MYH9 facilitates autoregulation of adipose tissue depot development.

White adipose tissue not only serves as a reservoir for energy storage but also secretes a variety of hormonal signals and modulates systemic metabolism. A substantial amount of adipose tissue develops in early postnatal life, providing exceptional access to the formation of this important tissue. Although a number of factors have been identified that can modulate the differentiation of progenitor cells into mature adipocytes in cell-autonomous assays, it remains unclear which are connected to physiological extracellular inputs and are most relevant to tissue formation in vivo. Here, we elucidate that mature adipocytes themselves signal to adipose depot-resident progenitor cells to direct depot formation in early postnatal life and gate adipogenesis when the tissue matures. Our studies revealed that as the adipose depot matures, a signal generated in mature adipocytes is produced, converges on progenitor cells to regulate the cytoskeletal protein MYH9, and attenuates the rate of adipogenesis in vivo.

TRPC4/TRPC5 channels mediate adverse reaction to the cancer cell cytotoxic agent (-)-Englerin A.

(-)-Englerin A (EA) is a natural product which has potent cytotoxic effects on renal cell carcinoma cells and other types of cancer cell but not non-cancer cells. Although selectively cytotoxic to cancer cells, adverse reaction in mice and rats has been suggested. EA is a remarkably potent activator of ion channels formed by Transient Receptor Potential Canonical 4 and 5 proteins (TRPC4 and TRPC5) and TRPC4 is essential for EA-mediated cancer cell cytotoxicity. Here we specifically investigated the relevance of TRPC4 and TRPC5 to the adverse reaction. Injection of EA (2 mg.kg-1 i.p.) adversely affected mice for about 1 hour, manifesting as a marked reduction in locomotor activity, after which they fully recovered. TRPC4 and TRPC5 single knockout mice were partially protected and double knockout mice fully protected. TRPC4/TRPC5 double knockout mice were also protected against intravenous injection of EA. Importance of TRPC4/TRPC5 channels was further suggested by pre-administration of Compound 31 (Pico145), a potent and selective small-molecule inhibitor of TRPC4/TRPC5 channels which did not cause adverse reaction itself but prevented adverse reaction to EA. EA was detected in the plasma but not the brain and so peripheral mechanisms were implicated but not identified. The data confirm the existence of adverse reaction to EA in mice and suggest that it depends on a combination of TRPC4 and TRPC5 which therefore overlaps partially with TRPC4-dependent cancer cell cytotoxicity. The underlying nature of the observed adverse reaction to EA, as a consequence of TRPC4/TRPC5 channel activation, remains unclear and warrants further investigation.

Picomolar, selective, and subtype-specific small-molecule inhibition of TRPC1/4/5 channels.

The concentration of free cytosolic Ca2+ and the voltage across the plasma membrane are major determinants of cell function. Ca2+-permeable non-selective cationic channels are known to regulate these parameters, but understanding of these channels remains inadequate. Here we focus on transient receptor potential canonical 4 and 5 proteins (TRPC4 and TRPC5), which assemble as homomers or heteromerize with TRPC1 to form Ca2+-permeable non-selective cationic channels in many mammalian cell types. Multiple roles have been suggested, including in epilepsy, innate fear, pain, and cardiac remodeling, but limitations in tools to probe these channels have restricted progress. A key question is whether we can overcome these limitations and develop tools that are high-quality, reliable, easy to use, and readily accessible for all investigators. Here, through chemical synthesis and studies of native and overexpressed channels by Ca2+ and patch-clamp assays, we describe compound 31, a remarkable small-molecule inhibitor of TRPC1/4/5 channels. Its potency ranged from 9 to 1300 pm, depending on the TRPC1/4/5 subtype and activation mechanism. Other channel types investigated were unaffected, including TRPC3, TRPC6, TRPV1, TRPV4, TRPA1, TRPM2, TRPM8, and store-operated Ca2+ entry mediated by Orai1. These findings suggest identification of an important experimental tool compound, which has much higher potency for inhibiting TRPC1/4/5 channels than previously reported agents, impressive specificity, and graded subtype selectivity within the TRPC1/4/5 channel family. The compound should greatly facilitate future studies of these ion channels. We suggest naming this TRPC1/4/5-inhibitory compound Pico145.

Activation of transient receptor potential vanilloid 3 channel suppresses adipogenesis.

The present study shows that activation of the transient receptor potential vanilloid 3 channel (TRPV3) suppresses adipocyte differentiation. We also found that a major functional catechin compound in green tea and cocoa, (-)-epicatechin, exerts antiadipogenic effects in the adipocytes through direct activation of TRPV3. TRPV3 was detected in the 3T3-L1 adipocytes using immunohistochemistry and semiquantitative PCR. TRPV3 activation by activators (-)-epicatechin and diphenylborinic anhydride was determined using live cell fluorescent Ca(2+) imaging and patch-clamp electrophysiology. Using RNA interference, immunoblotting, and Oil red O staining, we found that the TRPV3 agonists prevented adipogenesis by inhibiting the phosphorylation of insulin receptor substrate 1, the downstream phosphoinositide 3-kinase/Akt/forkhead box protein O1 axis, and the expression of the adipogenic genes peroxisome proliferator-activated receptor γ and CCAAT/enhancer-binding protein α. TRPV3 overexpression hindered adipogenesis in the 3T3-L1 cells. In vivo studies showed that chronic treatment with the TRPV3 activators prevented adipogenesis and weight gain in the mice fed on high-fat diets. Moreover, TRPV3 expression was reduced in the visceral adipose tissue from mice fed on high-fat diets and obese (ob/ob) and diabetic (db/m(+)) mice. In conclusion, our study illustrates the antiadipogenic role of TRPV3 in the adipocytes.

An upregulation in the expression of vanilloid transient potential channels 2 enhances hypotonicity-induced cytosolic Ca²âº rise in human induced pluripotent stem cell model of Hutchinson-Gillford Progeria.

Hutchinson-Gillford Progeria Syndrome (HGPS) is a fatal genetic disorder characterized by premature aging in multiple organs including the skin, musculoskeletal and cardiovascular systems. It is believed that an increased mechanosensitivity of HGPS cells is a causative factor for vascular cell death and vascular diseases in HGPS patients. However, the exact mechanism is unknown. Transient receptor potential (TRP) channels are cationic channels that can act as cellular sensors for mechanical stimuli. The aim of this present study was to examine the expression and functional role of TRP channels in human induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) from the patients with HGPS. The mRNA and protein expression of TRP channels in HGPS and control (IMR90) iPSC-ECs were examined by semi-quantitative RT-PCRs and immunoblots, respectively. Hypotonicity-induced cytosolic Ca²âº ([Ca²âº](i)) rise in iPSC-ECs was measured by confocal microscopy. RT-PCRs and immunoblots showed higher expressional levels of TRPV2 in iPSC-ECs from HGPS patients than those from normal individuals. In functional studies, hypotonicity induced a transient [Ca²âº](i) rise in iPSC-ECs from normal individuals but a sustained [Ca²âº](i) elevation in iPSC-ECs from HGPS patients. A nonselective TRPV inhibitor, ruthenium red (RuR, 20 µM), and a specific TRPV2 channel inhibitor, tranilast (100 µM), abolished the sustained phase of hypotonicity-induced [Ca²âº](i) rise in iPSC-ECs from HGPS patients, and also markedly attenuated the transient phase of the [Ca²âº](i) rise in these cells. Importantly, a short 10 min hypotonicity treatment caused a substantial increase in caspase 8 activity in iPSC-ECs from HGPS patients but not in cells from normal individuals. Tranilast could also inhibit the hypotonicity-induced increase in caspase 8 activity. Taken together, our data suggest that an up-regulation in TRPV2 expression causes a sustained [Ca²âº](i) elevation in HGPS-iPSC-ECs under hypotonicity, consequently resulting in apoptotic cell death. This mechanism may contribute to the pathogenesis of vascular diseases in HGPS patients.