A method of culturing coronary artery explants for measuring vascular smooth muscle proliferation in rainbow trout

1995 ◽  
Vol 73 (4) ◽  
pp. 623-631 ◽  
Author(s):  
Bang Q. Gong ◽  
Anthony P. Farrell
Keyword(s):  
Smooth Muscle ◽  
Rainbow Trout ◽  
Coronary Artery ◽  
Vascular Smooth Muscle ◽  
Thymidine Incorporation ◽  
Recovery Period ◽  
Treated Group ◽  
Smooth Muscle Proliferation

A standardized method of in vitro [3H]thymidine incorporation during the S phase of cell division was developed to study vascular smooth muscle proliferation in the coronary artery of rainbow trout (Oncorhynchus mykiss). We established a reliable medium system, an optimum [3H]thymidine dose, and optimum incubation conditions. Incorporation of the radiolabel into vascular smooth muscle nuclei was confirmed with autoradiography. To test the sensitivity of the assay system, the coronary artery was surgically exposed and gently rubbed in anesthetized rainbow trout. Following either a 1-, 2-, or 3-day recovery period, fish were sacrificed and the coronary artery explants were cultured with [3H]thymidine. Gentle rubbing of the coronary artery in vivo resulted in a significant increase in [3H]thymidine incorporation into the coronary artery explant compared with sham-operated and untreated control groups of fish. Peak incorporation of [3H]thymidine occurred at day 2 in the treated group, when incorporation was three times that in the sham-operated group. This technique has potential application in the study of coronary arteriosclerosis in salmonids, where vascular smooth muscle proliferation is a primary event.

Effects of polyunsaturated fatty acids and some of their metabolites on mitotic activity of vascular smooth muscle explants from the coronary artery of rainbow trout (Oncorhynchus mykiss)

1997 ◽  
Vol 75 (1) ◽  
pp. 80-86 ◽  
Author(s):  
B. Gong ◽  
R. Townley ◽  
A. P. Farrell
Keyword(s):  
Fatty Acids ◽  
Smooth Muscle ◽  
Rainbow Trout ◽  
Coronary Artery ◽  
Vascular Smooth Muscle ◽  
Polyunsaturated Fatty Acids ◽  
Oncorhynchus Mykiss ◽  
Thymidine Incorporation ◽  
Mitogenic Effect ◽  
Rainbow Trout Oncorhynchus Mykiss

This study is the first to examine the effects of polyunsaturated fatty acids and some of their metabolites on [3H]thymidine incorporation into vascular smooth muscle explants from the coronary artery of rainbow trout (Oncorhynchus mykiss). At a concentration of 120 μM, eicosapentaenoic acid (EPA; 20:5ω3), arachidonic acid (AA; 20:4ω6), and eicosatrienoic acid (ETA; 20:3ω6) all approximately doubled [3H]thymidine incorporation relative to controls. At a concentration of 20 μM, EPA had no significant effect, while ETA inhibited and AA caused an almost 5-fold increase in [3H]thymidine incorporation. The large mitogenic effect of 20 μM AA was completely inhibited by simultaneous addition of EPA to the culture medium. ETA only partially inhibited the mitogenic effect of 20 μM AA. Four AA-derived eicosanoids (or their stable analogues) were also tested. [3H]Thymidine incorporation was at least doubled with 1000 ng/mL carbacyclin (a prostacyclin analogue), 120 ng/mL prostaglandin F2α, and U-46619 (a thromboxane A2 analogue), but did not reach the level of stimulation produced by 20 μM AA. Leukotriene C4 had no significant effect. We conclude that dietary modulation of polyunsaturated fatty acids (PUFAs) in salmonids could have significant effects on coronary vascular smooth muscle mitosis through the incorporation of PUFAs into cell membranes and the production of eicosanoids.

Injury-induced expression of cytokeratins 8 and 18 by vascular smooth muscle cells requires concurrent activation of cytoskeletal and growth factor receptors

2008 ◽  
Vol 86 (5) ◽  
pp. 223-231 ◽  
Author(s):  
Michael C. Moon ◽  
Lorraine Yau ◽  
Brenda Wright ◽  
Peter Zahradka
Keyword(s):  
Smooth Muscle ◽  
Coronary Artery ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Femoral Artery ◽  
Porcine Coronary Artery ◽  
Cytokeratin Expression

Cytokeratins are not present in the vascular smooth muscle cells (VSMCs) of normal arteries, but they are detectable in the VSMCs of atherosclerotic lesions. A correlation between cytokeratin expression and VSMC phenotype is proposed, but an examination of VSMCs after mechanical injury has yet to be performed. Immunohistochemistry was used to monitor proteins in arterial sections. Western blotting enabled quantification of protein levels. Angioplasty of porcine femoral artery in vivo and porcine coronary artery in vitro served as models of vascular injury. Cytokeratins 8 and 18 were expressed by VSMCs in porcine femoral artery lesions 14 days after balloon angioplasty. Cytokeratins were also present in the neointima of porcine coronary artery segments placed into organ culture for 4 days. Cytokeratin expression was decreased in the presence of inhibitors that affect MAP kinase, PI3 kinase, Src kinase, and G protein, but not in the presence of an AT1 receptor antagonist. Cytokeratin expression also occurred when VSMCs were plated onto collagen in the presence of serum. We conclude that mechanical injury induces expression of cytokeratin 8 and 18 both in vitro and in vivo by synthetic VSMCs that migrate into the neointima. Furthermore, cytokeratin expression requires cellular attachment to extracellular matrix proteins in conjunction with mitogenic stimulation.

scholarly journals Effects of Hyperglycemia on Vascular Smooth Muscle Ca2+Signaling

2017 ◽  
Vol 2017 ◽  
pp. 1-16 ◽  
Author(s):  
Nahed El-Najjar ◽  
Rashmi P. Kulkarni ◽  
Nancy Nader ◽  
Rawad Hodeify ◽  
Khaled Machaca
Keyword(s):  
Insulin Resistance ◽  
Smooth Muscle ◽  
Sarcoplasmic Reticulum ◽  
Vascular Smooth Muscle ◽  
Complex Disease ◽  
Prolonged Exposure ◽  
In Vitro System ◽  
A7r5 Cells

Diabetes is a complex disease that is characterized with hyperglycemia, dyslipidemia, and insulin resistance. These pathologies are associated with significant cardiovascular implications that affect both the macro- and microvasculature. It is therefore important to understand the effects of various pathologies associated with diabetes on the vasculature. Here we directly test the effects of hyperglycemia on vascular smooth muscle (VSM) Ca2+signaling in an isolated in vitro system using the A7r5 rat aortic cell line as a model. We find that prolonged exposure of A7r5 cells to hyperglycemia (weeks) is associated with changes to Ca2+signaling, including most prominently an inhibition of the passive ER Ca2+leak and the sarcoplasmic reticulum Ca2+-ATPase (SERCA). To translate these findings to the in vivo condition, we used primary VSM cells from normal and diabetic subjects and find that only the inhibition of the ER Ca2+leaks replicates in cells from diabetic donors. These results show that prolonged hyperglycemia in isolation alters the Ca2+signaling machinery in VSM cells. However, these alterations are not readily translatable to the whole organism situation where alterations to the Ca2+signaling machinery are different.

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