Biology Reference
In-Depth Information
developed using information drawn from the international literature (PubMed).
Starting from these data, we created a database in Microsoft Office Excel 2007.
For each metabolic event, we specified the original molecule, the target molecule,
and the type of interaction (activation, inhibition, etc.). In addition, each molecule
was assigned an attribute, depending on the specific intracellular localization within
which it operates (cytosol, membrane, mitochondria, etc.). The network was
upgraded using Cytoscape 2.6.1 software, in which molecules were arranged
depending on the intracellular localization through the Cerebral plugin v.2.
6.3 Results
Biological results showed that inhibition of actin polymerization negatively affects
the status of capacitation. In fact, exposure of in vitro capacitated sperm to sZP led to
AR in 30% of CTR spermatozoa, while it caused the same reaction in only 10% of
those treated with CD [
p
0.01 vs. CTR, analysis of variance (ANOVA)]. However,
functional analyses performed on both groups of spermatozoa demonstrated no
differences with regard to signal transduction pathways. In particular, CTC staining
revealed a similar incidence of spermatozoa with fluorescence patterns typical of
capacitated sperm in both CTR cells and those treated with CD (35.3
<
4.3 vs.
36.7
0.05, ANOVA). In the same way, the kinetics and degree of protein
tyrosine phosphorylation were similar. Furthermore, in vitro incubation promoted
in both CTR sperm and CD cells a relocation of PLC
5.5,
p
>
1 with translocation of the
enzyme from the cytoplasm to the sperm membrane. Finally, basal intracellular
calcium levels were similar in CTR and CD sperm, and the rapid increase of [Ca
2+
],
induced by exposure to sZP, was detected in about 32% of analyzed cells, regard-
less of adopted culture conditions (
p
g
0.05 vs. CTR, ANOVA). To interpret these
experimental results, an analysis of the computational model was performed,
highlighting how actin polymerization is the node of the network that contacts
the largest number of intracytoplasmic districts, with the specific biological role of
integrating events taking place in different cellular compartments, such as cytosol,
membrane, cytoskeleton, mitochondria, and acrosome (Fig.
6.1
).
>
6.4 Discussion
The biological results obtained in this study showed that sperm incubated with CD
under capacitating conditions negatively affected the acquisition of fertilizing
power. In fact, the coincubation of sperm with solubilized zonae pellucidae,
physiological agonists of AR, was unable to promote exocytosis of acrosomal
contents. However, the study of the main signal transduction pathways involved
in this process showed that they were unaltered by the presence of CD but were
efficient. The functional interpretation of these data can be inferred by analyzing the
