Prolylcarboxypeptidase (PRCP) as a new target for obesity treatment

Recently, we serendipitously discovered that mice with the deficiency of the enzyme prolylcarboxypeptidase (PRCP) have elevated α-melanocyte-stimulating hormone (α-MSH) levels which lead to decreased food intake and weight loss. This suggests that PRCP is an endogenous inactivator of α-MSH and an appetite stimulant. Since a modest weight loss can have the most profound influence on reducing cardiovascular risk factors, the inhibitors of PRCP would be emerging as a possible alternative for pharmacotherapy in high-risk patients with obesity and obesity-related disorders. The discovery of a new biological activity of PRCP in the PRCP-deficient mice and studies of α-MSH function indicate the importance and complexity of the hypothalamic pro-opiomelanocortin (POMC) system in altering food intake. Identifying a role for PRCP in regulating α-MSH in the brain may be a critical step in enhancing our understanding of how the brain controls food intake and body weight. In light of recent findings, the potential role of PRCP in regulating fuel homeostasis is critically evaluated. Further studies of the role of PRCP in obesity are much needed

Cytosolic Calmodulin Is Increased in SK-N-SH Human Neuroblastoma Cells Due to Release of Calcium from Intracellular Stores

Muscarinic receptor stimulation elicits a redistribution of calmodulin (CaM) from the membrane fraction to cytosol in the human neuroblastoma cell line SK-N-SH. Increasing the intracellular Ca 2+ concentration with ionomycin also elevates cytosolic CaM. The aim of this study was to investigate the roles of extracellular and intracellular Ca 2+ pools in the muscarinic receptor-mediated increases in cytosolic CaM in SK-N-SH cells. Stimulus-mediated changes in intracellular Ca 2+ were monitored in fura-2-loaded cells, and CaM was measured by radioimmunoassay in the 100,000- g cytosol and membrane fractions. The influx of extracellular Ca 2+ normally seen with carbachol treatment in SK-N-SH cells was eliminated by pretreatment with the nonspecific Ca 2+ channel blocker Ni 2+ . Blocking the influx of extracellular Ca 2+ had no effect on carbachol-mediated increases in cytosolic CaM (168 18% of control values for carbachol treatment alone vs. 163 28% for Ni 2+ and carbachol) or decreases in membrane CaM. Similarly, removal of extracellular Ca 2+ from the medium did not affect carbachol-mediated increases in cytosolic CaM (168 26% of control). On the other hand, prevention of the carbachol-mediated increase of intracellular free Ca 2+ by pretreatment with the cell-permeant Ca 2+ chelator BAPTA/AM did attenuate the carbachol-mediated increase in cytosolic CaM (221 37% of control without BAPTA/AM vs. 136 13% with BAPTA/AM). The effect of direct entry of extracellular Ca 2+ into the cell by K + depolarization was assessed. Incubation of SK-N-SH cells with 60 m M K + elicited an immediate and persistent increase in intracellular free Ca 2+ concentration, but there was no corresponding alteration in CaM localization. On the contrary, in cells where intracellular Ca 2+ was directly elevated by thapsigargin treatment, cytosolic CaM was elevated for at least 30 min while particulate CaM was decreased. In addition, treatment with ionomycin in the absence of extracellular Ca 2+ , which releases Ca 2+ from intracellular stores, induced an increase in cytosolic CaM (203 30% of control). The mechanism for the CaM release may involve activation of the isozyme of protein kinase C, which was translocated from cytosol to membranes much more profoundly by thapsigargin than by K + depolarization. These data demonstrate that release of Ca 2+ from the intracellular store is important for the carbachol-mediated redistribution of CaM in human neuroblastoma SK-N-SH cells.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/66213/1/j.1471-4159.1998.70010139.x.pd

Effect of Continuous Phorbol Ester Treatment on Muscarinic Receptor-Mediated Calmodulin Redistribution in SK-N-SH Neuroblastoma Cells

Stimulation of muscarinic receptors by carbachol and activation of protein kinase C elicits the translocation of calmodulin (CaM) from membranes to cytosol in the human neuroblastoma cell line SK-N-SH. Our previous studies have suggested a role for protein kinase C in the regulation of CaM redistribution. To explore further the role of protein kinase C in carbachol-induced calmodulin translocation, we treated cells for 17 h with 12- O -tetradecanoylphorbol 13-acetate (TPA) to down-regulate protein kinase C isozymes or 72 h to differentiate the cells. Treatment of SK-N-SH cells for 17 h with 70 n M TPA nearly abolished the effect of carbachol on CaM redistribution. After 72 h of TPA, however, the cells appeared differentiated, and the ability of carbachol to increase cytosolic CaM levels was restored. In untreated control cells, the carbachol-mediated increase in cytosolic CaM content was mimicked by TPA and blocked by pretreatment with the selective protein kinase C inhibitor Ro 31-8220 at 10 µ M . In the 72-h TPA-treated cells, however, the ability of TPA to increase cytosolic CaM levels was significantly reduced, and the action of carbachol was no longer blocked by Ro 31-8220. The effect of prolonged TPA treatment on select protein kinase C isozymes was examined by immunoblotting. Treatment of cells for either 17 or 72 h abolished the Α-isozyme in the cytosol and reduced (17 h) or abolished (72 h) the content in the membranes. In both 17- and 72-h TPA-treated cells, the ε-isozyme was nearly abolished in the cytosol and slightly reduced in the membranes. Some protein kinase C activity may have been maintained during TPA treatment because the basal level of phosphorylation of the protein kinase C substrate myristoylated alanine-rich C kinase substrate was enhanced in cells treated for either 17 or 72 h with TPA. The potential dissociation of carbachol and protein kinase C in eliciting increases in cytosolic CaM content was a function of prolonged TPA treatment and not differentiation per se because carbachol-mediated increases in cytosolic CaM levels were inhibited by Ro 31-8220 in retinoic acid-differentiated SK-N-SH cells. This study demonstrates that continuous TPA treatment, although initially down-regulating the protein kinase C-mediated effect of carbachol on CaM redistribution, uncouples carbachol and protein kinase C at longer times.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65656/1/j.1471-4159.1997.68010040.x.pd

Recombinant prolylcarboxypeptidase activates plasma prekallikrein

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/106127/1/jth03969.pd

Scupa Or Fxii Stimulate Erk1/2 Or Akt Through Upar And Beta 1 Integrins

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/106134/1/jth00092.pd

Mapping the interaction between uPAR and high molecular weight kininogen

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/106070/1/jth03972.pd

Predicting the risk to develop preeclampsia in the first trimester combining promoter variant -98A/C of LGALS13 (placental protein 13), Black ethnicity, previous preeclampsia, obesity, and maternal age

BACKGROUND: We studied LGALS13 [Placental Protein 13 (PP13)] promoter DNA polymorphisms in preeclampsia (PE) prediction, given PP13’s effects on hypotension, angiogenesis and immunotolerance. METHODS: We retrieved 67 PE (49 term, 18 preterm) cases and 196 matched controls from first trimester plasma samples prospectively collected at King's College Hospital, London. Cell-free DNA was extracted and the four LGALS13 exons were sequenced after PCR amplification. Expression of LGALS13 promoter reporter constructs were determined in BeWo trophoblast-like cells with luciferase assays. RESULTS: A/C genotype in –98 position was the lowest in term PE compared to controls (p<0.032), similar to a South African cohort. Control but not all PE allele frequencies were in Hardy-Weinberg equilibrium (p=0.036). The Odds ratio for term PE calculated from prior risk, the A/A genotype and black ethnicity was 14 (p<0.001). In luciferase assays, the LGALS13 promoter “-98A" variant had 13% (p=0.04) and 26% (p<0.001) lower expression than the "-98C" variant in non-differentiated and differentiated BeWo cells, respectively. After 48-hour differentiation, there was 4.55- fold increase in expression of "-98C" variant versus 3.85-fold of "-98A" variant (p<0.001). CONCLUSION: Lower LGALS13 (PP13) expression by the "-98A/A" genotype appears to impose higher risk to develop PE and could aid in PE prediction

BepiColombo’s Cruise Phase: Unique Opportunity for Synergistic Observations

The investigation of multi-spacecraft coordinated observations during the cruise phase of BepiColombo (ESA/JAXA) are reported, with a particular emphasis on the recently launched missions, Solar Orbiter (ESA/NASA) and Parker Solar Probe (NASA). Despite some payload constraints, many instruments onboard BepiColombo are operating during its cruise phase simultaneously covering a wide range of heliocentric distances (0.28 AU–0.5 AU). Hence, the various spacecraft configurations and the combined in-situ and remote sensing measurements from the different spacecraft, offer unique opportunities for BepiColombo to be part of these unprecedented multipoint synergistic observations and for potential scientific studies in the inner heliosphere, even before its orbit insertion around Mercury in December 2025. The main goal of this report is to present the coordinated observation opportunities during the cruise phase of BepiColombo (excluding the planetary flybys). We summarize the identified science topics, the operational instruments, the method we have used to identify the windows of opportunity and discuss the planning of joint observations in the future

BepiColombo's Cruise Phase : Unique Opportunity for Synergistic Observations

The investigation of multi-spacecraft coordinated observations during the cruise phase of BepiColombo (ESA/JAXA) are reported, with a particular emphasis on the recently launched missions, Solar Orbiter (ESA/NASA) and Parker Solar Probe (NASA). Despite some payload constraints, many instruments onboard BepiColombo are operating during its cruise phase simultaneously covering a wide range of heliocentric distances (0.28 AU-0.5 AU). Hence, the various spacecraft configurations and the combined in-situ and remote sensing measurements from the different spacecraft, offer unique opportunities for BepiColombo to be part of these unprecedented multipoint synergistic observations and for potential scientific studies in the inner heliosphere, even before its orbit insertion around Mercury in December 2025. The main goal of this report is to present the coordinated observation opportunities during the cruise phase of BepiColombo (excluding the planetary flybys). We summarize the identified science topics, the operational instruments, the method we have used to identify the windows of opportunity and discuss the planning of joint observations in the future.Peer reviewe