Mammary duct luminal epithelium controls adipocyte thermogenic programme.

Sympathetic activation during cold exposure increases adipocyte thermogenesis via the expression of mitochondrial protein uncoupling protein 1 (UCP1)1. The propensity of adipocytes to express UCP1 is under a critical influence of the adipose microenvironment and varies between sexes and among various fat depots2-7. Here we report that mammary gland ductal epithelial cells in the adipose niche regulate cold-induced adipocyte UCP1 expression in female mouse subcutaneous white adipose tissue (scWAT). Single-cell RNA sequencing shows that glandular luminal epithelium subtypes express transcripts that encode secretory factors controlling adipocyte UCP1 expression under cold conditions. We term these luminal epithelium secretory factors 'mammokines'. Using 3D visualization of whole-tissue immunofluorescence, we reveal sympathetic nerve-ductal contact points. We show that mammary ducts activated by sympathetic nerves limit adipocyte UCP1 expression via the mammokine lipocalin 2. In vivo and ex vivo ablation of mammary duct epithelium enhance the cold-induced adipocyte thermogenic gene programme in scWAT. Since the mammary duct network extends throughout most of the scWAT in female mice, females show markedly less scWAT UCP1 expression, fat oxidation, energy expenditure and subcutaneous fat mass loss compared with male mice, implicating sex-specific roles of mammokines in adipose thermogenesis. These results reveal a role of sympathetic nerve-activated glandular epithelium in adipocyte UCP1 expression and suggest that mammary duct luminal epithelium has an important role in controlling glandular adiposity.

Overexpression of aryl hydrocarbon receptor (AHR) signalling pathway in human meningioma.

Aryl hydrocarbon receptor (AHR) is a ligand activated transcription factor and involved in tumorigenesis of many cancers. However there are no reports on AHR in human meningioma. Therefore we examined the status of the AHR and its signalling molecules in human meningioma by using tumor biopsy samples and autopsy control meninges. We report the up regulation of AHR pathway genes like aryl hydrocarbon receptor nuclear translocator (ARNT), aldehyde dehydrogenase1family memberA3 (ALDH1A3), cytochrome P450, family1, subfamily A polypeptide1 (CYP1A1) and TCCD induced poly ADP ribose polymerase (TIPARP) gene expression in human meningioma. Further, AHR protein expression was found to be up regulated in all grades of human meningioma. We found that AHR localized in the nucleus for high grade anaplastic meningioma through immunohistochemical analysis. Since AHR signalling pathway was known to involve in inhibition of apoptosis in cancer cells, we evaluated the cyclophilin D levels which maintains mitochondrial permeability transition pore a critical event during apoptosis. We report that cyclophilin D levels were upregulated in all grades of human meningioma compared to control meninges. Finally we also evaluated c-Fos protein levels as its levels were regulated by AHR. Here we report that c-Fos protein levels were down regulated in all grades of human meningioma compared to control meninges. To sum-up we found that AHR signalling pathway components were upregulated, as the grade of the meningioma progresses from low to high grade, suggesting an important role of AHR signalling pathway in human meningioma.

Altered tryptophan metabolism in human meningioma.

Meningiomas are the neoplasms that arise from the arachnoid cells of the meninges. It was reported that cancer cells escape from immune system through the metabolism of an aromatic essential amino acid tryptophan (TRP) via Kynurenine (KYN) pathway. However, the role of TRP metabolites such as, 5-Hydroxy tryptophan (5-HTP), 5-Hydroxy tryptamine (5-HT), N-acetyl serotonin (NAS), Melatonin (MEL), KYN, N-acetyl tryptamine, 5-Hydroxy indole acetic acid (5-HIAA) and 5-Methoxy indole acetic acid is not yet evaluated in human meningioma. Therefore, in the current study we have evaluated the levels of TRP and its metabolites in the progression of human meningioma using tumor biopsy samples and autopsy control meninges with Reverse Phase-HPLC. We here report that TRP metabolism favors towards KYN pathway in human meningioma and it could be due to increased indoleamine 2,3-dioxygenase 2 levels as we found its m-RNA levels to be up regulated in human meningioma. We observed significant increase in KYN and 5HIAA levels and significant decrease in TRP, 5-HTP, 5-HT, NAS and MEL levels in meningioma compared to control meninges. Since TRP metabolites regulate inducible nitric oxide synthase (INOS) gene expression and thereby nitric oxide (NO) production, we have also evaluated the INOS and NO levels. The INOS and NO levels were up regulated in human meningioma. The present data corroborates with existing data on TRP metabolism in tumor progression and may serve to target TRP metabolism as a therapeutic intervention.

Hepatokine ITIH3 protects against hepatic steatosis by downregulating mitochondrial bioenergetics and de novo lipogenesis.

Recent studies demonstrate that liver secretory proteins, also known as hepatokines, regulate normal development, obesity, and simple steatosis to non-alcoholic steatohepatitis (NASH) progression. Using a panel of ∼100 diverse inbred strains of mice and a cohort of bariatric surgery patients, we found that one such hepatokine, inter-trypsin inhibitor heavy chain 3 (ITIH3), was progressively lower in severe non-alcoholic fatty liver disease (NAFLD) disease states highlighting an inverse relationship between Itih3/ITIH3 expression and NAFLD severity. Follow-up animal and cell culture models demonstrated that hepatic ITIH3 overexpression lowered liver triglyceride and lipid droplet accumulation, respectively. Conversely, ITIH3 knockdown in mice increased the liver triglyceride in two independent NAFLD models. Mechanistically, ITIH3 reduced mitochondrial respiration and this, in turn, reduced liver triglycerides, via downregulated de novo lipogenesis. This was accompanied by increased STAT1 signaling and Stat3 expression, both of which are known to protect against NAFLD/NASH. Our findings indicate hepatokine ITIH3 as a potential biomarker and/or treatment for NAFLD.

Lipid-droplet associated mitochondria promote fatty-acid oxidation through a distinct bioenergetic pattern in male Wistar rats.

Mitochondria empower the liver to regulate lipid homeostasis by enabling fatty acid oxidation during starvation and lipogenesis during nutrient-rich conditions. It is unknown if mitochondria can seamlessly regulate these two distinct processes or if two discrete populations of mitochondria achieve these two functions in the liver. For the first time in the liver, we report the isolation of two distinct populations of mitochondria from male Wistar rats on an ad-libitum diet: cytoplasmic mitochondria and lipid droplet-associated mitochondria. Our studies show that while lipid droplet mitochondria exhibit higher fatty acid oxidation and are marked by enhanced levels of pACC2, MFN2, and CPT1 activity, cytoplasmic mitochondria are associated with higher respiration capacity. Notably, lipid droplet-associated mitochondria isolated from a non-alcoholic fatty liver disease (NAFLD) rat model are compromised for fatty acid oxidation. We demonstrate the importance of functional segregation of mitochondria as any aberration in lipid droplet-associated mitochondria may lead to NAFLD.

Kisspeptin preserves mitochondrial function by inducing mitophagy and autophagy in aging rat brain hippocampus and human neuronal cell line.

It has become amply clear that mitochondrial function defined by quality, quantity, dynamics, homeostasis, and regulated by mitophagy and mitochondrial biogenesis is a critical metric of human aging and disease. As a consequence, therapeutic interventions that can improve mitochondrial function can have a profound impact on human health and longevity. Kisspeptins are neuropeptides belonging to the family of metastasis suppressors that are known to regulate functions like fertility, reproduction, and metabolism. Using SKNSH cell line, hippocampus explant cultures and hippocampus of aging Wistar rat models, we show that Kisspeptin-10 (Kp) induces autophagy and mitophagy via calcium, Ca2+/CaM-dependent protein kinase kinase ß (CaMKKß), AMP-activated protein kinase (AMPK), and Unc-51 like autophagy activating kinase (ULK1) signaling pathway that is independent of mammalian target of rapamycin (mTOR). Intriguingly, Kp administration in vivo also results in the enhancement of mitochondrial number, complex I activity, and Adenosine Triphosphate (ATP) levels. This study uncovers potential effects of Kp in protecting mitochondrial health and as a possible therapeutic intervention to hippocampus associated impairments such as memory, cognitive aging, and other diseases linked to mitochondrial dysfunction.

Aging reduces kisspeptin receptor (GPR54) expression levels in the hypothalamus and extra-hypothalamic brain regions.

Aging leads to the diminished pulsatile secretion of hypothalamic gonadotropin-releasing hormone (GnRH). Kisspeptin (Kp), the upstream regulator of the hypothalamic-pituitary-gonadal (HPG) axis, regulates GnRH synthesis and release through its cognate receptor, G-protein coupled receptor 54 (GPR54). In turn, GnRH regulates GPR54 expression. GnRH administration into the third ventricle has been shown to induce neurogenesis in different brain regions in old age. However, aging-associated changes in hypothalamic and extra-hypothalamic GPR54 expression were unclear. Therefore, the expression levels of GPR54 were evaluated in various brain regions of adult (age, 3-4 months) and old (age, 20-24 months) male Wistar rats in the present study. In the hypothalamus, mRNA and protein levels of Kp and GPR54 were identified to be significantly decreased in old age. Furthermore, GnRH1 expression in the hypothalamus was analyzed to observe the functional consequence of a reduced Kp-GPR54 system in the hypothalamus. It was found that hypothalamic GnRH1 levels were significantly decreased in old age. As GnRH regulates GPR54 levels, GPR54 was examined in extra-hypothalamic regions. GPR54 levels were found to be significantly decreased in the hippocampus and medulla and pons in old-age rats when compared to adult rats. Notably, GPR54 expression was observed in the frontal lobe, cortex, midbrain and cerebellum of adult and old-age rats; however, the difference between the two groups was not statistically significant. To the best of our knowledge, this is the first study that provides the quantitative distribution of GPR54 in different brain regions during aging. Thus, the reduced levels of Kp and its receptor, GPR54 in the hypothalamus could be cumulatively responsible for reduced levels of GnRH observed in old age.

Glutathionylated and Fe-S cluster containing hMIA40 (CHCHD4) regulates ROS and mitochondrial complex III and IV activities of the electron transport chain.

Human MIA40, an intermembrane space (IMS) import receptor of mitochondria harbors twin CX9C motifs for stability while its CPC motif is known to facilitate the import of IMS bound proteins. Site-directed mutagenesis complemented by MALDI on in vivo hMIA40 protein shows that a portion of MIA40 undergoes reversible S-glutathionylation at three cysteines in the twin CX9C motifs and the lone cysteine 4 residue. We find that HEK293T cells expressing hMIA40 mutant defective for glutathionylation are compromised in the activities of complexes III and IV of the Electron Transport Chain (ETC) and enhance Reactive Oxygen Species (ROS) levels. Immunocapture studies show MIA40 interacting with complex III. Interestingly, glutathionylated MIA40 can transfer electrons to cytochrome C directly. However, Fe-S clusters associated with the CPC motif are essential to facilitate the two-electron to one-electron transfer for reducing cytochrome C. These results suggest that hMIA40 undergoes glutathionylation to maintain ROS levels and for optimum function of complexes III and IV of ETC. Our studies shed light on a novel post-translational modification of hMIA40 and its ability to act as a redox switch to regulate the ETC and cellular redox homeostasis.