Kim M, et al. Immune Netw. 2022 Feb;22(1):e8. https://doi.org/10.4110/in.2022.22.e8

Lymph node fibroblastic reticular cells (LN-FRCs) provide functional structure to LNs and play important roles in interactions between T cells and antigen-presenting cells. However, the direct impact of LN-FRCs on naive CD4+ T cell differentiation has not been explored. Here, we show that T cell zone FRCs of LNs (LN-TRCs) express CD25, the α chain of the IL-2 receptor heterotrimer. Moreover, LN-TRCs trans-present IL-2 to naive CD4+ T cells through CD25, thereby facilitating early IL-2–mediated signaling. CD25-deficient LN-TRCs exhibit attenuated STAT5 phosphorylation in naive CD4+ T cells during T cell differentiation, promoting T helper 17 (Th17) cell differentiation and Th17 response-related gene expression. In experimental autoimmune disease models, disease severity was elevated in mice lacking CD25 in LN-TRCs. Therefore, our results suggest that CD25 expression on LN-TRCs regulates CD4+ T cell differentiation by modulating early IL-2 signaling of neighboring, naive CD4+ T cells, influencing the overall properties of immune responses.

Objective: Pregnancy in cattle after embryo transfer (ET) is influenced by several factors, including embryo quality. Therefore, preparing high-quality embryos with the greatest developmental potential is essential for achieving a successful pregnancy after ET. Meanwhile, blastocysts produced by in vitro fertilization (IVF) procedure have different developmental speed during in vitro culture (IVC) and they exhibited different competence in the establishment of pregnancy.Methods: This study aimed to identify the comparative features of early-, mid-, and late-developing bovine IVF blastocysts, when they first appeared at Day 7, 8, and 9 during IVC, respectively. In addition, the correlations between their molecular features and pregnancy ability were analyzed.Results: The results showed no difference in the morphological characteristics, including total cell count and diameter, between the Day 7, 8, and 9 blastocysts. However, the pregnancy rate post-ET was significantly different between the groups at 51.7%, 36.7%, and 17.8% for Day 7, 8, and 9 blastocysts, respectively. During early embryo development, late-developing blastocysts demonstrated a reduced cell count in the inner cell mass and decreased expression of the early embryo developmental genes (Oct4 and Sox2) compared with the early- and mid-developing blastocysts. In addition, the number of apoptotic cells and apoptosis-related gene expression (increased Bax and decreased Bcl2) gradually elevated from the Day 7 to Day 9 blastocysts. However, there was no difference in mitochondrial activity and mitochondria-relevant gene expression (Tfam and Cox1) between the groups. Correlation analysis identified a significantly negative correlation between the pregnancy rate and the blastocysts’ degree of apoptosis, indicating that the low pregnancy ability of late-developing blastocysts was mainly caused by increased apoptosis.Conclusion: This study’s results may contribute to the field of animal biotechnology by assisting in establishing an improved strategy for bovine ET with IVF embryos.

Reference genes are crucial in molecular biological studies as an internal control for gene re-search as they exhibit consistent expression patterns across many tissue types. In canines, radiation therapy is the most important therapeutic tool to cure various diseases like cancer. However, when using radiation for therapeutic strategy, radiation exposure to healthy tissues leads to some possible side effects such as acute radiation-induced skin injury and alters gene expression. Therefore, the analysis of a change in reference gene expression during the skin recovery process after radiation therapy is essential in healthy canine tissue. In the present study, we analyzed eight reference genes (ACTB, GAPDH, YWHAZ, GUSB, HPRT1, RPL4, RPS5, and TBP) in canine dermal tissues at 0, 1, 2, 3, 4, 5, 7, and 9 weeks of radiation exposure that affected the skin condition of canines. The stability of reference genes is determined by evaluating radiation therapy’s effect on healthy canine dermal tissue. Epidermal marker, Keratin 10 expression varies each week after irradiation, and HPRT1 is found to be the most suitable for normalization of mRNA expression in radiation-exposed canine dermal tissues. Changes in the gene expression level were evaluated by using a reliable tool such as quantitative real-time polymerase chain reaction (qRT-PCR). In order to achieve a valid qRT-PCR result, the most stable reference genes used for normalization after the radiation exposure process are important. Therefore, the current study was designed to evaluate the most stable reference gene for the post-irradiation canine tissues. After radiation exposure, the alternation of reference gene expression was estimated by three algorithms (geNorm, Normfinder, and Bestkeeper). The RG validation programs (GeNorm and NormFinder) suggested that HPRT1, RPL4, and TBP were suitable for normalization in qRT-PCR. Furthermore, three algorithms suggested that HPRT1 was the most stable reference gene for normalization with qRT-PCR results, regardless of before and after radiation exposure. Whereas GAPDH was found to be the most unstable reference gene. In addition, the use of stable or unstable reference genes for the normalization of Keratin 10 expression showed statistical differences. Therefore, we observed that, to obtain accurate and suitable PCR results of the canine tissues with and without radiation exposure, the HPRT1 reference gene is recommended for normalization with its high stability. Additionally, the use of RGs such as HPRT1, RPL4, and TBP for normalization in qRT-PCR experiments is recommended for post-radiation canine tissues to generate more accurate and reliable data. These results will provide fundamental information regarding internal controls for gene expression studies and can be used for the analysis of gene patterns in regenerative medicine.

Abnormal epigenetic reprogramming of nuclear-transferred (NT) embryos leads to the limited efficiency of producing cloned animals. Trichostatin A (TSA), a histone deacetylase inhibitor, improves NT embryo development, but its role in histone acetylation in porcine embryos cloned with mesenchymal stem cells (MSCs) is not fully understood. This study aimed to compare the effects of TSA on embryo development, histone acetylation patterns, and key epigenetic-related genes between in vitro fertilization (IVF), NT-MSC, and 40 nM TSA-treated NT-MSC (T-NT-MSC). The results demonstrated an increase in the blastocyst rate from 13.7% to 32.5% in the T-NT-MSC, and the transcription levels of CDX2, NANOG, and IGF2R were significantly elevated in T-NT-MSC compared to NT-MSC. TSA treatment also led to increased fluorescence intensity of acH3K9 and acH3K18 during early embryo development but did not differ in acH4K12 levels. The expression of epigenetic-related genes (HDAC1, HDAC2, CBP, p300, DNMT3a, and DNMT1) in early pre-implantation embryos followed a pattern similar to IVF embryos. In conclusion, TSA treatment improves the in vitro development of porcine embryos cloned with MSCs by increasing histone acetylation, modifying chromatin structure, and enhancing the expression of key genes, resulting in profiles similar to those of IVF embryos.

Kim BG, et al. Immune Netw. 2024 Aug;24(4):e32. https://doi.org/10.4110/in.2024.24.e32