[Effects of Nd(2)O(3) exposure of rare earth particles on C57 BL/6J male mice sex hormone secretion and CYP11A1/PLZF/STRA8 protein expression].

Objective: To explore the effects of Nd(2)O(3) exposure to rare earth particles on the secretion of sex hormones, cytochrome P450 family member 11A1 (CYP11A1) , spermatogenesis markers promyelocytic leukemia zinc finger protein (PLZF) and retinoic acid stimulating gene 8 (STRA8) protein in C57 BL/6J male mice. Methods: In March 2021, Forty-eight male C57 BL/6J mice aged 6-8 weeks divided into control group and Nd(2)O(3) exposure low, medium and high dose groups (exposing doses of 62.5, 125.0, 250.0 mg/ml Nd(2)O(3)) , 12 per group. The mice in the Nd(2)O(3) groups were perfused with different doses of Nd(2)O(3) suspension by a one-time non-exposing tracheal instillation method, and the control group was perfused with an equal volume of normal saline, with a volume of 0.1 ml, to establish a mouse reproductive function injury model. After 28 days of exposure, the mice's body weight, testes and epididymis were weighed, and the organ coefficients were calculated; the two epididymis were taken to make a sperm suspension to determine the sperm count, survival rate, and deformity rate; inductively coupled plasma mass spectrometry (ICP-MS) method was used to detect the content of Nd in mouse testis tissue; HE staining was used to detect testicular tissue pathological changes and quantitative analysis; enzyme-linked immunosorbent assay (ELISA) method was used to detect serum luteinizing hormone (LH) and follicle stimulating hormone (FSH) and testosterone (T) content; western blot was used to detect the protein levels of CYP11A1, PLZF and STRA8 in testicular tissues. Results: Compared with the control group, with the increase of the exposure dose, the Nd content in the testis of the mice showed an increasing trend, the sperm survival rate and LH showed a decreasing trend, and the sperm deformity rate showed an increasing trend (P<0.05) ; Pathological showed that the number of sperm in the seminiferous tubules of the testicular tissue in the Nd(2)O(3) medium and high dose groups was significantly reduced, and the germinal epithelial disintegration, intraepithelial vacuolization, and exfoliation of spermatogenic cells and supporting cells occurred; The height of germinal epithelium was significantly reduced, and the percentage of damaged seminiferous tubules showed an increasing trend (P<0.05) ; FSH and T levels in serum in the middle and high dose groups of Nd(2)O(3), and CYP11A1, PLZF and STRA8 proteins in testicular tissues showed a downward trend with increasing dose (P<0.05) . Conclusion: The rare earth particulate Nd(2)O(3) may interfere with the expression of CYP11A1, PLZF and STRA8 protein, thereby causing the disorder of sex hormone secretion in the body, the maintenance of spermatogonia and the obstruction of the process of meiosis, causing reproductive function damage.

A comparison of the effects of fetal bovine serum and newborn calf serum on cell growth and maintenance of cryopreserved mouse spermatogonial stem cells.

Serum is a common supplement that is widely used to protect various cells and tissues from cryopreservation because it provides the necessary active components for cell growth and maintenance. In this study, we compared the effects of newborn calf serum (NCS) and fetal bovine serum (FBS) on the cryopreservation of mouse spermatogonial stem cells (SSCs). The isolated SSCs were cryopreserved in two groups: freezing medium that contained 10% DMSO (dimethyl sulfoxide) and 10% FBS in DMEM (Dulbecco's Modified Eagle's Medium) (group 1) and freezing medium that contained 10% DMSO and 10% NCS in DMEM (group 2). Real-time PCR was performed for stemness gene expression. The SSCs' viability was performed by trypan blue. We observed that the SSCs had increased viability in the NCS-freeze/thaw group (87.82%) compared to the FBS-freeze/thaw group (79.83%), but this increase was not statistically significant (P < 0.105). Promyelocytic leukemia zinc finger (Plzf) and Lin28 gene expression levels in the NCS-frozen/thawed SSCs were not significantly different compared to the FBS-frozen/thawed SSCs; however, Nanog gene expression increased considerably, and Dazl gene expression decreased significantly. The results in this study demonstrated that the presence of NCS in a solution of cryopreserved SSCs increased their viability after freeze/thawing and might promote the proliferation of cultivated SSCs in vitro by increasing the relative expression of Nanog.

Methylation of H3K27 and H3K4 in key gene promoter regions of thymus in RA mice is involved in the abnormal development and differentiation of iNKT cells.

Epigenetic modifications have been shown to be important for immune cell differentiation by regulating gene transcription. However, the role and mechanism of histone methylation in the development and differentiation of iNKT cells in rheumatoid arthritis (RA) mice have yet to be deciphered. The DBA/1 mouse RA model was established by using a modified GPI mixed peptide. We demonstrated that total peripheral blood, thymus, and spleen iNKT cells in RA mice decreased significantly, while iNKT1 in the thymus and spleen was increased significantly. PLZF protein and PLZF mRNA levels were significantly decreased in thymus DP T cells, while T-bet protein and mRNA were significantly increased in thymus iNKT cells. We found a marked accumulation in H3K27me3 around the promoter regions of the signature gene Zbtb16 in RA mice thymus DP T cells, and an accumulation of H3K4me3 around the promoters of the Tbx21 gene in iNKT cells. The expression levels of UTX in the thymus of RA mice were significantly reduced. The changes in the above indicators were particularly significant in the progressive phase of inflammation (11 days after modeling) and the peak phase of inflammation (14 days after modeling) in RA mice. Developmental and differentiation defects of iNKT cells in RA mice were associated with abnormal methylation levels (H3K27me3 and H3K4me3) in the promoters of key genes Zbtb16 (encoding PLZF) and Tbx21 (encoding T-bet). Decreased UTX of thymus histone demethylase levels resulted in the accumulation of H3K27me3 modification.

Coexpression of YY1 Is Required to Elaborate the Effector Functions Controlled by PLZF in NKT Cells.

The promyelocytic leukemia zinc-finger transcription factor (PLZF) is essential for nearly all of the unique, innate-like functions and characteristics of NKT cells. It is not known, however, if the activity of PLZF is regulated by other factors. In this article, we show that the function of PLZF is completely dependent on the transcription factor Yin Yang 1 (YY1). Mouse NKT cells expressing wild-type levels of PLZF, but deficient for YY1, had developmental defects, lost their characteristic "preformed" mRNA for cytokines, and failed to produce cytokine protein upon primary activation. Immunoprecipitation experiments showed that YY1 and PLZF were coassociated. Taken together, these biochemical and genetic data show that the broadly expressed transcription factor, YY1, is required for the cell-specific "master regulator" functions of PLZF.

Transcription factor YY1 is essential for iNKT cell development.

Invariant natural killer T (iNKT) cells develop from CD4+CD8+ double-positive (DP) thymocytes and express an invariant Vα14-Jα18 T-cell receptor (TCR) α-chain. Generation of these cells requires the prolonged survival of DP thymocytes to allow for Vα14-Jα18 gene rearrangements and strong TCR signaling to induce the expression of the iNKT lineage-specific transcription factor PLZF. Here, we report that the transcription factor Yin Yang 1 (YY1) is essential for iNKT cell formation. Thymocytes lacking YY1 displayed a block in iNKT cell development at the earliest progenitor stage. YY1-deficient thymocytes underwent normal Vα14-Jα18 gene rearrangements, but exhibited impaired cell survival. Deletion of the apoptotic protein BIM failed to rescue the defect in iNKT cell generation. Chromatin immunoprecipitation and deep-sequencing experiments demonstrated that YY1 directly binds and activates the promoter of the Plzf gene. Thus, YY1 plays essential roles in iNKT cell development by coordinately regulating cell survival and PLZF expression.