LUNG PHENOTYPES
METHACHOLINE_ED50 (mg/kg).
Methacholine challenge is commonly used to detect differences in airway reactivity between normal
and hypersensitive animals or human subjects. Methacholine stimulates muscarinic receptors that
mediate airway smooth muscle contraction, which can then be detected by measuring pulmonary
conductance. The ED-50 is the dose of methacholine that decreases the pulmonary conductance to
one half its pretreatment value as estimated by fitting a standard sigmoid function to the pulmonary
conductance vs. methacholine dose data.
R_FLOW100 (mmHgminkgml1)
The pulmonary vascular resistance (R) at a flow rate (F) of 100ml/min per kg body weight is used as
a measure of pulmonary vascular function, particularly with regard to the vascular remodeling
during hypoxia. Isolated lungs are ventilated (6% CO2 and 15 % O2 in N2) and perfused
(physiological salt solution containing 4 % bovine serum albumin). The flow rate (F) is increased to
40 ml/min and then progressively decreased to zero while the perfusion pressure (P) is monitored.
The equation P = ((1+B**F)1/51)*, derived from a distensible vessel model of the pulmonary
vasculature, is fit to the resulting pressure vs. flow data to estimate the vascular resistance (P/F) at F
= 100ml/min per kg body weight.
ALPHA (mmHg1)
The value of  obtained from the pressure vs. flow data used to estimate R_FLOW100 is a measure
of the contribution of the distensibility of the resistance vessels to the pressure vs. flow relationship.
ALPHA can be thought of as the fractional change in resistance vessel diameter per mmHg change
in intravascular pressure. A lower value of  indicates less distensible vessels.
FAPGG_MSAP_PRODUCT (mlmin1kg)
FAPGG (furylacrylolphenylalanylglycylglycine) is a substrate for angiotensin converting enzyme
(ACE), which is particularly abundant on the luminal surface of the pulmonary capillary
endothelium. ACE cleaves the glycylglycine, which decreases the absorbance of an FAPGG
solution at 340 nm. To measure the hydrolysis of FAPGG during transit through the lung, 0.6mM
FAPGG is added to the perfusate entering the pulmonary artery of the isolated lungs while the lungs
are perfused at a flow rate (F) of 10 ml/min. After 1 min the absorbance (340nm) of the perfusate
entering the pulmonary artery and exiting the pulmonary veins is measured, and the fraction (f) of
FAPGG hydrolyzed during passage through the lungs is calculated using the difference. To convert
f into a quantity proportional to the first order rate of hydrolysis, and normalizable to animal (lung)
size, the Crone equation is used to convert f to the metabolism-surface area product (MSAP) where
MSAP = F*ln(1f). MSAP has units of ml/min and is the perfusate flow rate at which f would
equal 0.63. Also MSAP/0.693 is the flow rate that would result in half the infused FAPGG being
hydrolyzed.
MB_MSAP1_PRODUCT (mlmin1kg)
and
MB_MSAP3_PRODUCT (mlmin1kg)
Methylene blue (MB+) is a hydrophilic redox indicator dye that is reduced on the endothelial cell
surface. The reduced, colorless, lipophilic form (MBH) is then able to permeate the cell membrane.
Once inside the cells the reduced form undergoes cyanide sensitive reoxidation and becomes trapped
within the cells. The extent to which these reduction and oxidation reactions take place provides an
index of the redox status of the cells. When infused into the pulmonary artery of the lungs a fraction
of the MB+ emerges in the venous effluent as the originally infused MB+, another fraction exits as
the reduced and colorless MBH, and another fraction remains in the lung. To measure these
fractions, MB+ (10 M) is added to the perfusate entering the pulmonary artery. After 1 min the
absorbance (665 nm) of the perfusate entering the pulmonary artery and exiting the pulmonary veins
is measured, and the fraction (f3) of MB+ lost (due to reduction and/or uptake into the tissue) during
passage through the lungs is calculated. Any MBH in the venous effluent is then allowed to autooxidize and the absorbance is measured again. The venous effluent MB+ absorbance after autooxidation compared to the inflow absorbance reveals the fraction (f1) that did not emerge from the
lungs. To convert f1 and f3 into quantities proportional to the first order rate of uptake or reduction,
and normalizable to animal (lung) size, the Crone equation is used to convert f1and f3 to the
metabolism-surface area products MB_MSAP1 and MB_MSAP3, respectively, where MB_MSAP1
= F*ln(1f1), and MB_MSAP3 = F*ln(1f3). The meaning of MSAP is the same as indicated for
FAPGG. The assumption is that all of the MB+ lost from the perfusate on passage through the lungs
is the result of reduction (the oxidized form being confined to the perfusate). Thus, MB_MSAP3 is
the measure of the net reduction rate, and MB_MSAP1 is the measure of the rate that the methylene
blue (as MBH) was taken up by the lung.
MB_MSAP3_FAPGG_MSAP_RATIO
The ratio of MB_MSAP3 to FAPGG_MSAP is presented as a phenotype because each of these two
individual phenotypes has the potential for reflecting a difference in either the intensive or extensive
properties (or both) of the reaction processes involved. For example, a difference could result from
more or less enzyme activity per unit endothelial surface area or from more or less endothelial
surface area. Examining the ratio of the rates of two processes having apparently unrelated intensive
properties but having a common extensive property (surface area), increases the chance that a
difference in one or the other can be attributed to the intensive property of interest. In other words,
if both MB_MSAP3_and FAPGG_MSAP were proportionately higher in one strain than another,
then a higher surface area would be suspected. On the other hand, divergence between the two
individual phenotypes reflected in a difference in the ratio would suggest a more specific
difference(s) with regard to the processes involved.
BODY_WEIGHT (kg)
LUNG_DRY_WT_BODY_WT_RATIO
A measure of relative lung size.
RV_LV_RATIO
A measure of right heart hypertrophy during exposure to hypoxia.
HEMATOCRIT (%)
A measure of the hemopoetic response to hypoxia.