Apoptosis of sperm - a quick flow cytometry test

Apoptosis is a natural and regulated process, which usually stands at the end of live cycle of most cells in the body, including sperms. Fragmentation of DNA during apoptosis excludes apoptotic sperms from ability to give a life to a baby. Apoptosis of sperms can be iniciated also by various stres factors, such as high temperature, infection, presence of reactive oxygen species etc. At the beginning of this irreversible process of autodestruction the early apoptotic sperm still looks like and behaves like a vital sperm, although there is no way back. Zygote formed by early apoptotic sperm with fragmented DNA is not able to develop into a viable embryo. It is thus important to analyse the amount of apoptotic sperms in the ejaculate, if there is a need of assisted reproduction.
 
The sperm apoptosis test is a quick flow cytometric method that is being used as a supplement to spermiogram. Early apoptotic cells express on their surface phosphatidylserine (PS), which can be detected by fluorescently labeled annexin V, but they are still resistant to nuclear staining with propidium iodide (PI). Viable cells do not express phosphatidylserine on their surface, and those cells, that are PS-positive and PI-positive are necrotic or late apoptotic cells. Percentage of the particular cases is being compared with standard values, like are the WHO recommendations (2010).
 
We have evaluated data from 100 man with normal spermiogram, and found that in the average there was around 8% of early apoptotic cells in their sperma. As WHO recommends that good ejaculate should contain at least 58% of acceptable sperms based on spermiogram, it means that good ejaculate should contain at least 50% of viable sperms based on sperm apoptosis test.


Fig. 1: Percentage of viable sperms (A = 27,24 %; B = 67,95 %) in the ejaculate of two normozoospermics as revealed by a quick and simple flow cytometry method.
Although both individuals had normal spermiogram, a significant portion of seemingly healthy sperms were revealed as early apoptotic or Late apoptptic / necrotic sperms. 
 

Zuzana Kratka, Ph.D.

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