, P  < 0.05; -63% , P  < 0.05) and mesoderm-specific transcript homolog protein (Mest) (-80%, P < 0.05; -72%, P  < 0.05) was lower in gonadal and subcutaneous white adipose tissue (WAT), respectively. Expression of insulin receptor substrate1 (Irs1) (-24%, P  < 0.05) was only lower in subcutaneous WAT in GLU + GAL-fed females. CONCLUSIONS: Partial replacement of glucose with galactose, resulting in a 1:1 ratio mimicking lactose, in a 3-wk postweaning diet lowered body weight, adiposity, HOMA-IR, and expression of WAT insulin signaling in HFD-challenged female mice in later life. This suggests that prolonged galactose intake may improve metabolic and overall health in later life.

[Round-cell sarcomas]. / Rundzellige Sarkome.

Round-cell sarcomas represent highly malignant tumors that occur predominantly in children, adolescents, and young adults. Round-cell sarcomas are caused by recurrent translocations that involve certain transcription factors. Ewing's sarcoma, Ewing-like sarcomas (e.g. CIC-DUX positive or BCOR positive sarcomas), desmoplastic small round-cell tumors (DSRCTs), and alveolar rhabdomyosarcomas (ARMs) are typical examples of this particular group of sarcomas. These entities differ in their tumor genetics, which is correlated with immunohistochemical expression profiles and with clinical phenotypes. Classification should be based on molecular findings. Immunohistochemistry may serve as a surrogate marker.

[Epithelioid, biphasic and mixed tumors of soft tissue]. / Epitheloide, biphasische und Mischtumoren des Weichgewebes.

Epithelioid soft tissue tumors consist exclusively of epithelioid tumor cells. Biphasic tumors are composed of both a spindle-cell and an epithelioid component. The rare mixed tumors of soft tissue show a broader variety of cellular and stromal differentiation but also include at least one, possibly several, epithelioid portions.The close morphological similarity of some of these entities with each other, as well as with the more frequent soft tissue metastases of carcinomas, carcinosarcomas, and melanomas, to malignant mesothelioma and certain lymphomas, can often make the correct diagnosis extremely difficult. Recent advances in the detection of certain molecular alterations (mostly chromosomal translocations) have contributed to changes in tumor classification but also to improved pathological diagnostics (e.g. through the development of potent diagnostic antibodies) and biological understanding.The present overview should help the pathologist in the diagnosis of these rare tumors through the classical approach of morphological pattern recognition. The most important entities are discussed and illustrated in more detail, with the incorporation of the latest immunohistochemical and molecular aspects and the differential diagnosis of similar tumors.

[Successful pembrolizumab therapy in metastasized adenosquamous carcinoma of the colon]. / Erfolgreiche Pembrolizumab-Therapie bei metastasiertem adenosquamösem Karzinom des Kolons.

Adenosquamous carcinoma (ASqC) is an exceedingly rare subtype of colorectal cancer without any known special guidelines for treatment. The biological behaviour and molecular background are widely unknown, although a few case studies report a worse prognosis compared to ordinary colorectal adenocarcinoma. We herein report for the first time the successful immune checkpoint inhibitor therapy in a 40-year-old patient suffering from metastasized right-sided colonic ASqC with unique molecular features, after having previously progressed under standard chemotherapy.

[Complexity of PEComas : Diagnostic approach, molecular background, clinical management (German version)]. / Komplexität von PEComen : Diagnose, Molekulargenetik, klinisches Management.

Perivascular epithelioid cell neoplasms (PEComas) are a family of mesenchymal neoplasms with features of both melanotic and smooth muscle differentiation. PEComa morphology is highly variable and encompasses epithelioid to spindle cells often with clear cytoplasm and prominent nucleoli. Molecularly, most PEComas are defined by a loss of function of the TSC1/TSC2 complex. Additionally, a distinct small subset of PEComas harboring rearrangements of the TFE3 (Xp11) gene locus has been identified. By presenting a series of three case reports with distinct features, we demonstrate diagnostic pitfalls as well as the importance of molecular work-up of PEComas because of important therapeutic consequences.

Discrete Organic Phosphorus Signatures are Evident in Pollutant Sources within a Lake Erie Tributary.

Phosphorus loads are strongly associated with the severity of harmful algal blooms in Lake Erie, a Great Lake situated between the United States and Canada. Inorganic and total phosphorus measurements have historically been used to estimate nonpoint and point source contributions, from contributing watersheds with organic phosphorus often neglected. Here, we used ultrahigh resolution mass spectrometry to characterize the dissolved organic matter and specifically dissolved organic phosphorus composition of several nutrient pollutant source materials and aqueous samples in a Lake Erie tributary. We detected between 23 and 313 organic phosphorus formulas across our samples, with manure samples having greater abundance of phosphorus- and nitrogen containing compounds compared to other samples. Manures also were enriched in lipids and protein-like compounds. The greatest similarities were observed between the Sandusky River and wastewater treatment plant effluent (WWTP), or the Sandusky River and agricultural edge of field samples. These sample pairs shared 84% of organic compounds and 59-73% of P-containing organic compounds, respectively. This similarity suggests that agricultural and/or WWTP sources dominate the supply of organic phosphorus compounds to the river. We identify formulas shared between the river and pollutant sources that could serve as possible markers of source contamination in the tributary.

Screening-relevant age threshold of 70 years and older is a stronger determinant for the choice of adjuvant treatment in breast cancer patients than tumor biology.

PURPOSE: The 70-year threshold determines whether patients are eligible or not for the breast cancer screening program in Germany. It is not known whether this age threshold also influences the choice of adjuvant treatment and ultimate outcome. METHODS: 3463 patients were analyzed from the clinical cancer registry Regensburg (Germany) with primary, non-metastatic invasive breast cancer diagnosed between 2000 and 2012. The distribution of tumor biological subtypes was evaluated in breast cancer patients both in those eligible for screening (ESG, 50-69 years) and those not eligible for screening (NESG, ≥70 years). Local and systemic therapies in different subtypes as well as overall survival (OS) were analyzed. RESULTS: 2171 patients (62.7%) pertained to the ESG and 1292 patients (37.3%) referred to the NESG. The distribution of the common subtypes Luminal A, Luminal B, HER2-like, and Basal-like was comparable in both groups. Treatment varied considerably with less systemic therapies in all subtypes in patients in the NESG. Regarding local therapies, patients in the NESG also received less surgery and less radiotherapy. As to Luminal A patients, best OS was seen in patients receiving endocrine therapy (ET) (7-year OS of 95.6%) and CHT plus ET (7-year OS of 93.1%) in the ESG. In the NESG, best OS was seen in patients receiving CHT plus ET (7-year OS of 95.2%), whereas patients receiving only ET had a 7-year OS of 73.9%. CONCLUSIONS: Despite similar tumor biology, elderly patients are undertreated regarding both systemic and local therapies compared to younger patients, leading to reduced OS.

Basophils Trigger Fibroblast Activation in Cardiac Allograft Fibrosis Development.

Fibrosis is a major component of chronic cardiac allograft rejection. Although several cell types are able to produce collagen, resident (donor-derived) fibroblasts are mainly responsible for excessive production of extracellular matrix proteins. It is currently unclear which cells regulate production of connective tissue elements in allograft fibrosis and how basophils, as potential producers of profibrotic cytokines, are involved this process. We studied this question in a fully MHC-mismatched model of heart transplantation with transient depletion of CD4(+) T cells to largely prevent acute rejection. The model is characterized by myocardial infiltration of leukocytes and development of interstitial fibrosis and allograft vasculopathy. Using depletion of basophils, IL-4-deficient recipients and IL-4 receptor-deficient grafts, we showed that basophils and IL-4 play crucial roles in activation of fibroblasts and development of fibrotic organ remodeling. In the absence of CD4(+) T cells, basophils are the predominant source of IL-4 in the graft and contribute to expansion of myofibroblasts, interstitial deposition of collagen and development of allograft vasculopathy. Our results indicated that basophils trigger the production of various connective tissue elements by myofibroblasts. Basophil-derived IL-4 may be an attractive target for treatment of chronic allograft rejection.

Muscle mitohormesis promotes cellular survival via serine/glycine pathway flux.

Recent studies on mouse and human skeletal muscle (SM) demonstrated the important link between mitochondrial function and the cellular metabolic adaptation. To identify key compensatory molecular mechanisms in response to chronic mitochondrial distress, we analyzed mice with ectopic SM respiratory uncoupling in uncoupling protein 1 transgenic (UCP1-TG) mice as model of muscle-specific compromised mitochondrial function. Here we describe a detailed metabolic reprogramming profile associated with mitochondrial perturbations in SM, triggering an increased protein turnover and amino acid metabolism with induced biosynthetic serine/1-carbon/glycine pathway and the longevity-promoting polyamine spermidine as well as the trans-sulfuration pathway. This is related to an induction of NADPH-generating pathways and glutathione metabolism as an adaptive mitohormetic response and defense against increased oxidative stress. Strikingly, consistent muscle retrograde signaling profiles were observed in acute stress states such as muscle cell starvation and lipid overload, muscle regeneration, and heart muscle inflammation, but not in response to exercise. We provide conclusive evidence for a key compensatory stress-signaling network that preserves cellular function, oxidative stress tolerance, and survival during conditions of increased SM mitochondrial distress, a metabolic reprogramming profile so far only demonstrated for cancer cells and heart muscle.