Millipore Sigma Vibrant Logo
Attention: We have moved. Merck Millipore products are no longer available for purchase on MerckMillipore.com.Learn More
 

non-gmp


62 Results Advanced Search  
Showing
Products (3)
Documents (22)
Site Content (37)

Narrow Your Results Use the filters below to refine your search

Document Type

  • (21)
  • (1)
Can't Find What You're Looking For?
Contact Customer Service

 

  • Insulin stimulates glucose transport via nitric oxide/cyclic GMP pathway in human vascular smooth muscle cells. 14615391

    In cultured human vascular smooth muscle cells, insulin increases cyclic GMP production by inducing nitric oxide (NO) synthesis. The aim of the present study was to determine whether in these cells the insulin-stimulated NO/cyclic GMP pathway plays a role in the regulation of glucose uptake.Glucose transport in human vascular smooth muscle cells was measured as uptake of 2-deoxy-d-[3H]glucose, cyclic GMP synthesis was checked by radioimmunoassay, and GLUT4 recruitment into the plasma membrane was determined by immunofluorescence. Insulin-stimulated glucose transport and GLUT4 recruitment were blocked by an inhibitor of NO synthesis and mimicked by NO-releasing drugs. Insulin- and NO-elicited glucose uptake were blocked by inhibitors of soluble guanylate cyclase and cyclic GMP-dependent protein kinase; furthermore, glucose transport was stimulated by an analog of cyclic GMP.Our results suggest that insulin-elicited glucose transport (and the corresponding GLUT4 recruitment into the plasma membrane) in human vascular smooth muscle cells is mediated by an increased synthesis of NO, which stimulates the production of cyclic GMP and the subsequent activation of a cyclic GMP-dependent protein kinase.
    Document Type:
    Reference
    Product Catalog Number:
    05-611
    Product Catalog Name:
    Anti-phospho-VASP (Ser239) Antibody, clone 16C2 (Anti-phospho-VASP (Ser239) Antibody, clone 16C2)

  • No long-term weight maintenance effects of gelatin in a supra-sustained protein diet. 20457173

    In the short-term, gelatin showed stronger hunger suppression and less energy intake compared with other proteins. This study investigated if a supra-sustained gelatin-milk protein (GMP) diet improves weight maintenance (WM) compared with a sustained milk protein (SMP) diet and supra-sustained milk protein (SSMP) diet during a 4-months WM period after 8-week weight loss (WL) in sixty-five healthy subjects (28.6+/-3.4kg/m(2); 44+/-10years). Absolute protein intake was kept constant (sustained) throughout per subject. Diets were: protein(P)/fat(F)/carbohydrate(C): 15/40/45% of energy (En%) (SMP) and 30/25/45 En% (SSMP or GMP) for weeks 9-16. Diets on weeks 17-24: P/F/C: 30/35/35 En% (SMP) and 60/5/35 En% (SSMP or GMP). From weeks 8 to 16, and weeks 16 to 24, changes in BMI were similar between the GMP (-0.4+/-0.6 and 0.3+/-0.7kg/m(2) respectively), and the SMP (-0.7+/-0.9 and 0.1+/-0.7kg/m(2) respectively) and SSMP (-0.6+/-0.6 and 0.3+/-0.6kg/m(2) respectively) diets. Sparing of fat free mass (FFM): increases/decreases in FFM%/fat-mass% from weeks 8 to 16 were similar between the GMP and both control diets, and maintained from weeks 16 to 24. In conclusion, all 3 diets resulted in a successful WM period, while a GMP diet does not improve body weight maintenance and related variables after weight loss compared with a SMP and SSMP diet.
    Document Type:
    Reference
    Product Catalog Number:
    Multiple
    Product Catalog Name:
    Multiple

  • Derivation and propagation of human embryonic stem cell lines from frozen embryos in an animal product-free environment. 22722371

    The protocols described here are comprehensive instructions for deriving human embryonic stem (hES) cell lines in xeno-free conditions from cryopreserved embryos. Details are included for propagation, cryopreservation and characterization. Initial derivation is on feeder cells and is followed by adaptation to a feeder-free environment; competent technicians can perform these simplified methods easily. From derivation to cryopreservation of fully characterized initial stocks takes 3-4 months. These protocols served as the basis for standard operating procedures (SOPs), with both operational and technical components, that we set to meet good manufacturing practice (GMP) and UK regulatory body requirements for derivation of clinical-grade cells. As such, these SOPs are currently used in our current GMP compliant facility to derive hES cell lines ab initio, in an animal product-free environment; these lines are suitable for research and potentially for clinical use in cell therapy. So far, we have derived eight clinical-grade lines, which will be freely available to the scientific community after submission/accession to the UK Stem Cell Bank.
    Document Type:
    Reference
    Product Catalog Number:
    MAB4381
    Product Catalog Name:
    Anti-TRA-1-81 Antibody, clone TRA-1-81 (Anti-TRA-1-81 Antibody, clone TRA-1-81)

  • Scalable production of transplantable dopaminergic neurons from hESCs and iPSCs in xeno-free defined conditions. 22872425

    Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) are potentially an unlimited cell source for cell replacement therapy and personalized medicine. Before hESC- and iPSC-based therapy can be moved from bench to bedside, however, it is essential to establish protocols for generating therapeutically relevant cells, like dopaminergic neurons in defined conditions that are suitable for scalable good manufacturing practice (GMP)-compliant protocols. Here, the derivation and differentiation of functional dopaminergic neurons from hESCs and iPSCs under xeno-free defined conditions are described. These protocols have been validated in multiple hESC and iPSC lines.
    Document Type:
    Reference
    Product Catalog Number:
    MAB342
    Product Catalog Name:
    Anti-Galactocerebroside Antibody, clone mGalC (Anti-Galactocerebroside Antibody, clone mGalC)

  • Novel protective role of endogenous cardiac myocyte P2X4 receptors in heart failure. 24622244

    Heart failure (HF), despite continuing progress, remains a leading cause of mortality and morbidity. P2X4 receptors (P2X4R) have emerged as potentially important molecules in regulating cardiac function and as potential targets for HF therapy. Transgenic P2X4R overexpression can protect against HF, but this does not explain the role of native cardiac P2X4R. Our goal is to define the physiological role of endogenous cardiac myocyte P2X4R under basal conditions and during HF induced by myocardial infarction or pressure overload.Mice established with conditional cardiac-specific P2X4R knockout were subjected to left anterior descending coronary artery ligation-induced postinfarct or transverse aorta constriction-induced pressure overload HF. Knockout cardiac myocytes did not show P2X4R by immunoblotting or by any response to the P2X4R-specific allosteric enhancer ivermectin. Knockout hearts showed normal basal cardiac function but depressed contractile performance in postinfarct and pressure overload models of HF by in vivo echocardiography and ex vivo isolated working heart parameters. P2X4R coimmunoprecipitated and colocalized with nitric oxide synthase 3 (eNOS) in wild-type cardiac myocytes. Mice with cardiac-specific P2X4R overexpression had increased S-nitrosylation, cyclic GMP, NO formation, and were protected from postinfarct and pressure overload HF. Inhibitor of eNOS, L-N(5)-(1-iminoethyl)ornithine hydrochloride, blocked the salutary effect of cardiac P2X4R overexpression in postinfarct and pressure overload HF as did eNOS knockout.This study establishes a new protective role for endogenous cardiac myocyte P2X4R in HF and is the first to demonstrate a physical interaction between the myocyte receptor and eNOS, a mediator of HF protection.
    Document Type:
    Reference
    Product Catalog Number:
    04-811
    Product Catalog Name:
    Anti-phospho-eNOS/NOS III (Thr495) Antibody, rabbit monoclonal (Anti-phospho-eNOS/NOS III (Thr495) Antibody, rabbit monoclonal)

  • Nerve growth factor inhibits PC12 cell PDE 2 phosphodiesterase activity and increases PDE 2 binding to phosphoproteins. 11181844

    Nerve growth factor (NGF) has been shown to increase cyclic AMP in PC12 cells and to potentiate the actions of other agents that raise cyclic AMP. In our studies, NGF causes over 50% loss of PDE 2 activity (cyclic GMP-stimulated cyclic nucleotide phosphodiesterase) in PC12 cells within 24 h. After 72 h of NGF treatment, cyclic AMP hydrolysis in PC12 extracts is no longer cyclic GMP-stimulated. NGF deprivation increases the phosphodiesterase activity of treated cells. NGF does not decrease either PDE 2 mRNA or immunoreactivity of PDE 2A2 protein. Incubation of whole cells with micromolar Na(3)VO(4) mimics NGF treatment, reducing PDE 2 activity in PC12 cells by over 50% after 24 h, suggesting a phosphoprotein-mediated regulation of PDE 2 activity. Protein kinase inhibitor effects were difficult to assess due to their direct interaction with the PDE in cell lysates. To study phosphorylation in PDE 2 regulation, PDE 2A2 was epitope-tagged, and stable clonal PC12 cell transfectants were isolated (PC12B cells). When combined with metabolically labeled (32)P-phosphoproteins in vivo or in vitro, phosphoproteins of 108, 90, 64, 43, 33 and 19 kDa coprecipitated with epitope-tagged PDE 2A2 in an NGF sensitive manner. A 23-kDa phosphoprotein containing immunoreactive phosphoserine associated with the complex in an NGF independent manner. Phosphothreonine plus phosphotyrosine immunoreactivity at 23, 24, and 64 kDa as well as the phosphotyrosine immunoreactivity at 108, 90, 64, 43, 33, and 19 kDa required NGF or orthovanadate treatment. These proteins are hypothesized to be part of an NGF-regulated complex controlling PDE 2A2 activity.
    Document Type:
    Reference
    Product Catalog Number:
    Multiple
    Product Catalog Name:
    Multiple

  • Alveolar epithelial CNGA1 channels mediate cGMP-stimulated, amiloride-insensitive, lung liquid absorption. 21559843

    Impairment of lung liquid absorption can lead to severe respiratory symptoms, such as those observed in pulmonary oedema. In the adult lung, liquid absorption is driven by cation transport through two pathways: a well-established amiloride-sensitive Na(+) channel (ENaC) and, more controversially, an amiloride-insensitive channel that may belong to the cyclic nucleotide-gated (CNG) channel family. Here, we show robust CNGA1 (but not CNGA2 or CNGA3) channel expression principally in rat alveolar type I cells; CNGA3 was expressed in ciliated airway epithelial cells. Using a rat in situ lung liquid clearance assay, CNG channel activation with 1 mM 8Br-cGMP resulted in an approximate 1.8-fold stimulation of lung liquid absorption. There was no stimulation by 8Br-cGMP when applied in the presence of either 100 μM L: -cis-diltiazem or 100 nM pseudechetoxin (PsTx), a specific inhibitor of CNGA1 channels. Channel specificity of PsTx and amiloride was confirmed by patch clamp experiments showing that CNGA1 channels in HEK 293 cells were not inhibited by 100 μM amiloride and that recombinant αβγ-ENaC were not inhibited by 100 nM PsTx. Importantly, 8Br-cGMP stimulated lung liquid absorption in situ, even in the presence of 50 μM amiloride. Furthermore, neither L: -cis-diltiazem nor PsTx affected the β(2)-adrenoceptor agonist-stimulated lung liquid absorption, but, as expected, amiloride completely ablated it. Thus, transport through alveolar CNGA1 channels, located in type I cells, underlies the amiloride-insensitive component of lung liquid reabsorption. Furthermore, our in situ data highlight the potential of CNGA1 as a novel therapeutic target for the treatment of diseases characterised by lung liquid overload.
    Document Type:
    Reference
    Product Catalog Number:
    MAB3132

  • Distribution of soluble guanylyl cyclase in rat retina. 17436468

    The nitric oxide (NO)-cGMP pathway is implicated in modulation of visual information processing in the retina. Despite numerous functional studies of this pathway, information about the retinal distribution of the major downstream effector of NO, soluble guanylyl cyclase (sGC), is very limited. In the present work, we have used immunohistochemistry and multiple labeling to determine the distribution of sGC in rat retina. sGC was present at high levels in inner retina but barely detectable in outer retina. Photoreceptors and horizontal cells, as well as Müller cells, were immunonegative, whereas retinal ganglion cells exhibited moderate staining for sGC. Strong immunostaining was found in subpopulations of bipolar and amacrine cells, but staining was weak in rod bipolar cells, and AII amacrine cells were immunonegative. Double labeling of sGC with neuronal nitric oxide synthase showed that the two proteins are generally located in adjacent puncta in inner plexiform layer, implying paracrine interactions. Our results suggest that the NO-cGMP pathway modulates the neural circuitry in inner retina, preferentially within the cone pathway.
    Document Type:
    Reference
    Product Catalog Number:
    MAB351
    Product Catalog Name:
    Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6 (Anti-Glutamate Decarboxylase Antibody, 65 kDa isoform, clone GAD-6)

  • Sequential activation of soluble guanylate cyclase, protein kinase G and cGMP-degrading phosphodiesterase is necessary for proper induction of long-term potentiation in C ... 15312984

    Long-term potentiation (LTP) is a long-lasting enhancement of synaptic transmission efficacy and is considered the base for some forms of learning and memory. Nitric oxide (NO)-induced formation of cGMP is involved in hippocampal LTP. We have studied in hippocampal slices the effects of application of a tetanus to induce LTP on cGMP metabolism and the mechanisms by which cGMP modulates LTP. Tetanus application induced a transient rise in cGMP, reaching a maximum at 10s and decreasing below basal levels 5 min after the tetanus, remaining below basal levels after 60 min. Soluble guanylate cyclase (sGC) activity increased 5 min after tetanus and returned to basal levels at 60 min. The decrease in cGMP was due to sustained tetanus-induced increase in cGMP-degrading phosphodiesterase activity, which remained activated 60 min after tetanus. Tetanus-induced activation of PDE and decrease of cGMP were prevented by inhibiting protein kinase G (PKG). This indicates that the initial increase in cGMP activates PKG that phosphorylates (and activates) cGMP-degrading PDE, which, in turn, degrades cGMP. Inhibition of sGC, of PKG or of cGMP-degrading phosphodiesterase impairs LTP, indicating that proper induction of LTP involves transient activation of sGC and increase in cGMP, followed by activation of cGMP-dependent protein kinase, which, in turn, activates cGMP-degrading phosphodiesterase, resulting in long-lasting reduction of cGMP content. Hyperammonemia is the main responsible for the neurological alterations found in liver disease and hepatic encephalopathy, including impaired intellectual function. Hyperammonemia impairs LTP in hippocampus by altering the modulation of this sGC-PKG-cGMP-degrading PDE pathway. Exposure of hippocampal slices to 1 mM ammonia completely prevents tetanus-induced decrease of cGMP by impairing PKG-mediated activation of cGMP-degrading phosphodiesterase. This impairment is responsible for the loss of the maintenance of LTP in hyperammonemia, and may be also involved in the cognitive impairment in patients with hyperammonemia and hepatic encephalopathy.
    Document Type:
    Reference
    Product Catalog Number:
    14-688
    Product Catalog Name:
    PKG1α Protein, active, 10 µg (PKG1α Protein, active, 10 µg)