Maternal emulsifier consumption programs offspring metabolic and neuropsychological health in mice.

Modern lifestyle is associated with a major consumption of ultra-processed foods (UPF) due to their practicality and palatability. The ingestion of emulsifiers, a main additive in UPFs, has been related to gut inflammation, microbiota dysbiosis, adiposity, and obesity. Maternal unbalanced nutritional habits during embryonic and perinatal stages perturb offspring's long-term metabolic health, thus increasing obesity and associated comorbidity risk. However, whether maternal emulsifier consumption influences developmental programming in the offspring remains unknown. Here, we show that, in mice, maternal consumption of dietary emulsifiers (1% carboxymethyl cellulose (CMC) and 1% P80 in drinking water), during gestation and lactation, perturbs the development of hypothalamic energy balance regulation centers of the progeny, leads to metabolic impairments, cognition deficits, and induces anxiety-like traits in a sex-specific manner. Our findings support the notion that maternal consumption of emulsifiers, common additives of UPFs, causes mild metabolic and neuropsychological malprogramming in the progeny. Our data call for nutritional advice during gestation.

Negative energy balance hinders prosocial helping behavior.

The internal state of an animal, including homeostatic requirements, modulates its behavior. Negative energy balance stimulates hunger, thus promoting a range of actions aimed at obtaining food. While these survival actions are well established, the influence of the energy status on prosocial behavior remains unexplored. We developed a paradigm to assess helping behavior in which a free mouse was faced with a conspecific trapped in a restrainer. We measured the willingness of the free mouse to liberate the confined mouse under diverse metabolic conditions. Around 42% of ad libitum-fed mice exhibited a helping behavior, as evidenced by the reduction in the latencies to release the trapped cagemate. This behavior was independent of subsequent social contact reward and was associated with changes in corticosterone indicative of emotional contagion. This decision-making process was coupled with reduced blood glucose excursions and higher Adenosine triphosphate (ATP):Adenosine diphosphate (ADP) ratios in the forebrain of helper mice, suggesting that it was a highly energy-demanding process. Interestingly, chronic (food restriction and type 2 diabetes) and acute (chemogenetic activation of hunger-promoting AgRP neurons) situations mimicking organismal negative energy balance and enhanced appetite attenuated helping behavior toward a distressed conspecific. To investigate similar effects in humans, we estimated the influence of glycated hemoglobin (a surrogate of long-term glycemic control) on prosocial behavior (namely charity donation) using the Understanding Society dataset. Our results evidenced that organismal energy status markedly influences helping behavior and that hypothalamic AgRP neurons are at the interface of metabolism and prosocial behavior.

Food craving-like episodes during pregnancy are mediated by accumbal dopaminergic circuits.

Preparation for motherhood requires a myriad of physiological and behavioural adjustments throughout gestation to provide an adequate environment for proper embryonic development1. Cravings for highly palatable foods are highly prevalent during pregnancy2 and contribute to the maintenance and development of gestational overweight or obesity3. However, the neurobiology underlying the distinct ingestive behaviours that result from craving specific foods remain unknown. Here we show that mice, similarly to humans, experience gestational food craving-like episodes. These episodes are associated with a brain connectivity reorganization that affects key components of the dopaminergic mesolimbic circuitry, which drives motivated appetitive behaviours and facilitates the perception of rewarding stimuli. Pregnancy engages a dynamic modulation of dopaminergic signalling through neurons expressing dopamine D2 receptors in the nucleus accumbens, which directly modulate food craving-like events. Importantly, persistent maternal food craving-like behaviour has long-lasting effects on the offspring, particularly in males, leading to glucose intolerance, increased body weight and increased susceptibility to develop eating disorders and anxiety-like behaviours during adulthood. Our results reveal the cognitively motivated nature of pregnancy food cravings and advocates for moderating emotional eating during gestation to prevent deterioration of the offspring's neuropsychological and metabolic health.

Hypothalamic pregnenolone mediates recognition memory in the context of metabolic disorders.

Obesity and type 2 diabetes are associated with cognitive dysfunction. Because the hypothalamus is implicated in energy balance control and memory disorders, we hypothesized that specific neurons in this brain region are at the interface of metabolism and cognition. Acute obesogenic diet administration in mice impaired recognition memory due to defective production of the neurosteroid precursor pregnenolone in the hypothalamus. Genetic interference with pregnenolone synthesis by Star deletion in hypothalamic POMC, but not AgRP neurons, deteriorated recognition memory independently of metabolic disturbances. Our data suggest that pregnenolone's effects on cognitive function were mediated via an autocrine mechanism on POMC neurons, influencing hippocampal long-term potentiation. The relevance of central pregnenolone on cognition was also confirmed in metabolically unhealthy patients with obesity. Our data reveal an unsuspected role for POMC neuron-derived neurosteroids in cognition. These results provide the basis for a framework to investigate new facets of POMC neuron biology with implications for cognitive disorders.

Mitochondrial cristae-remodeling protein OPA1 in POMC neurons couples Ca2+ homeostasis with adipose tissue lipolysis.

Appropriate cristae remodeling is a determinant of mitochondrial function and bioenergetics and thus represents a crucial process for cellular metabolic adaptations. Here, we show that mitochondrial cristae architecture and expression of the master cristae-remodeling protein OPA1 in proopiomelanocortin (POMC) neurons, which are key metabolic sensors implicated in energy balance control, is affected by fluctuations in nutrient availability. Genetic inactivation of OPA1 in POMC neurons causes dramatic alterations in cristae topology, mitochondrial Ca2+ handling, reduction in alpha-melanocyte stimulating hormone (α-MSH) in target areas, hyperphagia, and attenuated white adipose tissue (WAT) lipolysis resulting in obesity. Pharmacological blockade of mitochondrial Ca2+ influx restores α-MSH and the lipolytic program, while improving the metabolic defects of mutant mice. Chemogenetic manipulation of POMC neurons confirms a role in lipolysis control. Our results unveil a novel axis that connects OPA1 in POMC neurons with mitochondrial cristae, Ca2+ homeostasis, and WAT lipolysis in the regulation of energy balance.

Cell Plasticity-Related Phenotypes and Taxanes Resistance in Castration-Resistant Prostate Cancer.

The prostatic tumor cells plasticity is involved in resistance to hormone-therapy, allowing these cells to survive despite androgen receptor inhibition. However, its role in taxanes resistance has not been fully established. Gene expression of plasticity-related phenotypes such as epithelial-mesenchymal transition (EMT), stem cell-like and neuroendocrine (NE) phenotypes was studied in vitro, in silico, in circulating tumor cells (CTCs) (N=22) and in tumor samples (N=117) from taxanes-treated metastatic castration-resistant prostate cancer (mCRPC) patients. Docetaxel (D)-resistant cells presented a more pronounced EMT phenotype than cabazitaxel (CZ)-resistant cells. In silico analysis revealed ESRP1 down-regulation in taxane-exposed mCRPC samples. Cell plasticity-related changes occurred in CTCs after taxanes treatment. Tumor EMT phenotype was associated with lower PSA progression-free survival (PFS) to D (P<0.001), and better to CZ (P=0.002). High ESRP1 expression was independently associated with longer PSA-PFS (P<0.001) and radiologic-PFS (P=0.001) in D and shorter PSA-PFS in the CZ cohort (P=0.041). High SYP expression was independently associated with lower PSA-PFS in D (P=0.003) and overall survival (OS) in CZ (P=0.002), and high EZH2 expression was associated with adverse OS in D-treated patients (P=0.013). In conclusion, EMT profile in primary tumor is differentially associated with D or CZ benefit and NE dedifferentiation correlates with adverse taxanes clinical outcome.

Pro-opiomelanocortin (POMC) neuron translatome signatures underlying obesogenic gestational malprogramming in mice.

OBJECTIVE: Maternal unbalanced nutritional habits during embryonic development and perinatal stages perturb hypothalamic neuronal programming of the offspring, thus increasing obesity-associated diabetes risk. However, the underlying molecular mechanisms remain largely unknown. In this study we sought to determine the translatomic signatures associated with pro-opiomelanocortin (POMC) neuron malprogramming in maternal obesogenic conditions. METHODS: We used the RiboTag mouse model to specifically profile the translatome of POMC neurons during neonatal (P0) and perinatal (P21) life and its neuroanatomical, functional, and physiological consequences. RESULTS: Maternal high-fat diet (HFD) exposure did not interfere with offspring's hypothalamic POMC neuron specification, but significantly impaired their spatial distribution and axonal extension to target areas. Importantly, we established POMC neuron-specific translatome signatures accounting for aberrant neuronal development and axonal growth. These anatomical and molecular alterations caused metabolic dysfunction in early life and adulthood. CONCLUSIONS: Our study provides fundamental insights on the molecular mechanisms underlying POMC neuron malprogramming in obesogenic contexts.

Androgen Receptor and Its Splicing Variant 7 Expression in Peripheral Blood Mononuclear Cells and in Circulating Tumor Cells in Metastatic Castration-Resistant Prostate Cancer.

Androgen receptor (AR) signaling remains crucial in castration-resistant prostate cancer (CRPC). Since it is also essential in immune cells, we studied whether the expression of AR full-length (ARFL) and its splicing variant ARV7 in peripheral blood mononuclear cells (PBMC) predicts systemic treatment response in mCRPC in comparison with circulating-tumor cells (CTC). We measured ARFL and ARV7 mRNA in PBMC and CTC from patients prior to receiving abiraterone (AA), enzalutamide (E), or taxanes by a pre-amplification plus quantitative reverse-transcription PCR. They were also tested in LNCaP-ARV7-transfected and in 22RV1 docetaxel-resistant (22RV1DR) cells. We studied 171 PBMC from 136 patients and from 24 non-cancer controls, and 47 CTC from 22 patients. High PBMC ARV7 levels correlated with worse AA/E and better taxane response. In taxane-treated patients high PBMC ARFL also correlated with longer progression-free survival (PFS). High ARV7 and ARFL expression were independently associated with better biochemical-PFS. Conversely, high CTC ARV7 and ARFL correlated with shorter radiological-PFS and overall survival, respectively. High ARV7 in 22RV1DR and LNCaP-ARV7 cells correlated with taxane resistance. In conclusion, ARFL and ARV7 at PBMC or CTC have a different predictive role in the taxane response, suggesting a potential influence of the AR pathway from PBMC in such response modulation.

The influence of treatment sequence in the prognostic value of TMPRSS2-ERG as biomarker of taxane resistance in castration-resistant prostate cancer.

TMPRSS2-ERG expression in blood has been correlated with low docetaxel benefit in metastatic castration-resistant prostate cancer (mCRPC). This multicenter study aimed to prospectively asses its role as a taxane-resistance biomarker in blood and retrospectively in tumors, exploring also the impact of prior abiraterone/enzalutamide (A/E) in patients and in vitro. TMPRSS2-ERG was tested by quantitative reverse-transcription PCR. We included 204 patients (137 blood and 124 tumor samples) treated with taxanes. TMPRSS2-ERG expression was correlated with prostate-specific antigen (PSA)-progression-free survival (PFS), radiological-PFS (RX-PFS), and overall survival (OS). Independent association with survival was evaluated by multivariate Cox modeling. In vitro ERG knockdown and combinatorial and sequential experiments with enzalutamide and docetaxel were performed in VCaP cells. Prior A/E (HR 1.8, 95% CI 1.2-2.8) and blood TMPRSS2-ERG detection (HR 2, 95% CI 1.1-3.7) were independently associated to lower PSA-PFS. In patients without prior A/E, blood and tumor TMPRSS2-ERG independently predicted lower PSA-PFS (HR 3.3, 95% CI 1.4-7.9 and HR 1.8, 95% CI 1.02-3.3, respectively) to taxanes. When prior A/E was administered, TMPRSS2-ERG was not associated with outcome. There was a significant interaction between blood TMPRSS2-ERG and prior A/E related to PSA-PFS (p = 0.032) and RX-PFS (p = 0.009). In vitro stable ERG inhibition did not sensitize VCaP cells to docetaxel. Concomitant enzalutamide and taxanes were synergistic, but prior enzalutamide reduced docetaxel cytotoxicity in VCaP cells. Enzalutamide induced the expression of neuroendocrine markers and reduced that of E-cadherin. We conclude that prior hormone-therapy may influence taxanes response and TMPRSS2-ERG prognostic value. Thus, multiple and sequential biomarkers are needed in CRPC follow-up evaluation.

Endothelial cell rearrangements during vascular patterning require PI3-kinase-mediated inhibition of actomyosin contractility.

Angiogenesis is a dynamic process relying on endothelial cell rearrangements within vascular tubes, yet the underlying mechanisms and functional relevance are poorly understood. Here we show that PI3Kα regulates endothelial cell rearrangements using a combination of a PI3Kα-selective inhibitor and endothelial-specific genetic deletion to abrogate PI3Kα activity during vessel development. Quantitative phosphoproteomics together with detailed cell biology analyses in vivo and in vitro reveal that PI3K signalling prevents NUAK1-dependent phosphorylation of the myosin phosphatase targeting-1 (MYPT1) protein, thereby allowing myosin light chain phosphatase (MLCP) activity and ultimately downregulating actomyosin contractility. Decreased PI3K activity enhances actomyosin contractility and impairs junctional remodelling and stabilization. This leads to overstretched endothelial cells that fail to anastomose properly and form aberrant superimposed layers within the vasculature. Our findings define the PI3K/NUAK1/MYPT1/MLCP axis as a critical pathway to regulate actomyosin contractility in endothelial cells, supporting vascular patterning and expansion through the control of cell rearrangement.

Therapeutic Benefit of Selective Inhibition of p110α PI3-Kinase in Pancreatic Neuroendocrine Tumors.

PURPOSE: Mutations in the PI3K pathway occur in 16% of patients with pancreatic neuroendocrine tumors (PanNETs), which suggests that these tumors are an exciting setting for PI3K/AKT/mTOR pharmacologic intervention. Everolimus, an mTOR inhibitor, is being used to treat patients with advanced PanNETs. However, resistance to mTOR-targeted therapy is emerging partially due to the loss of mTOR-dependent feedback inhibition of AKT. In contrast, the response to PI3K inhibitors in PanNETs is unknown. EXPERIMENTAL DESIGN: In the current study, we assessed the frequency of PI3K pathway activation in human PanNETs and in RIP1-Tag2 mice, a preclinical tumor model of PanNETs, and we investigated the therapeutic efficacy of inhibiting PI3K in RIP1-Tag2 mice using a combination of pan (GDC-0941) and p110α-selective (GDC-0326) inhibitors and isoform-specific PI3K kinase-dead-mutant mice. RESULTS: Human and mouse PanNETs showed enhanced pAKT, pPRAS40, and pS6 positivity compared with normal tissue. Although treatment of RIP1-Tag2 mice with GDC-0941 led to reduced tumor growth with no impact on tumor vessels, the selective inactivation of the p110α PI3K isoform, either genetically or pharmacologically, reduced tumor growth as well as vascular area. Furthermore, GDC-0326 reduced the incidence of liver and lymph node metastasis compared with vehicle-treated mice. We also demonstrated that tumor and stromal cells are implicated in the antitumor activity of GDC-0326 in RIP1-Tag2 tumors. CONCLUSIONS: Our data provide a rationale for p110α-selective intervention in PanNETs and unravel a new function of this kinase in cancer biology through its role in promoting metastasis. Clin Cancer Res; 22(23); 5805-17. ©2016 AACR.

Somatic activating mutations in Pik3ca cause sporadic venous malformations in mice and humans.

Venous malformations (VMs) are painful and deforming vascular lesions composed of dilated vascular channels, which are present from birth. Mutations in the TEK gene, encoding the tyrosine kinase receptor TIE2, are found in about half of sporadic (nonfamilial) VMs, and the causes of the remaining cases are unknown. Sclerotherapy, widely accepted as first-line treatment, is not fully efficient, and targeted therapy for this disease remains underexplored. We have generated a mouse model that faithfully mirrors human VM through mosaic expression of Pik3ca(H1047R), a constitutively active mutant of the p110α isoform of phosphatidylinositol 3-kinase (PI3K), in the embryonic mesoderm. Endothelial expression of Pik3ca(H1047R)resulted in endothelial cell (EC) hyperproliferation, reduction in pericyte coverage of blood vessels, and decreased expression of arteriovenous specification markers. PI3K pathway inhibition with rapamycin normalized EC hyperproliferation and pericyte coverage in postnatal retinas and stimulated VM regression in vivo. In line with the mouse data, we also report the presence of activating PIK3CA mutations in human VMs, mutually exclusive with TEK mutations. Our data demonstrate a causal relationship between activating Pik3ca mutations and the genesis of VMs, provide a genetic model that faithfully mirrors the normal etiology and development of this human disease, and establish the basis for the use of PI3K-targeted therapies in VMs.