A high temperature silicon on insulator pressure sensor utilising a Ti/TiN/Pt/Au electrode is presented for improving the thermal stability of ohmic contacts, which can work stably at high temperatures of up to 500°C. To analyse the characteristics of the electrode at high temperatures, a special test structure is measured using the linear transmission line method and Auger electron spectroscopy. To solve the measurement problem, a novel calibration setup is designed to calibrate the absolute pressure sensor at extremely high temperatures. The measurement results have shown that the pressure sensor has a nonlinearity error of 0.17%FS and a sensitivity of 0.24 mV/kPa with a measurement range of 30–150 kPa at 500°C, indicating the good thermal stability of the ohmic contacts.
As a critical sphingolipid metabolite, sphingosine-1-phosphate (S1P) plays an essential role in immune and vascular systems. There are five S1P receptors, designated as S1PR1 to S1PR5, encoded in the human genome, and their activities are governed by endogenous S1P, lipid-like S1P mimics, or nonlipid-like therapeutic molecules. Among S1PRs, S1PR1 stands out due to its nonredundant functions, such as the egress of T and B cells from the thymus and secondary lymphoid tissues, making it a potential therapeutic target. However, the structural basis of S1PR1 activation and regulation by various agonists remains unclear. Here, we report four atomic resolution cryo-electron microscopy (cryo-EM) structures of Gi-coupled human S1PR1 complexes: bound to endogenous agonist d18:1 S1P, benchmark lipid-like S1P mimic phosphorylated Fingolimod [(S)-FTY720-P], or nonlipid-like therapeutic molecule CBP-307 in two binding modes. Our results revealed the similarities and differences of activation of S1PR1 through distinct ligands binding to the amphiphilic orthosteric pocket. We also proposed a two-step “shallow to deep” transition process of CBP-307 for S1PR1 activation. Both binding modes of CBP-307 could activate S1PR1, but from shallow to deep transition may trigger the rotation of the N-terminal helix of Gαi and further stabilize the complex by increasing the Gαi interaction with the cell membrane. We combine with extensive biochemical analysis and molecular dynamic simulations to suggest key steps of S1P binding and receptor activation. The above results decipher the common feature of the S1PR1 agonist recognition and activation mechanism and will firmly promote the development of therapeutics targeting S1PRs.
Abstract As a critical sphingolipid metabolite, sphingosine-1-phosphate (S1P) plays an essential role in immune and vascular systems. There are five S1P receptors, designated as S1PR1-5, encoded in the human genome, and their activities are governed by endogenous S1P, lipid-like S1P mimics, or non-lipid-like therapeutic molecules. Among S1PRs, S1PR1 stands out due to its non-redundant functions, such as the egress of T and B cells from the thymus and secondary lymphoid tissues, making it a potential therapeutic target. However, the structural basis of S1PR1 activation and regulation by various agonists remains unclear. Here we reported four atomic resolution cryo-EM structures of Gi-coupled human S1PR1 complexes: bound to endogenous agonist d18:1 S1P, benchmark lipid-like S1P mimic phosphorylated Fingolimod ((S)-FTY720-P), or non-lipid-like therapeutic molecule CBP-307 in two binding modes. Our results revealed the similarities and differences of activation of S1PR1 through distinct ligands binding to the amphiphilic orthosteric pocket. We also proposed a two-step “shallow to deep” transition process of CBP-307 for S1PR1 activation. Both binding modes of CBP-307 could activate S1PR1, but from shallow to deep transition may trigger the rotation of the N-terminal helix of G αi and further stabilize the complex by increasing the G αi interaction with the cell membrane. We combine with extensive biochemical analysis and molecular dynamic simulations to suggest key steps of S1P binding and receptor activation. The above results decipher the common feature of the S1PR1 agonist recognition and activation mechanism and will firmly promote the development of therapeutics targeting S1P receptors.
To investigate the role of calcitonin gene related peptide(CGRP) and endothelin-1 (ET-1) in cirrhotic cardiomyopathy (CCM).We measured the level of CGRP and ET-1 in the samples of rat heart collected from 15 liver cirrhosis rats and 15 controls by using radio immunoassay.The data showed that the levels of CGRP (74.2130 +/- 10.3776 pg/mg protein) and ET-1 level (1.4780 +/- 0.9235 pg/mg protein) were significantly higher in the cirrhotic rat hearts than those in controls (P < 0.05). The increase of ET-1 in the cirrhotic rat hearts was closely associated with the severity of liver cirrhosis (P = 0.004); whereas no significant association was seen between the CGRP concentration and the severity of liver cirrhosis (P = 0.307).We infer that the increasing of CGRP level in the cirrhotic rat heart may be a protective or antagonistic reaction to ET-1 or other pathogenic factors for cardiac dysfunction. The disturbance of the balance between CGRP and ET-1 in the liver cirrhosis rat hearts may contribute to the pathologic process of CCM.
Calcitonin gene-related peptide (CGRP) plays a crucial role in the modulation of orofacial pain, and we hypothesized that CGRP mediated a neuron-glia crosstalk in orofacial pain. The objective of this study was to elucidate the mechanisms whereby CGRP mediated trigeminal neuron-glia crosstalk in modulating orofacial pain. Orofacial pain was elicited by ligating closed-coil springs between incisors and molars. Trigeminal neurons and satellite glial cells (SGCs) were cultured for mechanistic exploration. Gene and protein expression were determined through immunostaining, polymerase chain reaction, and Western blot. Orofacial pain was evaluated through the rat grimace scale. Our results revealed that the expressions of CGRP were elevated in both trigeminal neurons and SGCs following the induction of orofacial pain. Intraganglionic administration of CGRP and olcegepant exacerbated and alleviated orofacial pain, respectively. The knockdown of CGRP through viral vector-mediated RNA interference was able to downregulate CGRP expressions in both neurons and SGCs and to alleviate orofacial pain. CGRP upregulated the expression of inducible nitric oxide synthase through the p38 signaling pathway in cultured SGCs. In turn, L-arginine (nitric oxide donor) was able to enhance orofacial pain by upregulating CGRP expressions in vivo. In cultured trigeminal neurons, L-arginine upregulated the expression of CGRP, and this effect was diminished by cilnidipine (N-type calcium channel blocker) while not by mibefradil (L-type calcium channel blocker). In conclusion, CGRP modulated orofacial pain through upregulating the expression of nitric oxide through the p38 signaling pathway in SGCs, and the resulting nitric oxide in turn stimulated CGRP expression through N-type calcium channel in neurons, building a CGRP-mediated positive-feedback neuron-glia crosstalk.
To obtain a comprehensive understanding of proteins involved in mitochondrion-sarcoplasmic reticulum (SR) linking, a catalog of proteins from mitochondrion-associated membrane (MAM) of New Zealand white rabbit skeletal muscle were analyzed by an optimized shotgun proteomic method. The membrane fractions were prepared by differential centrifugation and separated by 1D electrophoresis followed by a highly reproducible, automated LC-MS/MS on the hybrid linear ion trap (LTQ)-Orbitrap mass spectrometer. By integrating as low as 1% false discovery rate as one of the features for quality control method, 459 proteins were identified from both of the two independent MAM preparations. Protein pI value, molecular weight range, and transmembrane region were calculated using bioinformatics softwares. One hundred one proteins were recognized as membrane proteins. This protein database suggested that the MAM preparations composed of proteins from mitochondrion, SR, and transverse-tubule. This result indicated mitochondria physically linked with SR in rabbit skeletal muscle, voltage-dependent anion channel 1 (VDAC1), VDAC2, and VDAC3 might participate in formation of the tethers between SR and mitochondria.
'/%3e%3cuse xlink:href='%23a' transform='rotate(23.036 .093 25.536)'/%3e%3cuse xlink:href='%23a' transform='rotate(45.87 1.273 16.18)'/%3e%3cuse xlink:href='%23a' transform='rotate(69.945 .996 12.078)'/%3e%3cuse xlink:href='%23a' transform='rotate(20.66 -19.689 31.932)'/%3e%3c/svg%3e)
'/%3e%3c/svg%3e)