Laminin gamma2 fragments are increased in the circulation of patients with early phase acute lung injury

Abstract

Objective: Laminin-5, a cell adhesive molecule expressed solely by epithelium, is known to enhance epithelial cell migration and repair of injured epithelium, after its essential component gamma2-chain is processed proteolytically. Our previous study revealed circulating levels of amino-terminal fragment of laminin gamma2-chain (G2F) reflect epithelial tumor invasiveness in carcinoma patients, but its physiological role in alveolar epithelial injury remains unknown.

Design: Sampling of epithelial lining fluids or pulmonary edema fluids from patients with acute lung injury (ALI) or related diseases was performed. Plasma samples were obtained from them at the time of disease onset or later. G2F concentrations were determined by immunoassay constructed by ourselves.

Results: We found a significantly higher amount of G2F in pulmonary edema and epithelial lining fluids of patients with ALI, as compared with those with the other respiratory diseases. Their plasma levels were also elevated significantly early at the onset of ALI (mean +/- SD; 147 +/- 82 ng/ml in non-surviving and 90 +/- 56 in surviving patients) as compared with those in the patients with cardiogenic pulmonary edema (59 +/- 36) or idiopathic pulmonary fibrosis (37 +/- 17), indicating alveolar epithelium rapidly secrete laminin-5 in ALI. At 5 days after onset, non-surviving patients maintained higher plasma concentrations (152 +/- 84), but in contrast, the levels in surviving patients declined (71 +/- 35), suggesting secretion of laminin-5 was suppressed, associated with recovery from ALI.

Conclusion: Circulating G2F may be a biomarker for alveolar laminin-5 secreted early at disease onset in ALI, potentially regulating alveolar re-epithelialization.

Fig. 1

Schematic representation of ELF collection procedure. Detailed procedure is described in “Materials and methods.” Briefly, a flexible fiberoptic bronchoscope was inserted into the lungs. The bronchial micro-sampling probe, with the cotton probe as absorptive materials, was inserted into the lungs through the bronchoscope for absorbing the ELFs. The wet inner probe was sectioned, introduced in a pre-weighed tube and weighed. The diluted solution for measurements of biochemical constituents was prepared by adding 3 ml of saline to the tube containing the triplicate probes. The probe was then dried and weighed again to measure the ELF volume recovered, and the dilution factor was calculated

Fig. 2

Levels of G2Fs in ELF of subjects in the early phase of ALI. The box–whisker plots show the 25th and 75th percentiles, the median (horizontal line within the box), and the 10th and 90th percentiles (whiskers). Individual values below the 10th percentile and above the 90th percentile are shown as open circles. The mean G2F levels in the ELF samples collected from 25 patients with ALI, at the onset time of the disease (day 0), were significantly higher (*P < 0.0003) than those from 15 control subjects or those from 12 COPD patients. They were also significantly elevated (**P < 0.02) rather than those from 5 patients with CPE or those from 11 patients with IPF

Fig. 3

Comparison of G2F levels in PEF of the patients with CPE and ALI. The box–whisker plots show the 25th and 75th percentiles, the median (horizontal line within the box), and the 10th and 90th percentiles (whiskers). Individual values below the 10th percentile and above the 90th percentile are shown as open circles. The mean G2F levels in the PEF samples collected from 21 patients with ALI were moderately higher (*P < 0.05) than those from 11 CPE patients. The highest G2F value (32,197 ng/ml) was out of the range

Fig. 4

Levels of G2Fs in plasma of clinical subjects. Number of the subjects in each column was indicated in parentheses. The box–whisker plots show the 25th and 75th percentiles, the median (horizontal line within the box), and the 10th and 90th percentiles (whiskers). Individual values below the 10th percentile and above the 90th percentile are shown as open circles. The ALI patients were classified into the five groups (day 0, 1, 3, 5 or 7 after disease onset), and each of the groups was further divided into survivors (white box) and non-survivors (shadowed box). The plasma G2F levels of 15 control subjects were indicated as a black box in the left end. Statistical differences between survivors and non-survivors are indicated as follows: *P < 0.02, **P < 0.003, or ***P < 0.0002. The levels in the patients with CPE or IPF were not significantly higher than those in the control subjects