Acute Crit Care.  2021 Feb;36(1):29-36. 10.4266/acc.2020.00969.

Feasibility study of incident dark-field video microscope for measuring microcirculatory variables in the mouse dorsal skinfold chamber model

• Kang C
• Cho AR
• Lee HJ
• Kim HJ
• Kim EJ
• Jeo S
• Hong JM
• Moon D

• KMID: 2513251
• DOI: http://doi.org/10.4266/acc.2020.00969

Abstract

Background
Despite the importance of microcirculation in organ function, monitoring microcirculation is not a routine practice. With developments in microscopic technology, incident dark field (IDF) microscopy (Cytocam) has allowed visualization of the microcirculation. Dorsal skinfold chamber (DSC) mouse model has been used to investigate microcirculation physiology. By employing Cytocam-IDF imaging with DSC model to assess microcirculatory alteration in lipopolysaccharide (LPS)-induced endotoxemia, we attempted to validate availability of Cytocam-IDF imaging of microcirculation.
Methods
DSC was implanted in eight BALB/c mice for each group; control and sepsis. Both groups were given 72 hours to recover from surgery. The sepsis group had an additional 24-hour period of recovery post-LPS injection (4 mg/kg). Subsequently, a video of the microcirculation was recorded using Cytocam. Data on microcirculatory variables were obtained. Electron microscopy was implemented using lanthanum fixation to detect endothelial glycocalyx degradation.
Results
The microcirculatory flow index was significantly lower (control, 2.8±0.3; sepsis, 2.1±0.8; P=0.033) and heterogeneity index was considerably higher (control, 0.10±0.15; sepsis, 0.53±0.48; P=0.044) in the sepsis group than in the control group. Electron microscopy revealed glycocalyx demolishment in the sepsis group.
Conclusions
Cytocam showed reliable ability for observing changes in the microcirculation under septic conditions in the DSC model. The convenience and good imaging quality and the automatic analysis software available for Cytocam-IDF imaging, along with the ability to perform real-time in vivo experiments in the DSC model, are expected to be helpful in future microcirculation investigations.

Keyword

dorsal skinfold chamber model; glycocalyx; incident dark field; microcirculation; sepsis

Figure

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  Figure 1. Markings made prior to dorsal skinfold chamber (DSC) implantation (A). Completion of DSC implantation; note the placement of glass cover slip below the retaining ring (B).

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  Figure 2. Image taken after removal of the retaining ring and glass cover slip (A). Direct contact of the observational window with green light emitting microscope (B).

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  Figure 3. Electron microscopy of a cross section of dorsal skin vessel; control (A) and sepsis (B). ×20,000; white arrow, lanthanum nitrate-stained glycocalyx layer.

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