Vertical and middle band banding in alveolar type II cell surfactant protein granules is shown here (hopefully this is SP-A). Using the same image as posted as the quintessential surfactant protein granule in alveolar type II cells, the image has been copied, inverted, and offset just a few pixels, to cast a shadow and give dimension to the “edges” in the image, a kind of embossing technique.

Embossing vertically highlights the bumpiness of ribosomes and other protein “lumps” within the granule, while embossing by moving the inverted image below, causes the ribosomes to appear to be concave.  Both accentuate different features (see below). The embossing upward as well as moving the inverted image a few pixels to the left highlight the banding seen within the granule which is secondary to the electron dense bands that run parallel to the long dimension of the granule… the vertical lines are not present in the surrounding cytoplasm, thereby indicating that the vertically lines within the granule are not section artifact or fixation artifact but actually a part of the specific alignment of the granule proteins.  The third image is a flattened image, cropped, increased in brightness and contrast, then vertical lines were superimposed where I saw definition.  Ribosomes, as an internal control for magnification are about the size of the red marker (27 nm estimated diameter), black line of 300 nm = the height of the three banding periods comprising this particular granule, 100 nm bars mark out the middle dense layer (with the blue dots where periodicity shows up) and also shows that about 4-5 protein densities occur in the middle layer of each 100 nm period.

Original ferret alveolar type II cell RER granule electron micrograph, untouched, from which these processed images were derived can be found here.

Negative 9879 block 23494 ferret # 16, untreated. Electron micrograph of an alveolar type II cell with a surfactant protein granule (which in some ways is similar to other collectin, c-type lectin, style granules, where organization of these proteins is highly oligomerized–e,g, Birbeck granules).  Embossed upward, and down ward images below. Bottom image is a cropped and enlarged from middle image, showing vertical lines which may be 4 or 5 per 100 nm and can transcend all three periods of this granule; ribosomes (as a size marker of @27 nm); periodicity of the medium dense layer of this granule (blue dots and 100 nm bar marker).

Alveolar type II cell intra-RER protein organization: ferret, more hexagonal structures per the last two posts, there is more order here,  actual linear densities perpendicular to the long axis of the layered granule, and these blend out into the tangential plane as hexagonal structures with a central dense area (either 1 or 2 dots — in this case mostly 1 dot).

Top electron micrograph (unretouched, except to show the bounding box of the inset image) is an enlargement from a negative (9855) of a ferret type II cell, of just one of the layered granules present which is particularly well suited to examination of its tangential-orientation and detection of patterns that might indicate that it is surfactant protein A.

Bottom electron micrograph is inset, enlarged, and the patterning perpendicular to the long axis of the granule layering (here, tangentially cut) is seen, and the erase tool has been used to outline what could be hexagonal patterns for the surfactant protein A 18-mer bouquet.  You are free to decide if this looks right. My question is that they are small, but also would represent just 1/4 of the 100 nm width of the real pattern-period, of mirrored and stacked (4 molecules) of SP-A 18-mers. I have not highlighted all the vertical lines, nor have i highlighted all the hexagonal structures.

More tangential sections of RER layered protein from ferret alveolar type II cells seen with electron microscopy. I have chosen one of the more easily identifiable granules with the light portion of the banding ‘periods’ spread out so that any order seen on from the “top down” might be highlighted.  I have outlined in red (this is a “partial erase” in photoshop, against a red (and green) layer), no other manipulation of the micrograph has occurred.

The red outlines are pretty obvious hexagonal structures, seemingly mostly with a central dense area, but way to often for chance, two central dots.  I have seen this so many times that it becomes necessary to call attention to it, and likely it means something in the organization of this surfactant protein (which I am calling surfactant protein A). On the other hand, the size of a ribosomes in these micrographs would make the hexagons a little too small….  maybe only 75- 80 nm across, so this is an issue to reconcile.

Red outlines, within the tangentially spread inbetween layers), green outlines, an area which might be still within this particular RER profile, but might actually be cytoplasmic.  I have compared the incidence of cytoplasmic heaxagonal structures, there are some, no question, and I don’t want to read more into the hexagonal molecular organization than is warranted.  To me however, it is significantly greater within the membrane of the RER when tangentially sectioned as happened here, and the previous post (same animal, same micrograph, same magnification, different site.

The granule in the alveolar type II cell of guinea pig, ferret and dog is unique, in that it appears to have wonderful shapes, concentric, U-shaped, linear, branching (even highly branching mostly at perpendicular planes to the original layering). I was trying to figure out what could cause such variation in shape. One keeps in mind that these are cross-sectional images, 2D of the 3D cytoplasmic granules, and therefore can be difficult to interpret as a whole granule.

To note first: there are often parallel “jets” (which I have called periods, just to be proper ) for the production of surfactant protein A (yep, I am calling it that with nothing but circumstantial evidence). Sometimes there are 10 or 15 such nidi of production, each perpendicular to the long axis of the layering (banding) and having about 4 ribosomes (the 4th ribosome in each period serves as the 1st in the adjacent layer).  The amount of resistance at the opposite end of the granule to protein synthesis, and the rate of protein synthesis (at the growing end(s) of the granule where the ribosomes are ) will determine whether this “jet” will spew out SP-A in an unhindered stream to create a linear granule or whether it will buckle, twist, spin or curl.

I began searching extrusion images on google, and these below (easily-found set of 6   extrusion images which illustrate the point) do a really good job of matching what is seen electron microscopically, of what becomes of the structure of protein layering during SP-A production when the protein produced meets with resistance.  Awesome… ha ha, a natural physical phenomenon, I will bet. What scientific journal will publish my diagram… ha ha, probably not.  Too funny.