Tacrolimus suppresses tumour necrosis factor-alpha and protects against splanchnic artery occlusion shock

Abstract

1. Tumour necrosis factor (TNF-alpha) is a pleiotropic cytokine which is deeply involved in the pathogenesis of splanchnic artery occlusion (SAO) shock. Tacrolimus, formerly known as FK506, is a macrolide antibiotic, that blocks the transcription of several proinflammatory cytokines including TNF-alpha. 2. Male anaesthetized rats were subjected to clamping of the splanchnic arteries for 45 min. This surgical procedure resulted in an irreversible state of shock (SAO shock). Sham operated animals were used as controls. SAO shocked rats had a decreased survival rate (0% at 4 h of reperfusion, while sham shocked rats survived more than 4 h), enhanced serum TNF-alpha concentrations (415+/-12 U ml(-1)), decreased mean arterial blood pressure (MAP), leukopenia and increased ileal leukocyte accumulation studied by means of myeloperoxidase activity (MPO=7.5+/-0.3 U g(-1) tissue). Moreover aortic rings from shocked rats showed a marked hyporeactivity to phenylephrine (PE, 1 nM - 10 microM), reduced responsiveness to acetylcholine (ACh, 10 nM - 10 microM) and increased staining for intercellular adhesion molecule-1 (ICAM-1). Furthermore increased mRNA for TNF-alpha was observed in peritoneal macrophages of SAO shocked rats. 3. Tacrolimus (100 microg kg(-1), 5 min after splanchnic arteries occlusion) increased survival rate (SAO + Tacrolimus = 100% at 4 h of reperfusion), reverted the marked hypotension, reduced serum TNF-alpha (15+/-3 U ml(-1)), ameliorated leukopenia, reduced ileal MPO (0.9+/-0.01 U g(-1) tissue), restored to control values the hyporeactivity to PE. improved the reduced responsiveness to ACh and blunted the enhanced immunostaining for ICAM-1 in the aorta. Finally tacrolimus suppressed cytokine mRNA levels in peritoneal macrophages. 4. The data suggest that tacrolimus may represent a new therapeutic approach in circulatory shock.

Figure 1

Macrophage TNF-α mRNA expression in peritoneal macrophages harvested at different time points in rats subjected to splanchnic artery occlusion (SAO) shock and treated with tacrolimus (100 μg kg⁻¹ as an i.v. infusion of 0.5 ml kg⁻¹min−1) or vehicle (NaCl 0.9%; 0.5 ml kg⁻¹ min⁻¹ for 15 min as an i.v. infusion), both injected 5 min following the onset of reperfusion.

Figure 2

Effects of tacrolimus (100 μg kg⁻¹ as an i.v. infusion of 0.5 ml kg⁻¹ min⁻¹) or vehicle (NaCl 0.9%; 0.5 ml kg⁻¹ min⁻¹ for 15 min as an i.v. infusion), both injected 5 min following the onset of reperfusion, on immunohistochemical staining for ICAM-1 in aortic (Aorta) and superior mesenteric artery (SMA) endothelium from rats subjected to splanchnic artery occlusion (SAO) shock. Each point represents the mean±s.e.mean of seven experiments. ^*P<0.01 vs SAO+vehicle.

Figure 3

Relaxant effects of acetylcholine (ACh) in aortic rings (contracted with phenylephrine, 100 nM) of sham operated rats and rats subjected to splanchnic artery occlusion (SAO) shock treated with tacrolimus (100 μg kg⁻¹ as an i.v. infusion of 0.5 ml kg⁻¹ min⁻¹) or vehicle (NaCl 0.9%; 0.5 ml kg⁻¹ min⁻¹ for 15 min as an i.v. infusion), both injected 5 min following the onset of reperfusion. Each point represents the mean±s.e.mean from six experiments. ^*P<0.01 vs SAO+vehicle.

Figure 4

Contractile response to cumulative doses of phenylephrine (PE) in endothelium denuded aortic rings from sham-operated rats and rats subjected to splanchnic artery occlusion (SAO) shock treated with tacrolimus (100 μg kg⁻¹ as an i.v. infusion of 0.5 ml kg⁻¹ min⁻¹) or vehicle (NaCl 0.9%; 0.5 ml kg⁻¹ min⁻¹ for 15 min as an i.v. infusion), both injected 5 min following the onset of reperfusion. Each point represents the mean±s.e.mean of seven experiments. P<0.02 vs SAO+vehicle.

Figure 5

Effects of tacrolimus (100 μg kg⁻¹ as an i.v. infusion of 0.5 ml kg⁻¹ min⁻¹) or vehicle (NaCl 0.9%; 0.5 ml kg⁻¹ min⁻¹ for 15 min as an i.v. infusion), both injected 5 min following the onset of reperfusion, on mean arterial blood pressure (MAP) of sham operated rats and rats subjected to the splanchnic artery occlusion (SAO) shock. Each point represents the mean±s.e.mean of six experiments. ^*P<0.01 vs SAO+vehicle.