Abolition by Myosin and Heavy Meromyosin of the Inhibitory Effect of Smooth-Muscle Actomyosin Antibodies on Cell Aggregation In Vitro

1973 ◽  
Vol 12 (2) ◽  
pp. 631-639
Author(s):  
R. B. KEMP ◽  
B. M. JONES ◽  
U. GRÖSCHEL-STEWART
Keyword(s):  
Smooth Muscle ◽  
Striated Muscle ◽  
Cell Aggregation ◽  
Inhibitory Effect ◽  
Regulatory Function ◽  
Rabbit Serum ◽  
Heavy Meromyosin ◽  
Γ Globulins ◽  
Dissociated Cells

The ability of anti-chicken smooth-muscle actomyosin γ-globulins (anti-GAM) to inhibit the aggregation of dissociated cells from the skeletal muscle and liver of chick embryos was abolished by pretreatment of the anti-GAM with either myosin or heavy meromyosin (HMM). When the same cells were treated with HMM at a concentration of 1 mg per 2 x 106 cells/ml Eagle's MEM they aggregated as readily as untreated cells. The negative electrophoretic mobility of the embryonic chick fibroblastic cells was significantly reduced by the globulin fraction of anti-GAM but not of HMM-treated anti-GAM or non-immunized rabbit serum. Anti-chicken striated muscle actomyosin γ-globulins slightly reduced negative mobility but HMM had no effect. The experiments show that the inhibitory effect on cell aggregation of anti-GAM preparations is produced by the anti-myosin antibodies. They also provide support for the theory that a surface-localized myosin-like protein has a regulatory function in cell adhesion.

Aggregative Behaviour of Embryonic Chick Cells in the Presence of Anti-Bodies Directed Against Actomyosins

1971 ◽  
Vol 9 (1) ◽  
pp. 103-122
Author(s):  
R. B. KEMP ◽  
B. M. JONES ◽  
U. GRÖSCHEL-STEWART
Keyword(s):  
Smooth Muscle ◽  
Cell Surface ◽  
Striated Muscle ◽  
Calf Serum ◽  
Fluorescein Isothiocyanate ◽  
Muscle Cells ◽  
Cell Aggregation ◽  
Inhibitory Effect ◽  
Human Umbilical Cord ◽  
Gyratory Shaker

Skeletal muscle and liver tissue from 9-day-old chick embryos were dissociated into separate cells using 0.25 % (w/v) crude trypsin. The effect of rabbit anti-actomyosin sera on the aggregation of these cells was estimated by the gyratory shaker and turbidimetric methods. Studies were also undertaken on the ability of fluorescein isothiocyanate-labelled rabbit anti-uterine actomyosin serum (FITC-labelled anti-UAM) to stain the cell surface and on the type specificity and species specificity of rabbit anti-chicken actomyosin sera. Antisera against chicken gizzard smooth-muscle actomyosin (anti-GAM) and against chicken pectoralis striated muscle actomyosin (anti-PAM) both gave single precipitin bands with their respective actomyosins on diffusion through agar. The antisera neither reacted with their heterologous actomyosin nor with gizzard tropomyosin; they were type-specific. Serial sections of human cervix were stained in a similar pattern with both anti-UAM and anti-GAM, showing that anti-smooth muscle actomyosin sera were not species-specific. The fibrocytes of the human umbilical cord and human platelets were stained by FITC-labelled anti-UAM serum but not by labelled anti-human PAM. The aggregation of muscle and liver cells over a 24-h period in the presence of antisera against human or chicken PAM was not significantly different from the controls incubated on a gyratory shaker in Eagle's minimum essential medium (MEM) containing 10% (v/v) rabbit non-immunized serum (NIS) or calf serum. However, anti-UAM and anti-GAM inhibited both the rate of aggregation of liver and muscle cells and the size of aggregates attained in 24 h. This effect could not be simulated with specific rabbit antisera against human plasma proteins. The globulin-enriched fraction of anti-GAM markedly inhibited the aggregation of liver and muscle cells in a range of concentrations between 50 and 500 µg per 2 x 106 cells/ml Eagle's MEM. In contrast, the aggregation of cells incubated with globulin-enriched anti-PAM was similar to the controls. The addition of anti-GAM globulins at 1 or 2 h to muscle cells rotated by the turbidimetric method reduced the aggregative competence of the cells over the remainder of a 4-h period. The possibility that the inhibitory effect of anti-UAM and anti-GAM on cell aggregation is due to impurities in the antisera or to a general reaction with cell surface ATPases is discussed but, in the light of evidence, rejected in favour of a reaction between the antisera and an actomyosin of the smooth-muscle type at the cell surface.

Studies on the Mechanism Underlying the Inhibition by Puromycin of Cell Aggregation In Vitro

1970 ◽  
Vol 7 (2) ◽  
pp. 557-573
Author(s):  
M. J. DUNN ◽  
E. OWEN ◽  
R. B. KEMP
Keyword(s):  
Carbon Dioxide ◽  
Protein Synthesis ◽  
Cellular Metabolism ◽  
Cell Aggregation ◽  
Experimental Period ◽  
Inhibitory Effect ◽  
Dissociated Cells ◽  
Gyratory Shaker ◽  
Reduced Rate

Cells dissociated with 0.25% crude trypsin from the muscle tissue of 9-day-old chick embryos were employed to investigate the effect of puromycin on cellular metabolism. Parallel studies were also made, using the gyratory shaker, to confirm the effectiveness of puromycin in inhibiting cell aggregation and protein synthesis. Puromycin when introduced at a concentration of 10µg/ml into a suspension of cells in Eagle's MEM did not completely inhibit cell aggregation. Small aggregates were formed in the first 4 h of the experiment. Protein synthesis of the rotated cells, as measured by the incorporation of L-[α-14C]leucine into proteins, was arrested by 91.7% within 15 min of introducing puromycin into a cell suspension. The antibiotic retained its inhibitory effect on protein synthesis for the 24-h period of rotation. Puromycin inhibited the cellular oxygen uptake and carbon dioxide evolution of the rotated cells by 40% within 4 h of its introduction. However, treated cells were still respiring, though at a much reduced rate, at the end of the 24-h experimental period. The release of radioactive carbon dioxide by puromycin-treated cells was also inhibited by 40% at the 4-h stage but after 8 h no further 14CO2 was evolved. The presence of the antibiotic markedly inhibited the uptake of glucose by trypsin-dissociated cells. The level of glycogen and lactate in cells suspended in Eagle's MEM was reduced very considerably over a 24-h period. The presence of puromycin accelerated glycogen utilization over the first 6 h of rotation but at 24 h there was a difference of only 0.6% between the glycogen content of treated cells and controls. At 24 h 11.3% less lactate remained in the puromycin-treated cells than in the controls. The ATP/ADP ratio of trypsin-dissociated cells decreased from an initial value of 2.59 to 1.45 after rotation for 24 h. In the presence of puromycin the ATP/ADP ratio was 0.62 at 4 h and had further declined to 0.48 by 24 h. The effects of puromycin on the aggregation, protein synthesis and cellular metabolism of trypsin-dissociated cells are discussed in relation to cellular adhesive mechanisms.

scholarly journals Contractile and Endothelium-Dependent Dilatory Responses of Cerebral Arteries at Various Extracellular Magnesium Concentrations

1991 ◽  
Vol 11 (1) ◽  
pp. 161-164 ◽  
Author(s):  
Mária Faragó ◽  
Csaba Szabó ◽  
Eörs Dóra ◽  
Ildikó Horváth ◽  
Arisztid G. B. Kovách
Keyword(s):  
Smooth Muscle ◽  
Vascular Reactivity ◽  
Calcium Influx ◽  
Cerebral Arteries ◽  
Inhibitory Effect ◽  
Contractile Responses ◽  
Middle Cerebral Arteries ◽  
The Right ◽  
Endothelial Receptor

To clarify the effect of extracellular magnesium (Mg2+) on the vascular reactivity of feline isolated middle cerebral arteries, the effects of slight alterations in the Mg2+ concentration on the contractile and endothelium-dependent dilatory responses were investigated in vitro. The contractions, induced by 10−8-10−5 M norepinephrine, were significantly potentiated at low Mg2+ (0.8 m M v. the normal, 1.2 m M). High (1.6 and 2.0 m M) Mg2+ exhibited an inhibitory effect on the contractile responses. No significant changes, however, in the EC50 values for norepinephrine were found. The endothelium-dependent relaxations induced by 108–10−5 M acetylcholine were inhibited by high (1.6 and 2.0 m M) Mg2+. Lowering of the Mg2+ concentration to 0.8 m M or total withdrawal of this ion from the medium failed to alter the dilatory potency of acetylcholine. The changes in the dilatory responses also shifted the EC50 values for acetylcholine to the right. The present results show that the contractile responses of the cerebral arteries are extremely susceptible to the changes of Mg2+ concentrations. In response to contractile and endothelium-dependent dilatory agonists, Mg2+ probably affects both the calcium influx into the endothelial and smooth muscle cells as well as the binding of acetylcholine to its endothelial receptor. Since Mg2+ deficiency might facilitate the contractile but not the endothelium-dependent relaxant responses, the present study supports a role for Mg2+ deficiency in the development of the cerebral vasospasm.

Effects of Cytochalasins on the Initial Aggregation in Vitro of Embryonic Chick Cells

1974 ◽  
Vol 14 (1) ◽  
pp. 187-196
Author(s):  
J. C. APPLETON ◽  
R. B. KEMP
Keyword(s):  
Cytochalasin B ◽  
Cell Aggregation ◽  
Cellular Injury ◽  
Cellular Motility ◽  
Carbon Dioxide Evolution ◽  
Site Of Action ◽  
Dissociated Cells ◽  
Cytochalasin A ◽  
Final Confirmation

The initial aggregation of trypsin-dissociated cells from the skeletal muscle tissue of 9-day-old chick embryos in the presence of cytochalasins A and B was studied in order to discover the effects of these agents on contact and adhesion. Cytochalasin B (3 µg/ml) had a negligible effect on the rate of aggregation of cells over an 8-h period, but cytochalasin A at concentrations between 3 and 20 µg/ml markedly inhibited aggregation. Both agents altered the shape and size of aggregates and caused cells at their periphery to appear more spherical. The oxygen uptake of the treated cells was not noticeably different from that of the controls, despite the severe inhibition of isotopic carbon dioxide evolution. The effect of cytochalasin B on cell aggregation was reversible and although the cytochalasin A effect could not be abolished on return to medium free of A, the unaltered oxygen consumption was taken as an indication that permanent cellular injury did not occur. The effect of the cytochalasins on aggregate structure was interpreted on the basis of arrested cellular motility, but the singular inhibition by cytochalasin A of the rate of aggregation must await final confirmation of its site of action.

scholarly journals LDL Eliminates the Inhibitory Effect of Human Aortic Elastin on Cultured Rat Aortic Smooth Muscle Cell Migration In Vitro

1992 ◽  
Vol 20 (8-9) ◽  
pp. 703-707 ◽  
Author(s):  
Toshiro OOYAMA ◽  
Hiroshi SAKAMOTO ◽  
Katsunori FUKUDA ◽  
Hiromi ODA ◽  
Youko MURAI ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Cell Migration ◽  
Smooth Muscle Cell ◽  
Muscle Cell ◽  
Inhibitory Effect ◽  
Aortic Smooth Muscle Cell ◽  
Aortic Smooth Muscle ◽  
Smooth Muscle Cell Migration

Ultrastructural Evidence for Myosin of the Smooth Muscle Type at the Surface of Trypsin-Dissociated Embryonic Chick Cells

1974 ◽  
Vol 15 (2) ◽  
pp. 279-289
Author(s):  
I. AP GWYNN ◽  
R. B. KEMP ◽  
B. M. JONES ◽  
U. GRÖSCHEL-STEWART
Keyword(s):  
Smooth Muscle ◽  
Plasma Membrane ◽  
Striated Muscle ◽  
Smooth Muscle Myosin ◽  
Rabbit Serum ◽  
Embryonic Chick ◽  
Antibody Preparation ◽  
Rabbit Igg ◽  
Antibody Conjugate ◽  
Muscle Myosin

Cells dissociated from embryonic chick muscle tissue using trypsin were rotated in the presence of globulin-enriched rabbit antisera against both smooth and striated muscle actomyosins originating from chicken gizzard (GAM) and pectoralis (PAM) muscles respectively. The presence of the rabbit antibodies was demonstrated using peroxidase-labelled sheep anti-rabbit λ-globulins, the enzyme-antibody conjugate being located by electron-microscope histochemistry. Anti-GAM λ-globulins reacted strongly with the plasma membrane. Judging from the complete absence of staining, λ-globulins from non-immunized rabbit serum did not interact with the membrane.When λ-globulins of sheep anti-rabbit IgG serum were applied alone, that is in the absence of pretreatment with rabbit λ-globulin, there was an observable reaction with the cell surface. Preincubation of anti-GAM with the heavy meromyosin fraction from smooth-muscle myosin inhibited the interaction of the antibodies with the membrane, as evidenced by the absence of staining. A weak positive reaction obtained with anti-PAM was due to components of the antibody preparation which were reactive with actin and not with PAM. It was concluded that a smooth-muscle myosin-like protein is an integral part of the plasma membrane of embryonic chick muscle cells.

Role of endothelial cells in regulating hemoglobin-induced changes in lymphatic pumping

1992 ◽  
Vol 263 (6) ◽  
pp. H1880-H1887 ◽  
Author(s):  
R. M. Elias ◽  
J. Eisenhoffer ◽  
M. G. Johnston
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Inhibitory Effect ◽  
Direct Inhibitory Effect ◽  
Induced Changes ◽  
Lymphatic Pumping ◽  
Pumping Activity

Studies with a sheep isolated duct preparation in vivo demonstrated that the route of administration of hemoglobin was important in demonstrating its inhibitory effect on lymphatic pumping. With autologous oxyhemoglobin administered intravenously (final plasma concentration 5 x 10(-5) M), pumping was not inhibited. However, the addition of oxyhemoglobin (5 x 10(-5) M) into the reservoir (lumen of the duct) resulted in > 95% inhibition of pumping. The extraluminal administration of oxyhemoglobin (10(-5) M) to bovine mesenteric lymphatics in vitro resulted in a 40% inhibition of pumping, whereas the introduction of oxyhemoglobin (10(-5) M) into the lumen of the vessels suppressed pumping 95%. In vessels mechanically denuded of endothelium, intraluminal oxyhemoglobin inhibited pumping 50%. These results suggested that oxyhemoglobin depressed pumping through an effect on both smooth muscle and endothelium. Once pumping was inhibited with oxyhemoglobin administration, stimulation of the duct with elevations in transmural pressure restored pumping activity when endothelial cells were present. However, in the absence of endothelium, pumping decreased with increases in distending pressures. We conclude that oxyhemoglobin has a direct inhibitory effect on lymphatic smooth muscle. The ability of oxyhemoglobin to alter the pressure range over which the lymph pump operates appears to be dependent on an intact endothelium.

Inhibitory Effect of β3-Adrenoceptor Agonist in Lower Esophageal Sphincter Smooth Muscle: In Vitro Studies

2003 ◽  
Vol 304 (1) ◽  
pp. 48-55 ◽  
Author(s):  
D. N. K. Sarma ◽  
Kuldip Banwait ◽  
Ashim Basak ◽  
Anthony J. DiMarino ◽  
Satish Rattan
Keyword(s):  
Smooth Muscle ◽  
Lower Esophageal Sphincter ◽  
Inhibitory Effect ◽  
In Vitro Studies ◽  
Adrenoceptor Agonist ◽  
Esophageal Sphincter
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