Morphometric Description of Spermatogonial Stem Cells and Expansion of Their Clonal Derivatives - Male Germline Stem Cells Developmental and Regenerative Potential - page 94

Fig. 4.3 Schematic of a seminiferous tubule showing the wave of spermatogenesis through which
epithelial stages ( roman numerals ) follow each other sequentially. The differentiating type of
spermatogonia (A2, A3, A4 are indicated) also follow each other and go through the phases of the
cell cycle in an epithelial stage related order. G1, G2, S, M - phases of the cell cycle
cell cycle, somewhat further along the tubule one can see these cells become
bigger as they carry out S phase and still further they will be in G2 phase and
show some heterochromatin at the nuclear membrane. Subsequently, the cells
enter mitosis and A4 spermatogonia are formed. These A4 spermatogonia are
small at first as they are in G1 phase and then again further along the tubule, they
will become bigger and so on. This synchronization originates not only from the
intercellular bridges between cells from the same clone but also from a synchro-
nization of the cell cycle progress of all clones in the same area (Lok and de
Rooij 1983a ). Hence, following the A1-B spermatogonial generations along the
length of a tubule, one encounters relatively extended fields in which all sper-
matogonia from an A1-B spermatogonial cell type are in late G2 phase of the
cell cycle or are in mitosis. In contrast, the clones of A s,pr,al spermatogonia always
proliferate at random, and separate clones in a tubule area do not cycle synchro-
nously with each other and do not follow the behavior of the differentiating type
spermatogonia. These phenomena can be used to easily discern differentiating
type A spermatogonia from A s,pr,al spermatogonia (de Rooij 1973 ; Lok et al.
1982 ; Tegelenbosch and de Rooij 1993 ).
Hence, A s,pr,al spermatogonia can best studied in those areas in which the genera-
tions of differentiating type A spermatogonia are synchronously in G2 phase of the
cell cycle or in mitosis (Fig. 4.3 ). In these areas the type of differentiating type A
spermatogonia present are big and show some heterochromatin or are in mitosis
and the A s,pr,al spermatogonia stand out from the differentiating A spermatogonia as
they generally have much smaller nuclei because they are not in G2/M. The cells
composing the occasional clones of A s,pr,al spermatogonia that also happen to be in
G2/M are much closer to each other than the differentiating type A spermatogonia.
An additional advantage of this procedure is that when one studies A s,pr,al sper-
matogonia in areas in which the subsequent generations of A1, A2, A3, A4, In, and
B spermatogonia are in G2/M, one studies these cells at regular intervals of time
because the cell cycle time of the A1-B spermatogonia is always similar (see
below). Cell counts are usually made using the numbers of Sertoli cells present in
the same area as a reference, and numbers are then usually expressed per, for
example, 1,000 Sertoli cells.