Substance P receptor blockade decreases stretch-induced lung cytokines and lung injury in rats

Abstract

Overdistension of lung tissue during mechanical ventilation causes cytokine release, which may be facilitated by the autonomic nervous system. We used mechanical ventilation to cause lung injury in rats, and studied how cervical section of the vagus nerve, or substance P (SP) antagonism, affected the injury. The effects of 40 or 25 cmH(2)O high airway pressure injurious ventilation (HV(40) and HV(25)) were studied and compared with low airway pressure ventilation (LV) and spontaneous breathing (controls). Lung mechanics, lung weight, gas exchange, lung myeloperoxidase activity, lung concentrations of interleukin (IL)-1 beta and IL-6, and amounts of lung SP were measured. Control rats were intact, others were bivagotomized, and in some animals we administered the neurokinin-1 (NK-1) receptor blocking agent SR140333. We first determined the durations of HV(40) and HV(25) that induced the same levels of lung injury and increased lung contents of IL-1 beta and IL-6. They were 90 min and 120 min, respectively. Both HV(40) and HV(25) increased lung SP, IL-1 beta and IL-6 levels, these effects being markedly reduced by NK-1 receptor blockade. Bivagotomy reduced to a lesser extent the HV(40)- and HV(25)-induced increases in SP but significantly reduced cytokine production. Neither vagotomy nor NK-1 receptor blockade prevented HV(40)-induced lung injury but, in the HV(25) group, they made it possible to maintain lung injury indices close to those measured in the LV group. This study suggests that both neuronal and extra-neuronal SP might be involved in ventilator-induced lung inflammation and injury. NK-1 receptor blockade could be a pharmacological tool to minimize some adverse effects of mechanical ventilation.

Figure 1. The preliminary study

IL-1β and IL-6 concentrations in lung tissue from 8 series of rats ventilated with airway pressures of 40 (HV₄₀) or 25 (HV₂₅) cmH₂O for 30, 60, 90 or 120 min. n= 3–7 observations per series for 30 and 60 min epochs and 10 observations per series for 90 and 120 min epochs. The corresponding survival rate, at each period, was calculated as the ratio of rats alive to the total number of rats. Bars represent mean ±s.e.m. cytokine concentrations (left axis); lines represent the survival rates (right axis), filled circles/bars correspond to HV₄₀ ventilation, open circles/bars represent HV₂₅ ventilation.

Figure 2

Time course of (A), airway pressure (P_aw) (B), and arterial blood pressure (ABP) (C), in animals receiving low-pressure ventilation (LV), high-pressure ventilation at 40 cmH₂O airway pressure (HV₄₀-sham) and high-pressure ventilation at 25 cmH₂O airway pressure (HV₂₅-sham). Asterisks indicate significant differences versus baseline (**P < 0.01, ***P < 0.001). # indicates significant differences between HV₄₀-sham and HV₂₅-sham groups (#P < 0.05, ##P < 0.01).

Figure 4. Quantification of SP detection by measurement of the area under the SP peaks in lung extracts

Controls: rats breathing spontaneously; LV: rats ventilated with low-pressure ventilation; HV₄₀: rats with high-stretch ventilation using 40 cmH₂O P_aw, injected with saline (HV₄₀-sham) or bivagotomized (HV₄₀-vagotomy), or after pretreatment with NK-1 receptor blockade (HV₄₀-SPB); HV₂₅: rats with high-stretch ventilation using 25 cmH₂O P_aw, injected with saline (HV₂₅-sham) or bivagotomized (HV₂₅-vagotomy), or after pretreatment with NK-1 receptor blockade (HV₂₅-SPB). Data were normally distributed. Bars represent mean ±s.e.m. Asterisks indicate significant intergroup differences (*P < 0.05; ***P < 0.001).

Figure 3

Examples of HPLC profiles expressed as absorbance intensity at 230 nm (mV) versus retention time (minutes) chromatograms in each rat group: spontaneously breathing rats (Controls), low-pressure ventilation (LV), rats ventilated with 25 cmH₂O (HV₂₅-sham) or 40 cmH₂O airway pressure (HV₄₀-sham), bivagotomized rats (HV₄₀-vagotomy and HV₂₅-vagotomy) and rats pretreated with NK-1 receptor blockade (HV₄₀-SPB, HV₂₅-SPB). The arrows indicate the peaks corresponding to substance P.

Figure 5. Individual (circles) and median (horizontal bars) lung concentrations of IL-1β and IL-6 measured by ELISA

Controls: rats breathing spontaneously; LV: rats ventilated with low-pressure ventilation; HV₄₀: rats with high-stretch ventilation using 40 cmH₂O P_aw, injected with saline (HV₄₀-sham) or bivagotomized (HV₄₀-vagotomy), or after pretreatment with NK-1 receptor blockade (HV₄₀-SPB); HV₂₅: rats with high-stretch ventilation using 25 cmH₂O P_aw, injected with saline (HV₂₅-sham) or bivagotomized (HV₂₅-vagotomy), or after pretreatment with NK-1 receptor blockade (HV₂₅-SPB). Asterisks indicate significant intergroup differences using the non-parametric test (*P < 0.05, **P < 0.01; ***P < 0.001).

Figure 6

A, comparison of respiratory system pressure–volume curves during inflation, showing the absence (NK-1 receptor blockade), or the attenuation of the rightward shift of the curve and absence of lower inflection point (LIP) in rats exposed to high-stretch ventilation with 25 cmH₂O airway pressure when vagotomized (HV₂₅-vago, grey triangles) or pretreated with NK-1 receptor blockade (HV₂₅-SPB, grey circles) in comparison with non-pretreated rats (HV₂₅-sham, black circles). Rats with low-pressure ventilation are also represented for comparison (LV, white circles). B and C, comparison of lung weight indexed to body weight (B) and lung myeloperoxidase activity (MPO; C) in rats submitted to low-pressure ventilation (LV), high-stretch ventilation with 25 cmH₂O airway pressure without (HV₂₅-sham) or after vagotomy (HV₂₅-vago) or pretreatment with NK-1 receptor blockade (HV₂₅-SPB). For box plots, the top and the bottom of each box represent the 75th and 25th percentiles; median values are represented by thin horizontal bars.