A Note on Differentiation and Divisibility of Alveolar Epithelial Cells

should extend vertically from the apical to the basolateral cell surface, a process which seems easily possible in the cuboidal type 2 cell or in the simple squamous type 1 cell (Fig la, Ib, 2a). However, in the topologically more complex type 1 cell (Fig Ic, Id, 2b), it would be very difficult to conceive how a nuclear division could be followed by a cytoplasmic division, which adequately divides the apical and basoalteral cell surfaces. This is rendered even more difficult by the fact that the cytoplasmic flaps of type 1 cells extend over long distances from the nucleus. In contrast, the type 2 cell is predominantly a simple cuboidal cell, although some cells with two or even three apical faces have also been observed, but never with the same level of complexity as the type 1 cells." For the majority of these cells there is, therefore, no difficulty in adequately dividing, whereby the division plane passes across the apical and the basolateral face." Their cytoplasmic diHerentiation does not seem to be an obstacle, since it can be shown that the specific lamellar bodies are retained in mitotic cells and distributed among the two daughter cells. S In conclusion, we have demonstrated that type 2 cells have a high level of cytoplasmic but a low level of topologic diHerentiation, whereas the type 1 cells combine a low level of cytoplasmic with an exceptionally high level of topologic diHerentiation. Considering the process of mitotic cell division, it seems plausible that

type 1 cells should have great difficulty in proper division of its elements of the surface membrane. This could explain the fact that type 1 cells cannot divide, but that damaged type 1 cells are replaced by division and subsequent transdiHerentiation of type 2 cells. If this hypothesis can be proved, it would imply that the level of topologic rather than that of cytoplasmic diHerentiation is the major obstacle to mitotic divisibility.

DISCUSSION

trations may have produced significant ultrastructural alterations, especially in the mitochondria. I have been evaluating the structural alterations or damage which can be produced in rat liver cells if the tonicity of the fixing solutions is altered (Acta histochem 35: 1-17, 1970). It seems reasonable to assume that similar damage could be induced in pulmonary cells also. If the buDer used with formaldehyde was isotonic or slightly hypertonic, best results were achieved. However, when dilute buffer (0.05-0.1 M) was used with 4 percent formaldehyde or glutaraldehyde preceding the osmium tetroxide postfixation, marked differences, including dilated endoplasmic reticulum, swollen mitochondria and even ruptured mitochondrial membranes were observed in the specimens. This was true even if the tonicity of the fixative brought the osmolarity of the entire solution to isotonic or hypertonic levels. This observation diHers from the usual custom of carefully controlling the tonicity of the buffer and fixative together, ie keeping it nearly isotonic. I found that during the initial fixation, the combined tonicity of buffer and fixative was less critical than the concentration of the buller alone. Postfixation osmium tetroxide could not reverse the damage induced by the earlier use of a dilute buffer. Dr. Weibel: These are critical considerations, about which we have been very concerned also. We measure the osmolarity of each batch of fixative prepared, by the method of freezing point depression, and routinely use an osmolarity of 350 mosm/L. Another point which should be made is that the buffer employed in the fixative is important; we have found, for example, that glutaraldehyde buffered with collidine may produce holes in the tissue sections.

Dr. Stanford: Using the Cowdry terminology, are type 1 alveolar epithelial cells fixed postmitotic cells, or can they, under certain conditions, revert to the status of a dividing cell, similar to hepatocytes? Dr. Weibel: I will refer the question to Dr. Evans, who has done a great deal of work in this area recently. Dr. Evans: We have seen occasional cells with the morphologic features of type 1 cells 'labeled with tritiated thymidine label in young animals, but as yet not in adult animals, even under pathologic conditions. I would like to ask Dr. Weibel a question: What was the regional distribution of the lesions you described in your patient's lungs? Dr. Weibel: The lesions were multifocal, but we could not find any particular relationship to the pulmonary acinus or lobule. Dr. MitcheU: Is cellular proliferation in the pulmonary interstitium characteristic of oxygen toxicity? Dr. Weibel: Not necessarily; it may.bepresent in that and a wide variety of other pathologic processes, as Dr. Stanford mentioned in the introductory remarks this morning, and as I have shown in this series of patients who did not receive high-concentration or longtenn oxygen. Dr. Winter: What was the immediate cause of death in your patients? Dr. Weibel: Acute respiratory failure with impaired gaseous diffusion and consequent systemic hypoxemia. Dr. Paegle: I am concerned that during your presentation and those of several preceding speakers, who performed ultrastructural morphometric studies, relatively little was said about the precautions taken to exclude the possibility that variations in buffer and fixative concen-

CHEST 65: 4, APRIL, 1974 SUPPLEMENT

1 Bachofen M, Weibel ER: Basic pattern of tissue repair in

human lungs following unspecific injury. Chest (this issue) 2 Carrington CB, Green TJ: Granular pneumocytes in early repair of diffuse alveolar injury. Arch Intern Moo 126:464465, 1970 3 Evans MJ, Cabral LJ, Stephens RJ, et al: Renewal of alveolar epithelium in the rat following exposure to N02. Am J Pathol 70:175-198, 1973 4 KauHman SL, Burri PH, Weibel ER: The postnatal growth of the rat lung. Part 2: Autoradiography. (Submitted to Anat Rec) 5 Kolliker A: Zur Kenntnis des Bans der Lunge des Menschen. Verhandl Physiol Moo Ges (Wiirzburg) 16:1,

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6 Meyrick B, Reid L: The alveolar brush cell in rat lung-a third pneumocyte. J Ultrastruct Res 23:71-80, 1968 7 Weibel ER: The mystery of "non-nucleated plates" in the alveolar epithelium of the lung explained. Acta Anat (Basel) 78:425-443, 1971

18TH ASPEN LUNG CONFERENCE 215