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O. Austen, C. Demoliou Mason, A. D. Hughes, P-418: Effect of permissive and non-permissive temperatures on responses of temperature sensitive (TS) SV40 transformed human vascular smooth muscle cells, American Journal of Hypertension, Volume 14, Issue S1, April 2001, Pages 170A–171A, https://doi.org/10.1016/S0895-7061(01)01576-X
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Abstract
In view of the limited availability of human vascular smooth muscle cells, we recently developed an immortalized cell line (SM1) by infection of saphenous vein-derived smooth muscle cells with a temperature sensitive (ts) SV40 LT-antigen [Hsieh et al., Arterioscler Thromb Vasc Biol 2000;20:973-81.]. Although transformed, these cells retain a number of cytoskeletal characteristics typical of smooth muscle, particularly when cultured at a non-permissive temperature (39°C) that inhibits expression of LT-antigen. The object of these studies was to examine the reponses to agonists in this cell line. Increases in intracellular Ca2+([Ca2+]i) in response to agonists were measured in SM1 cells cultured in DMEM containing 15% FCS at 37°C or 39°C for 5-7 days. Prior to experiments cells were loaded with 5μM fura-2AM for 1h. Cells were then washed, trypsinized and resuspended in modified Kreb[scquote]s buffer. [Ca2+]i was measured as previously described [Hughes & Schachter, Cell Calcium 1994;15:317-330.]. Unstimulated levels of [Ca2+]i were 90±7nM (n=30) and 97±8nM (n=24) at 37°C and 39°C respectively. Increases in [Ca2+]i (nM) in response to stimuli are shown (Table). Following exposure to 39°C responses to platelet-derived growth factor-BB (PDGF; 10ng/ml), epidermal growth factor (EGF; 100ng/ml) phenylephrine (PE; 10μM), angiotensin II (AII; 1μM) and endothelin (ET-1; 100nM) were increased. The response to bradykinin (BK; 100nM) did not change significantly.
| . | PDGF . | EGF . | PE . | BK . | AII . | ET-1 . |
|---|---|---|---|---|---|---|
| 37°C | 40±15 (n=6) | 57±20 (n=6) | 32±16 (n=6) | 252±36 (n=6) | 74±19 (n=6) | 47±13 (n=6) |
| 39°C | 149±48 (n=3) | 207±43 (n=3) | 169±15 (n=3) | 165±41 (n=6) | 157±37 (n=6) | 252±42 (n=3) |
| . | PDGF . | EGF . | PE . | BK . | AII . | ET-1 . |
|---|---|---|---|---|---|---|
| 37°C | 40±15 (n=6) | 57±20 (n=6) | 32±16 (n=6) | 252±36 (n=6) | 74±19 (n=6) | 47±13 (n=6) |
| 39°C | 149±48 (n=3) | 207±43 (n=3) | 169±15 (n=3) | 165±41 (n=6) | 157±37 (n=6) | 252±42 (n=3) |
| . | PDGF . | EGF . | PE . | BK . | AII . | ET-1 . |
|---|---|---|---|---|---|---|
| 37°C | 40±15 (n=6) | 57±20 (n=6) | 32±16 (n=6) | 252±36 (n=6) | 74±19 (n=6) | 47±13 (n=6) |
| 39°C | 149±48 (n=3) | 207±43 (n=3) | 169±15 (n=3) | 165±41 (n=6) | 157±37 (n=6) | 252±42 (n=3) |
| . | PDGF . | EGF . | PE . | BK . | AII . | ET-1 . |
|---|---|---|---|---|---|---|
| 37°C | 40±15 (n=6) | 57±20 (n=6) | 32±16 (n=6) | 252±36 (n=6) | 74±19 (n=6) | 47±13 (n=6) |
| 39°C | 149±48 (n=3) | 207±43 (n=3) | 169±15 (n=3) | 165±41 (n=6) | 157±37 (n=6) | 252±42 (n=3) |
Inhibition of LT-antigen expression in SM1 cells is associated with a pattern of responses to stimulants more typical of untransformed human saphenous vein smooth muscle cells. SM1 cells cultured at non-permissive temperatures may be a useful model of human vascular smooth muscle cells.
- phenylephrine
- endothelin-1
- blood platelets
- smooth muscle
- angiotensin ii
- saphenous vein
- calcium
- central nervous system stimulants
- muscle, smooth, vascular
- antigens
- buffers
- cell lines
- cytoskeleton
- endothelins
- epidermal growth factor
- mechlorethamine
- new mexico
- platelet-derived growth factor
- simian virus 40
- infections
- bradykinin
- temperature
- agonists
- myocytes, smooth muscle
