Local oxygen concentration reversal from hyperoxia to hypoxia monitored by optical-resolution photoacoustic microscopy in inflammation-resolution process

Abstract

Current consensus suggests a simultaneous occurrence of hypoxia and inflammation. For the first time, we observed a hyperoxia state during the initiation stage of gouty arthritis (GA) via optical-resolution photoacoustic microscopy. GA as a paradigm of acute sterile inflammation, has been regarded as a single process. However, our experimental results demonstrated that the onset-resolution inflammation process composed of two sub-processes with different features. In the initial sub-process, inflammation and resolution events appear in hyperoxia state (1st-5th days). In the subsequent sub-process, post-resolution events appear in hypoxia state (6th-15th days), which is related with the second wave of immune response. Furthermore, we demonstrated that the inflammatory cytokines together with hyperoxia levels in initial sub-process can be downregulated by photobiomodulation, resulting in the hypoxia levels in subsequent sub-process were inhibited. Our results unveiled the detailed progress of GA and provided potential non-invasive monitoring and treatment strategies.

Keywords: Acute sterile inflammation; Gouty arthritis; Hyperoxia; Optical-resolution photoacoustic microscopy; Photobiomodulation; Spontaneous resolution.

Fig. 1

(a) Optical-resolution photoacoustic microscopy was used to monitor blood vessel morphology and oxygen saturation of SD rat ankle for 15 days. A ring-shape LED light source of 627 nm was used to irradiate rat ankle. (b) Acute inflammation was triggered by MSU crystal deposition with vasodilatation and inflammation cytokines accumulated. (c) An external light irradiation composed of LED alleviated the inflammation symptoms.

Fig. 2

Optical path diagram of OR-PAM system. HWF: half-wave plate, NDF: neutral density filters, PBS: polarizing beam splitter, M: mirror; FC: fiber coupler, DM: dichroic mirror, MMF: graded-index multimode fiber, PM-SMF: polarization-maintaining single-mode fiber, FMF: few-mode optical fiber, OL: optical lens, UST: ultrasonic transducer, AL: acoustic lens, WT: water tank.

Fig. 3

Observation of GA development in SD rats in 24 h. (a) Experimental schedule of monitoring. (b) Photos of rat ankle before and after MSU injection. (c) Swelling index and paw withdrawal threshold (PWT) of Von frey of GA ankle in 24 h. d) PA images of ankle blood vessels within 24 h. (e) Diameter and density of blood vessels in GA ankles. (f) sO₂ images of ankle blood vessels within 24 h. (g) sO₂ level of ankle blood vessels within 24 h with dashed line indicating the normoxia level. (h) Ultrasonography scan of GA ankles at various time points with dashed line indicating MSU area. (i-k) Concentration of inflammatory cytokines in blood serum: IL-1β, IL-6 and TNF-α respectively. Animal number n = 6, mean ± SD; *p < 0.05, * *p < 0.01, * **p < 0.001, * ** *p < 0.0001.

Fig. 4

Observation of GA inflammation-resolution in rats in 15 days. (a) Experimental schedule of the observation. (b) Photos of rat ankle. (c) OR-PAM images of morphology of ankle blood vessels. (d) sO₂ images of ankle blood vessels. (e) Swelling index of GA ankle. (f) PWT value of von Frey filament test. (g) Averaged diameter of blood vessels in GA ankles. (h) Averaged density of blood vessels in GA ankles. (i) Averaged s sO₂ level of ankle blood vessels with dashed line indicating the normoxia level. (j-l) Concentration of inflammatory cytokines: IL-1β, IL-6 and TNF-α. Animal number n = 6, mean ± SD; *p < 0.05, * *p < 0.01, * **p < 0.001, * ** *p < 0.0001.

Fig. 5

Effect of external light irradiation on GA rats in 15 days. (a) Experimental schedule of the external light irradiation. (b) Photos of rat ankle in GA+Light group. (c) OR-PAM images of morphology of ankle blood vessels. (d) sO₂ images of ankle blood vessels. (e) Ankle swelling index of GA rats. (f) PWT value of von Frey filament test. (g) Comparison of vessel diameter changes between GA and GA+Light group. (h) Comparison of changes in vascular density between GA and GA+Light group. (i) Comparison of blood oxygen saturation between GA and GA+Light group, showing a hyperoxia and hypoxia sub-processes, with dashed line indicating the normoxia level. The statistical analysis of e-i is presented in Tab. S1–5. Animal number n = 6, mean ± SD.

Fig. 6

Effect of external light irradiation on GA rats in 15 days. (a, b) Ultrasonography scan of GA and GA+Light group over 15 days. Yellow dashed lines represent MSU accumulation. (c) MSU area of GA and GA+Light group in 15 days. (d-f) Analysis of serum inflammatory cytokines IL-1β, IL-6 and TNF-α at various time points. Animal number n = 6, mean ± SD; *p < 0.05, * *p < 0.01, * **p < 0.001, * ** *p < 0.0001.