Videomorphometric Analysis of Hypoxic Pulmonary Vasoconstriction of Intra-pulmonary Arteries Using Murine Precision Cut Lung Slices

Abstract

Acute alveolar hypoxia causes pulmonary vasoconstriction (HPV) -also known as von Euler-Liljestrand mechanism -which serves to match lung perfusion to ventilation. Up to now, the underlying mechanisms are not fully understood. The major vascular segment contributing to HPV is the intra-acinar artery. This vessel section is responsible for the blood supply of an individual acinus, which is defined as the portion of lung distal to a terminal bronchiole. Intra-acinar arteries are mostly located in that part of the lung that cannot be selectively reached by a number of commonly used techniques such as measurement of the pulmonary artery pressure in isolated perfused lungs or force recordings from dissected proximal pulmonary artery segments 1,2 . The analysis of subpleural vessels by real-time confocal laser scanning luminescence microscopy is limited to vessels with up to 50 µm in diameter 3 .We provide a technique to study HPV of murine intra-pulmonary arteries in the range of 20-100 µm inner diameters. It is based on the videomorphometric analysis of cross-sectioned arteries in precision cut lung slices (PCLS). This method allows the quantitative measurement of vasoreactivity of small intra-acinar arteries with inner diameter between 20-40 µm which are located at gussets of alveolar septa next to alveolar ducts and of larger pre-acinar arteries with inner diameters between 40-100 µm which run adjacent to bronchi and bronchioles. In contrast to real-time imaging of subpleural vessels in anesthetized and ventilated mice, videomorphometric analysis of PCLS occurs under conditions free of shear stress. In our experimental model both arterial segments exhibit a monophasic HPV when exposed to medium gassed with 1% O 2 and the response fades after 30-40 min at hypoxia.

CD146-HIF-1α hypoxic reprogramming drives vascular remodeling and pulmonary arterial hypertension

“…The hypoxia- or drug-induced vasoconstriction of small PAs was determined by videomorphometric analysis of cross-sectioned arteries in precision cut lung slices 61 . In brief, the lungs were filled with low melting point agarose and cut into 200 μm thick precision cut lung slices (PCLS) using vibratome.…”

Phosphocholine-Modified Lipooligosaccharides of Haemophilus influenzae Inhibit ATP-Induced IL-1β Release by Pulmonary Epithelial Cells

“…571_M). Mouse PCLS were prepared using a modified protocol described previously [ 56 , 57 , 76 ]. Briefly, lungs were taken from 8 to 12 weeks old female C57BL/6NRj mice.…”

“…After 24 h of incubation, PCLS were treated with LPS (100 ng/mL), recombinant human IFN-γ (20 ng/mL; R&D Systems, Minneapolis, MN, USA) and recombinant human TNF-α (10 ng/mL; R&D Systems) for additional 24 h. As a control PCLS were left untreated for the same period of time. The functional integrity of PCLS produced according to the same protocol in the same laboratory was published recently [ 76 ]. Some of the untreated PCLS were fixed for 24 h in 4% paraformaldehyde (Carl Roth, Karlsruhe, Germany) and lightly stained with Richardson’s blue as described before [ 77 ].…”

Pre-clinical assessment of a water-in-fluorocarbon emulsion for the treatment of pulmonary vascular diseases

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