On the other hand, the prevalence of gaps in the septal interstitium of the two-week specimen was strikingly increased over that of controls (Fig 4; Table 4). Most of these gaps were occupied by type to severe emphysema which were produced by papain.
Gil and Weibel and Brain et al have shown the necessity of the vascular perfusion method of lung fixation, in order to preserve the alveolar surface lining and to maintain the alveolar macrophages in situ in the alveolar surface lining, findings we also have confirmed. This of course necessitates the removal of erythrocytes and leukocytes from the pulmonary vasculature, including the alveolar capillaries.
Montages of serial sections of whole alveoli were found to be essential for identifying relationships between septal gaps, epithelial cells and macrophages because a gap related to an alveolar cell or macrophage might appear on only one or two sections, whereas the cell often could be identified over an extent of 15 or more sections. This method severely limited the number of samples and hence the amount of lung tissue which could be studied because it is so laborintensive. The use of India ink as a marker allowed us to sample those areas of the lung parenchyma which the papain (or saline) had demonstrably reached.
Figure 4. Relative prevalence of type 2 cells and of type 2 cells in interstitial septal gaps per 1,000 ftm of alveolar surface area. C = control animals; S = saline control animal; P= papain-treated animals (with duration of time after treatment until sacrifice indicated below P). The solid line connects points representing the prevalence of type 2 cells in each specimen and the dashed line indicates the prevalence of type 2 cells in interstitial septal gaps. Note the similarity in patterns between total number of type 2 cells and number of type 2 cells in interstitial septal gaps.
Table 4—Prevalence of Pores and Gaps
|Untreated Control A||Untreated Control B||SalineControl||Papain 4 h||Papain 2 Weeks||Papain 4 Weeks|
|No. associated with cells||19||13||49||23||71||83||24||10||46||85||22||75|
|No. located at alveolar||62||43||110||51||57||66||105||43||44||81||22||75|
|Other Septal Gaps|
|No. associated with cells||19||86||27||49||15||52||29||64||100||86||45||92|
|No. located at alveolar||12||55||29||.53||15||52||27||60||90||78||45||92|