Soluble polycations and cationic amphiphiles inhibit volume-sensitive K-Cl cotransport in human red cell ghosts

1994 ◽  
Vol 266 (4) ◽  
pp. C997-C1005 ◽  
Author(s):  
J. R. Sachs
Keyword(s):  
Kinase Activity ◽  
Membrane Surface ◽  
Protein Kinase Activity ◽  
Red Cell ◽  
Effective Inhibitor ◽  
Inner Membrane ◽  
Cationic Amphiphiles ◽  
Activity Inhibition ◽  
High K ◽  
K Concentration

We have measured the effect of soluble polycations (spermine and methylglyoxal) and cationic amphiphiles (sphingosine and tetracaine) on K-Cl cotransport in shrunken and swollen red cell ghosts. All substances inhibited cotransport, and for each agent, the concentration at which inhibition was half-maximal was about the same for swollen and shrunken ghosts. Acetylspermine was a much less effective inhibitor than spermine, which demonstrates that inhibition depends on the cationic groups of spermine. Spermine was a more effective inhibitor in ATP-free ghosts than in ghosts containing ATP, which eliminates the possibility that inhibition of cotransport activity results from inhibition of protein kinase activity. Inhibition by spermine is as effective in K-free ghosts as in high-K ghosts; spermine does not inhibit cotransport by reducing the effective K concentration at the inner membrane surface. We conclude that regulation of K-Cl cotransport involves negative charges (phosphatidylserine or phosphatidylinositides) at the inner membrane surface and suggest a model that accounts for our findings.

scholarly journals Human red cells protein kinase in normal subjects and patients with hereditary spherocytosis, sickle cell disease, and autoimmune hemolytic anemia

Blood ◽  
1976 ◽  
Vol 48 (6) ◽  
pp. 887-898 ◽  
Author(s):  
E Beutler ◽  
E Guinto ◽  
C Johnson
Keyword(s):  
Protein Kinase ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Kinase Activity ◽  
Hereditary Spherocytosis ◽  
Cell Protein ◽  
Protein Kinase Activity ◽  
Red Cell ◽  
Cell Disease ◽  
Membrane Phosphorylation

Abstract A somewhat simplified modification of a previously described method for the measurement of red cell membrane phosphorylation by ATP has been devised. Phosphorylation of membranes was linear with time for only 5- 10 min, and linearity with membrane concentration was observed only when assays were limited to short incubation times. Protein kinase activity of hereditary spherocytosis (HS) membranes was found to be normal. However, the average phosphorylation after 60 min incubation was less in HS membranes than in normal membranes. Findings similar to those in HS membranes were observed in sickle cell disease. The Km of red cell protein kinase for ATP is approximately 10(-5) M. Membrane phosphate binding sites are not saturated in either HS or normal membranes after 1 hr incubation with ATP. Approximately 27% of phosphorylating activity is lost after 1 hr incubation at 37 degrees C. GTP is a very inefficient phosphate donor. Under the conditions of measurement employed, the enzyme is slightly stimulated by 1 muM cAMP, but is not stimulated by 1 muM cGMP. Dephosphorylation of red cell membranes after labeling occurs at a similar rate in HS as in normal membranes. Although a mild abnormally in membrane phosphorylation is observed in HS, this could not be demonstrated to be due to a decrease in protein kinase activity or in alterations of its kinetic properties. The abnormally seen is not specific for HS.

Effects of ionic calcium on the responses of canine testicular polymodal receptors to algesic substances

1989 ◽  
Vol 62 (1) ◽  
pp. 119-125 ◽  
Author(s):  
J. Sato ◽  
K. Mizumura ◽  
T. Kumazawa
Keyword(s):  
Hypertonic Saline ◽  
Membrane Surface ◽  
High Concentration ◽  
Simultaneous Application ◽  
High K ◽  
Ionic Calcium ◽  
K Concentration ◽  
Free Media ◽  
Dependent Processes

1. To explore possible mechanisms of the responses to algesic substances (bradykinin, hypertonic saline, and high K+ solution) of polymodal receptors in the canine testis, the Ca2+ concentration was varied in vitro. 2. After 1 min in Ca2+-free media, the responses to both high K+ solution (60 mM K+) and hypertonic saline (0.6 M Na+) were significantly augmented and tended to increase further with time; return to normal Ca2+ concentration quickly reversed these changes. These augmenting effects were blocked by the substitution of Mg2+ for Ca2+. The excitation produced by 60 mM K+ was decreased by increasing Ca2+ in a concentration-dependent fashion. 3. Reducing the bath concentration of K+ decreased responses evoked by 9 X 10(-8) M bradykinin (BK), whereas increased K+ concentration had the opposite effect. 4. The excitatory effects of BK were significantly suppressed in extracellular Ca2+-free condition. The suppression was not affected by the addition of Mg2+. Prostaglandin E2, which has been known to be released by BK and to augment the BK response, failed to restore the suppressed response by either preapplication (2.8 X 10(-7) M) or simultaneous application in high concentration (1.4 X 10(-5) M). 5. On the basis of these observations, it was postulated that Ca2+ concentration-dependent changes of the responses to 60 mM K+ and 0.6 M Na+ results from Ca2+-dependent "membrane surface potential" changes. The suppressed response to BK by Ca2+ depletion may be explained by the intervention of Ca2+-dependent processes other than PG production.

scholarly journals Flavonoids: Molecular Mechanism Behind Natural Chemoprotective Behavior-A Mini Review

2021 ◽  
Vol 12 (5) ◽  
pp. 5983-5995
Keyword(s):  
Anticancer Agents ◽  
Molecular Mechanisms ◽  
Kinase Activity ◽  
Protein Kinase Activity ◽  
Apoptosis Induction ◽  
Antioxidative Effect ◽  
Pharmacological Activities ◽  
Activity Inhibition ◽  
Metabolic Action ◽  
The Given

The given review summarizes the various molecular mechanisms responsible for the metabolic action of flavonoids as anticancer agents. Various types of flavonoids have proven to show biological & pharmacological activities like anti-inflammatory, antimicrobial, antioxidant, anticancer, anti-allergic, and antidiarrheal activities. The chemoprotective nature of flavonoids is also discussed due to their ability to inhibit topoisomerase enzymes at various stages of cancer, such as tumor initiation, promotion, and progression. The various biomolecular activities which are responsible for their role as the chemopreventive agent may be due to their antioxidative effect, anti-angiogenic properties, induction of protective enzymes, inhibitory action on the cell-like protein kinase activity inhibition, spreading of tumor cells, apoptosis induction, tumor cell invasion to name a few. There has been much-emerging evidence based on the versatility of flavonoids, their complex mechanism of action, lesser side effects, and varied pharmacological properties that make them potential anticancer agents. Challenges associated with their use in extraction, isolation, purification, and checking bioefficacy are also discussed.

scholarly journals Human red cells protein kinase in normal subjects and patients with hereditary spherocytosis, sickle cell disease, and autoimmune hemolytic anemia

Blood ◽  
1976 ◽  
Vol 48 (6) ◽  
pp. 887-898
Author(s):  
E Beutler ◽  
E Guinto ◽  
C Johnson
Keyword(s):  
Protein Kinase ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Kinase Activity ◽  
Hereditary Spherocytosis ◽  
Cell Protein ◽  
Protein Kinase Activity ◽  
Red Cell ◽  
Cell Disease ◽  
Membrane Phosphorylation

A somewhat simplified modification of a previously described method for the measurement of red cell membrane phosphorylation by ATP has been devised. Phosphorylation of membranes was linear with time for only 5- 10 min, and linearity with membrane concentration was observed only when assays were limited to short incubation times. Protein kinase activity of hereditary spherocytosis (HS) membranes was found to be normal. However, the average phosphorylation after 60 min incubation was less in HS membranes than in normal membranes. Findings similar to those in HS membranes were observed in sickle cell disease. The Km of red cell protein kinase for ATP is approximately 10(-5) M. Membrane phosphate binding sites are not saturated in either HS or normal membranes after 1 hr incubation with ATP. Approximately 27% of phosphorylating activity is lost after 1 hr incubation at 37 degrees C. GTP is a very inefficient phosphate donor. Under the conditions of measurement employed, the enzyme is slightly stimulated by 1 muM cAMP, but is not stimulated by 1 muM cGMP. Dephosphorylation of red cell membranes after labeling occurs at a similar rate in HS as in normal membranes. Although a mild abnormally in membrane phosphorylation is observed in HS, this could not be demonstrated to be due to a decrease in protein kinase activity or in alterations of its kinetic properties. The abnormally seen is not specific for HS.

Hair cell changes after cationic stimulation of corti's organ

1989 ◽  
Vol 47 ◽  
pp. 798-799
Author(s):  
Cesar D. Fermin ◽  
Hans-Peter Zenner
Keyword(s):  
Organ Of Corti ◽  
Cross Sectional ◽  
Surface Areas ◽  
High K ◽  
Anatomical Arrangement ◽  
Cell Cytosol ◽  
High Concentrations ◽  
K Concentration ◽  
Cell Changes

Contraction of outer and inner hair cells (OHC&IHC) in the Organ of Corti (OC) of the inner ear is necessary for sound transduction. Getting at HC in vivo preparations is difficult. Thus, isolated HCs have been used to study OHC properties. Even though viability has been shown in isolated (iOHC) preparations by good responses to current and cationic stimulation, the contribution of adjoining cells can not be explained with iOHC preparations. This study was undertaken to examine changes in the OHC after expossure of the OHC to high concentrations of potassium (K) and sodium (Na), by carefully immersing the OC in either artifical endolymph or perilymph. After K and Na exposure, OCs were fixed with 3% glutaraldehyde, post-fixed in osmium, separated into base, middle and apex and embedded in Araldite™. One μm thick sections were prepared for analysis with the light and E.M. Cross sectional areas were measured with Bioquant™ software.Potassium and sodium both cause isolated guinea pig OHC to contract. In vivo high K concentration may cause uncontrolled and sustained contractions that could contribute to Meniere's disease. The behavior of OHC in the vivo setting might be very different from that of iOHC. We show here changes of the cell cytosol and cisterns caused by K and Na to OHC in situs. The table below shows results from cross sectional area measurements of OHC from OC that were exposed to either K or Na. As one would expect, from the anatomical arrangement of the OC, OHC#l that are supported by rigid tissue would probably be displaced (move) less than those OHC located away from the pillar. Surprisingly, cells in the middle turn of the cochlea changed their surface areas more than those at either end of the cochlea. Moreover, changes in surface area do not seem to differ between K and Na treated OCs.

Effect of Thrombin on Phosphorylation of Platelet Membrane Proteins

1976 ◽  
Vol 35 (03) ◽  
pp. 635-642 ◽  
Author(s):  
M Steiner
Keyword(s):  
Protein Kinase ◽  
Kinase Activity ◽  
Qualitative Change ◽  
Protein Kinase Activity ◽  
Protein Substrates ◽  
Phosphoprotein Phosphatase ◽  
Progressive Loss ◽  
Platelet Membranes ◽  
Endogenous Protein ◽  
Considerable Loss

SummaryThe effect of thrombin on the phosphorylating activity of platelet membranes was compared to that of trypsin. Preincubation of non-32P phosphorylated platelet membranes with or without either of these two enzymes resulted in a considerable loss of membrane protein kinase activity which was most severe when trypsin was used. Protein kinase activity and endogenous protein acceptors decreased in parallel. 32P-phosphorylated membranes showed a slow but progressive loss of label which was accelerated by trypsin. Thrombin under these conditions prevented the loss of 32P-phosphate. These results are interpreted to indicate a thrombin-induced destruction of a phosphoprotein phosphatase. The protein kinase activity of phosphorylated platelet membranes using endogenous or exogenous protein substrates showed a significant reduction compared to non-phosphorylated membranes suggesting a deactivation of protein kinase by phosphorylation of platelet membranes. Neither thrombin nor trypsin caused a qualitative change in the membrane polypeptides accepting 32P-phosphate but resulted in quantitative alterations of their ability to become phosphorylated.

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