What is acrosome?
The acrosome is an organelle that develops over the front half of the head in the spermatozoa (sperm) of many animals, including humans. It is a cap-like structure derived from the Golgi apparatus.
In eutheran mammals, the acrosome contains degrading enzymes (including hyaluronidase and acrosine). These enzymes break down the egg’s outer membrane called the zona pellucida and allow the haploid nucleus in the sperm cell to bond with the haploid nucleus in the egg.
This release of the acrosome or the acrosome reaction can be stimulated in vitro by substances that a sperm cell can naturally encounter, such as progesterone or follicular fluid and the most commonly used calcium ionophore A23187.
This can be done to serve as a positive control in assessing the acrosome reaction of a sperm sample by flow cytometry or fluorescence microscopy. This is usually done after staining with a fluoresceinated lectin such as FITC-PNA, FITC-PSA, FITC-ConA, or a fluoresceinated antibody such as FITC-CD46.
Functions of the Acrosome
Transformation of the Sperm Head
The acrosome is involved in the transformation (shaping) of the sperm head by the acrosome-acroplaxoma-Manchette complex. These are two important structures: acroplaxon and manchette.
The acroplaxon (made up of actin and keratin) overlays the acrosome, and its complex with manchette contributes to:
- Development of an acrosome sac
- Anchoring of the acrosome to the nuclear envelope
- Transformation of the sperm cell
By shaping the acrosome, the complex enables the acrosome to shape the sperm head during spermogenesis.
In animals such as frogs and sea urchins, proteins called primary ligands have been shown to play an important role in the recognition of gametes. Although these ligands are usually found on the plasma membrane surface, some have been identified on the acrosome, thereby involving the organelle in gamete recognition.
Once the primary ligands identify specific proteins in the canister surrounding the egg, they initiate binding.
The acrosome reaction is a critical step during gamete interaction in all species, including humans. It allows sperm to enter the zona pellucida and fuse with the oocyte membrane. Spermatozoa that cannot react to acrosomes do not fertilize intact egg cells (oocytes).
The acrosomal reaction usually takes place in the ampulla of the fallopian tube (fertilization site) when the sperm enters the secondary oocyte. Some events precede the actual acrosome reaction.
The sperm cell receives a “hyperactive motility pattern” through which its flagellum creates powerful, whip-like movements that propel the sperm through the cervical canal and the uterine cavity until it reaches the isthmus of the fallopian tube.
The sperm approaches the ovum in the ampulla of the fallopian tube through various mechanisms, including chemotaxis. Glycoproteins on the outer surface of the sperm then bind with glycoproteins on the ovum’s zona pellucida.
Sperm that do not trigger the acrosome reaction before reaching the zona pellucida cannot penetrate the zona pellucida. Since the acrosome reaction has already taken place, the mechanical action of the tail allows sperm to enter the zona pellucida, not the acrosome reaction itself.
The first stage is the invasion of corona radiata by releasing hyaluronidase from the acrosome to digest the cumulus cells surrounding the oocyte and expose acrosin bound to the inner membrane of the sperm.
The cumulus cells are embedded in a gel-like substance, which consists mainly of hyaluronic acid and develops in the ovary with the egg and helps it grow. The acrosome reaction must take place before the sperm cell reaches the zona pellucida.
Acrosin digests the zona pellucida and the membrane of the oocyte. Part of the sperm cell membrane then fuses with the egg cell membrane and the contents of the head sink into the egg. In the mouse, ZP3, one of the proteins that make up the zona pellucida, has been shown to bind to a partner molecule (the β1,4-galactosyltransferase receptors) on the sperm.
This lock and key-type mechanism is species-specific and prevents sperm and egg cells from different species from merging. The zona pellucida also releases Ca granules to prevent other sperm from binding.
There is evidence that this bond causes the acrosome to release the enzymes that allow the sperm to fuse with the egg. A similar mechanism is likely to occur in other mammals, but the diversity of zona proteins between species means that the relevant protein and receptor may differ.
Recent scientific evidence shows that the acrosomal reaction is necessary to expose a protein called IZUMO1 on the sperm: Without the reaction, the sperm can still penetrate the egg membrane through the zona pellucida but cannot fuse.
As seen in mouse studies, IZUMO1 binds to the egg protein JUNO, and once it is bound together, the sperm and egg fuse into two pronuclei. These pronuclei supply the zygote with the genetic material necessary for the formation of an embryo. Once the sperm-egg fusion is complete, phospholipase C-zeta is released from the sperm.
If everything happens normally during intrusion, the process of egg activation occurs and the oocyte is said to have been activated. It is believed that this is induced by a specific protein phospholipase c zeta. It goes through its secondary meiotic division and the two haploid nuclei (paternal and maternal) merge into a zygote.
To prevent polyspermy and minimize the possibility of a triploid zygote forming, several changes in the egg cell’s cell membranes make it impermeable shortly after the first sperm enters the egg (e.g. the rapid loss of JUNO).
Spontaneous acrosome reaction
Spermatozoa can trigger the acrosomal reaction long before they reach the zona pellucida and in vitro in a suitable culture medium. This is known as the spontaneous acrosome reaction (SAR).
It is now known that this phenomenon is, in a sense, physiologically normal in mammalian species. The acrosome reaction is induced by the passage through the Cumulus oophorus cells, mediated by the hormones secreted by them (such as progesterone, LPA, LPC).
However, the physiological role of a truly spontaneous acrosomal response occurring long before this point in the female reproductive tract or in vitro is a separate phenomenon.
In mice it has been shown to be physiologically normal and frequent. Mouse sperm that have undergone a completely spontaneous acrosome reaction can continue to fertilize eggs. In addition, the rate of spontaneous acrosome reaction is higher in more promiscuous species such as Apodemus sylvaticus, which are exposed to high levels of sperm competition.
In humans, however, it remains controversial where, due to experimental limitations, the acrosome reaction is triggered during physiological fertilization (animal studies, for example, can use transgenic mice with fluorescent sperm, while studies on humans cannot).
Studies have been conducted with the intention of relating the in vitro SAR rate in human sperm to sperm quality and the rate of fertilization. However, the overall results are mixed and do not seem to be clinically useful as of 2018.
In in vitro fertilization
When using intracytoplasmic sperm injection (ICSI) for IVF, the implantation rate in oocytes injected with spermatozoa that have undergone an acrosome reaction (~ 40%) is higher than in oocytes that have been injected with unreacted spermatozoa (~ 10%). The implantation rate is ~ 25% when both converted and unreacted sperm are injected. The delivery rate per cycle follows the same trend.
The acrosome reaction can be stimulated in vitro by substances that a sperm cell can naturally encounter, such as progesterone or follicular fluid, and the most commonly used calcium ionophore A23187.
Birefringence microscopy, flow cytometry, or fluorescence microscopy can be used to assess acrosome release or the “acrosome response” of a sperm sample. Flow cytometry and fluorescence microscopy are usually performed after staining with a fluoresceinated lectin such as FITC-PNA, FITC-PSA, FITC-ConA, or a fluoresceinated antibody such as FITC-CD46.
The antibodies / lectins have a high specificity for different parts of the acrosome region and only bind to a certain point (acrosome content / inner / outer membrane). When bound to a fluorescent molecule, regions in which these probes are bound can be visualized. Sperm with artificially induced acrosome reactions can serve as positive controls.
For fluorescence microscopy, a swab of washed sperm is taken, air-dried, permeabilized, and then stained. Such a slide is then viewed under light of a wavelength that causes the probe to fluoresce when bound to the acrosomal region.
At least 200 cells are randomly considered and classified either as an intact acrosome (fluorescent light green) or as acrosome-reacted (no probe available or only in the equatorial region). It is then expressed as a percentage of the cells counted.
For assessment with flow cytometry, the washed cells are incubated with the selected probe, possibly passed through again and then removed in a flow cytometer. After gating the cell population for forward and side scatter, the resulting data can be analyzed (e.g., mean fluorescence compared).
This technique could also include a viability probe such as propidium iodide (PI) to exclude dead cells from acrosome scoring, as many sperm cells spontaneously lose their acrosome when they die.