Activation of SAPKs/JNKs and p38s In Vitro

互联网2014-02-13

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The stress-activated protein kinases (SAPKs)/c-Jun N₂ -terminal kinases (JNKs) and the p38 mitogen-activated protein kinases (MAPKs) are two MAPK groups activated predominantly by stresses and proinflammatory stimuli. Table 1 lists some of the stimuli known to recruit both the SAPKs/JNKs and p38s. In general, the same stimuli that activate the SAPKs/JNKs will also activate the p38s. One exception is ischemia-reperfusion. In this instance, the SAPKs/JNKs are activated exclusively during reperfusion, while the p38s are activated during ischemia and remain active during reperfusion (1 ).

Table 1 Representative Stimuli That Activate SAPK/JNK and p38 In Vivo ^a

Stimulus (ref.)	Target kinase(s)	Sensitive cell lines	Amount used	Comments
Anisomycin (6 )	SAPK/JNK, p38	All cells	1–10 μg/mL, 1–60 min
UV-C (12 )	SAPK/JNK, p38	All adherent cells	40 J/m² , 20 s^-1	Drain medium from cells, and wash with PBS before exposure.
γ Radiation (13 )	SAPK/JNK, p38	All cells (see comments)	10–20 gy, 137Cs source at 0.76 gy/min	This is controversial and has not been replicated in some laboratories. After treatment, wait between 20 min and 4 h before harvest.
cis platinum (CDDP) or cytosine arabinoside (ara-C) (13 )	SAPK/JNK, p38	All cells	50–300 μM , 2–6 h
Methyl methane sulfonate (MMS) (13 )	SAPK/JNK, p38	All cells	1 mM , 3 h
Oxidant stress (H₂ O₂ )(9 )	SAPK/JNK, p38	All cells	2–20 mM , 2–60 min
TNF (6 )	SAPK/JNK, p38	HepG2, CCD18-CO, HEK293, COS, U937, HL-60, Jurkat, L929, HeLa, primary thymocytes or hepatocytes	2–100 ng/mL, 1–60 min	Try to pick a cell type in which TNF is known to induce a biologic effect.
IL-1 (6 )	SAPK/JNK, p38	EL-4, HepG2, Jurkat, HL-60, KB	2–100 ng/mL, 1–60 min	Try to pick a cell type in which IL-is known to induce a biologic effect.
Endothelin (14 , 15 )	SAPK/JNK, p38	Endothelial, mesangial, pulmonary fibroblast, vascular smooth muscle, cardiomyocyte	10–200 nM , 2–60 min	Try to pick a cell type in which endothelin is known to induce a biologic effect.
Angiotensin II (14 )	SAPK/JNK	Endothelial, mesangial, pulmonary fibroblast, vascular smooth muscle, cardiomyocyte,	20–500 nM , 2–60 min	Try to pick a cell type in which the type II antiotensin II receptor is expressed and in which angiotensin II is known to induce a biologic effect.
Ischemia/reperfusion (16 )	SAPK/JNK, p38	Kidney, heart, brain	40 min ischemia, 2–120 min reperfusion	SAPK/JNK is activated only during reperfusion.

^a This table is meant to be a guide for stimuli that recruit SAPK/JNK and p38. The lists of stimuli and cells used are in no way inclusive. Theconditions for stimulation are also meant to serve as a guide. We strongly recommend that time courses and dose responses be performed whenever possible.