emad reza

Sox2 (SRY box2) is an essential transcription factor that plays a vital role in spermatogenesis and regulates the genes in this process. Sox2 is important for pluripotency, self-renewal, and even spermatogonial stem cell differentiation. This gene is found in pluripotent and specialized cells, and it is involved in their biological activities.

Protein-protein interaction (PPI) network analysis was performed during spermatogenesis using NCBI, STRING, and Cytoscape databases. Then, after isolating spermatogonial stem cells from 6 C57BL/6 mice, mouse embryonic stem cells and ES-like cells were prepared. In the following, Sox2 expression was examined in differentiated and undifferentiated spermatogonia by immunohistochemistry (IMH), immunocytochemistry (ICC), and Fluidigm PCR (polymerase chain reaction). Finally, the results were compared using the Kruskal-Wallis and Dunn tests at the significance level of p<0.05.

The results of this experiment showed that contrary to expectations, Sox2 has cytoplasmic expression in undifferentiated cells and nuclear expression in differentiated cells in in vitro conditions. In addition, the expression of Sox2 increased during differentiation. Fluidigm PCR showed a significantly higher expression of Sox2 (p<0.05) in differentiated compared to undifferentiated spermatogonia. Sox2 has an interaction with other genes during spermatogenesis such as Oct4, Nanog, Klf4, Stra8, Smad1, Tcf3, and Osm.

Sox2, which is known as a pluripotency marker, has a vital role in spermatogenesis and could be a differential marker. Sox2 has strong connections with other genes such as Oct4, Nanog, Klf4, Tcf3, Osm, Stra8, Lim2, Smad1, Gdnf, and Kit.

Alkaline phosphatase, BMP, and GATA proteins are important factors in the process of spermatogenesis. This study aims to investigate the effect of alkaline phosphatase, GATA, and BMP expression on spermatogenic stem cells, embryonic cells, and embryonic stem-like cells (ES-like) in C57BL mice.

In this experimental study, spermatogonial stem cells were isolated from three heads of 4-week-old C57BL mice, and embryonic stem cells and ES-like cells were prepared. Alkaline phosphatase staining test was performed on spermatogenic stem cells, embryonic cells, and ES-like cells. The expression of BMP and GATA genes was analyzed using Fluidigm PCR. Protein-protein interaction networks were isolated and drawn using databases.

Positive alkaline phosphatase expression in stem cells and negative expression in testicular Sertoli cells indicated the presence of this enzyme in pluripotent cells. The gene expression of BMP and GATA in spermatogonial stem cells (6.3 and 2.7, respectively), embryonic cells (3.2 and 4.4, respectively), and ES-like cells (8.5 and 2.5, respectively) was positive, but not statistically significant. Bioinformatics studies showed the regulatory role of these genes and their direct effect on alkaline phosphatase.

BMP and GATA genes, along with alkaline phosphatase enzymes, play a crucial role in controlling embryonic and spermatogonial stem cells, maintaining their pluripotency, and guiding them towards differentiated cells.
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One of the vital enzymes during spermatogenesis, which is one of the pluripotency factors of stem cells and contributes to maintaining their pluripotency is alkaline phosphatase. ZBTB16 and ZFP proteins are critical elements in stem cells which are expressed in pluripotent stem cells and maintain their pluripotency due to their role in messaging pathways.

The separation of the spermatogonial stem cells is the initial stage, followed by the preparation of the mouse embryonic stem cells and ES-like cells. The protein-protein network and the connections between genes were then investigated after alkaline phosphatase staining. Then, using the Fluidigm PCR method, the expression of the ZBTB16 and ZFP genes was determined. Finally, spermatogonia was immunohistochemically stained for the ZBTB16 protein.

Alkaline phosphatase was expressed positively by stem cell types; moreover, it was absent in Sertoli and fibroblast cells. Additionally, the specificity of these factors was demonstrated by the positive expression of the ZBTB16 and ZFP genes in these three cells and their negative expression in Sertoli cells. The connection and effect of these genes on the other genes are shown by the Protein-Protein network analysis.

Alkaline phosphatase, as well as ZBTB16 and ZFP genes are expected to be among the special markers of pluripotent stem cells and the critical elements of embryonic stem-like cells.

Spermatogenesis is a well-organized process that is influenced by a variety of factors. Alkaline phosphatase, and Gfra1, Lin28, and Sall4 genes are among the key players in this interconnected process. This study aimed to investigate the expression levels of Gfra1, Lin28, and Sall4 genes in embryonic, spermatogonial, and embryonic stem-like (ES-like) cells in mice.

First, spermatogonial stem cells were isolated, and then mouse embryonic stem cells and ES-like cells were prepared. Alkaline phosphatase expression was determined by alkaline phosphatase staining. After that, a bioinformatics analysis of directly related genes was done. Then the expression levels of the Gfra1, Lin28, and Sall4 genes were measured using Fluidigm PCR, and immunohistochemistry was done for Gfra1 expression.

In pluripotent stem cells and germ cells, the expression levels of Gfra1, Lin28, and Sall4 genes were found to be significantly positive (P<0.05). Alkaline phosphatase was also expressed positively in germ stem cells and pluripotent stem cells, but not in Sertoli cells or fibroblasts, indicating that this enzyme is exclusive to these cell types.

Because of their interaction and expression in pluripotent stem cells and germ cells, alkaline phosphatase, Gfra1, Lin28, and Sall4 genes can be employed as detecting markers for these cell lines.