Ventilator induced lung injury (VILI) results from mechanical trauma and secondary inflammatory responses. Lung protective ventilation attenuates cytokine concentrations and inflammatory infiltrates in the lungs [1, 2]. Previous imaging studies have focused on the effects of ventilator settings on structural or functional changes; however, no MRI studies have been conducted to elucidate the metabolic changes that occur during VILI. In this study, the conversion of HP [1-¹³C]-pyruvate and to [1-¹³C]-lactate was investigated as an in-vivo marker of lung inflammation was investigated in a two-hit model of acid aspiration and VILI.Five Sprague-Dawley rats (402 ± 85g) were ventilated with tidal volume (TV) of 6mL/kg and positive end-expiratory pressure (PEEP) of 10cmH₂O (protective ventilation) for t = 60min, until tracheal hydrochloric acid (pH 1.5, 2.0 mL/kg) instillation at t = 70min. The ventilation scheme was maintained until t = 150min. Then n = 2 rats were ventilated for 90 minutes with TV = 12mL/kg and PEEP=3cmH₂O to induce VILI; the remaining 3 rats continued with protective ventilation until the end of the experiment. A summary of the experimental setup is shown in Figure 3. Rats were imaged in supine position using a ¹H/¹³C Quadrature birdcage coil (m2m) at 4.7T (Varian Inc.). Proton images were acquired using a multi-slice gradient echo (TR/TE = 120/4ms, α=20°) images (128x128 voxels). HP [1-¹³C]-pyruvate imaging was conducted at t = 60min (baseline), t = 150min (post-HCl), and t = 240min (post-second-hit). ~22 µL of [1-¹³C]-pyruvate (Cambridge Isotopes Inc.) was polarized using HyperSense DNP polarizer (Oxford Instruments) to over 15%. The sample was melted using 4mL of a dissolution buffer (40mM Trizma, 160mM NaOH, 50mM NaCl, 0.1 g/L EDTA) at 180°C to yield an isotonic solution of 80mM [1-¹³C]-pyruvate with neutral pH at 37°C. 10 seconds after the dissolution 3mL of the hyperpolarized agent was administered via the tail-vein over 12 seconds. Three single-slice axial carbon-13 chemical shift images were acquired using a 16x16 single-point CSI sequence with (TR/TE=110/0.35ms, α=12°), 25 seconds after the start of the injection (in-plane FOV = 50x50mm², 25mm slice thickness). This yielded an in-plane resolution of 3.1x3.1mm². A 1.5M [1-¹³C]-sodium propionate (Cambridge Isotopes) sample (185ppm) was placed inside the coil to ensure proper overlay of the proton and carbon images. Three 10s breath-holds were applied to minimize the respiratory motion artifact during carbon imaging (28s). Blood oxygen level was monitored using a pulse oximeter during the scan to ensure sufficient oxygenation (SPO₂ > 90%). A custom outward spiral k-space phase-encoding order was used to acquire the center of the k-space every eleventh acquisition. Multiple FIDs acquired at k_xy = 0 were used to partially compensate the signal loss due to T₁ decay and mitigate the associated blurring artifact to improve localization of the spectra. Metabolites were quantified by integrating the peaks in the real spectra after zero and first order phasing and 4^th-order polynomial baseline correction. Spectroscopic data were processed using custom routines in MATLAB2014b (MathWorks Inc.). Processed carbon images were converted to DICOM format and overlaid on the proton image using OsiriX 7.0 (Figure 1).Figure 1 shows images of the metabolites in a rat post HCl injury. The intensity of each image is normalized to its own maximum intensity. The images suggest global injury in the lungs. Although the signal intensity in the heart is dominant (and in the muscle for alanine), the combination of breath-hold and de-blurring using the custom k-space trajectory allows for proper localization of metabolites to the lungs (Figure 2. Lactate-to-pyruvate ratio was measured from the averaged spectra in the voxels over the lungs (highlighted voxels in Figure 2). The average lactate-to-pyruvate ratio as a function of time is shown in Figure 3. Our measurements show an increase of 20.8 ± 12.1% in the lactate-to-pyruvate ratio after HCl instillation, compared to the baseline. The observed effect size for this measurement was 1.79 (Cohen’s d). This initial effect was likely due to the presence of inflammatory infiltrate [3]. Furthermore, our results indicate a further increase in the lactate-to-pyruvate ratio after exacerbation of the injury due to injurious ventilation leading to VILI (43.0 ± 21.1% greater than HCl cohort). This increase was significantly smaller in rats that underwent protective ventilation (14.1 ± 10.8% greater than HCl instillation).This study demonstrates that HP [1-¹³C]-pyruvate imaging can be used as a marker for metabolic changes due to lung injury in this two-hit model of acid aspiration and VILI, providing critical information for detection and management of acute lung injury that is otherwise difficult to acquire from other conventional modalities.