Mesenchymal stem cells (MSCs) are the most promising seed cells for cell therapy. Comparing the biological and transcriptome gene characteristics of MSCs from different sources provides an important basis for the screening of clinically used cells. The main purpose of this experiment was to establish methods for the isolation and culture of MSCs from five different canine sources, including adipose tissue, bone marrow, umbilical cord, amniotic membrane, and placenta, and compare biological and transcriptome characteristics of MSCs, in order to provide a basis for the clinical application of canine MSCs. MSCs were isolated from Chinese pastoral dogs, and the following experiments were performed: (1) the third, sixth, and ninth generations of cells were counted, respectively, and a growth curve was plotted to calculate the MSC population doubling time; (2) the expression of CD34 and CD44 surface markers was studied by immunofluorescence; (3) the third generation of cells were used for osteogenetic and adipogenic differentiation experiments; and (4) MSC transcriptome profiles were performed using RNA sequencing. All of the five types of MSCs showed fibroblast-like adherent growth. The cell surface expressed CD44 instead of CD34; the third-generation MSCs had the highest proliferative activity. The average population doubling time of adipose mesenchymal stem cells (AD-MSCs), placenta mesenchymal stem cells (P-MSCs), bone marrow mesenchymal stem cells (BM-MSCs), umbilical cord mesenchymal stem cells (UC-MSCs), and amniotic mesenchymal stem cells (AM-MSCs) were 15.8 h, 21.2 h, 26.2 h, 35 h, and 41.9 h, respectively. All five types of MSCs could be induced to differentiate into adipocytes and osteoblasts in vitro, with lipid droplets appearing after 8 days and bone formation occurring 5 days after AD-MSC induction. However, the multilineage differentiation for the remaining of MSCs was longer compared to that of the AD-MSCs. The MSC transcriptome profiles showed that AD-MSC and BM-MSCs had the highest homology, while P-MSCs were significantly different compared to the other four types of MSCs. All the isolated MSCs had the main biological characteristics of MSCs. AD-MSCs had the shortest time for proliferation, adipogenesis, and osteogenic differentiation.

Decidualization is necessary for the successful establishment of early pregnancy in rodents and humans. Disturbed decidualization results in recurrent implantation failure, recurrent spontaneous abortion, and preeclampsia. Tryptophan (Trp), one of the essential amino acids in humans, has a positive effect on mammalian pregnancy. Interleukin 4-induced gene 1 (IL4I1) is a recently identified enzyme that can metabolize L-Trp to activate aryl hydrocarbon receptor (AHR). Although IDO1-catalyzed kynurenine (Kyn) from Trp has been shown to enhance human in vitro decidualization via activating AHR, whether IL4I1-catalyzed metabolites of Trp are involved in human decidualization is still unknown. In our study, human chorionic gonadotropin stimulates IL4I1 expression and secretion from human endometrial epithelial cells through ornithine decarboxylase-induced putrescine production. Either IL4I1-catalyzed indole-3-pyruvic acid (I3P) or its metabolite indole-3-aldehyde (I3A) from Trp is able to induce human in vitro decidualization by activating AHR. As a target gene of AHR, Epiregulin induced by I3P and I3A promotes human in vitro decidualization. Our study indicates that IL4I1-catalyzed metabolites from Trp can enhance human in vitro decidualization through AHR-Epiregulin pathway.

Acrylamide is a class 2A carcinogen with neurotoxicity and genotoxicity. In addition to industrial production, it is ubiquitous in high-temperature heated high-carbohydrate foods. Numerous studies have confirmed the toxicity of ACR on reproduction. Implantation and decidualization are crucial processes during the establishment of pregnancy in rodents and humans. However, its effect on uterine implantation and decidualization remains poorly understood. The objective of this study is to elucidate the mechanism by which ACR affects implantation and decidualization in mice. ACR is exposed in the daily drinking water of female mice, and the dose is calculated according to the body weight of the mice. After 3 months of administration at concentrations of 0, 20, and 30 mg ACR/kg/d, female mice are mated with male mice to induce pregnancy. Compared to the control group, ACR treatment significantly reduces the number of embryo implantations and litter size. ACR treatment leads to abnormal expression of endometrial receptivity-related molecules in the luminal epithelium on day 4 of pregnancy, including a decrease in p-STAT3 level and an increase in MUC1 and MSX1 levels. The level of decidualization-related molecules is obviously downregulated by ACR. Furthermore, ACR treatment results in abnormality of oxidative stress- and ferroptosis-related protein levels at the implantation site on day 5. In conclusion, acrylamide can impair mouse implantation and decidualization by disrupting oxidative stress and ferroptosis.

The supply of amino acids is essential to embryo survival and successful pregnancy. The accumulating evidence indicates that arginine, a semi-essential amino acid, plays a key role during early pregnancy. Arginase (ARG2) is a key enzyme for catalyzing arginine into ornithine and urea. However, the expression, regulation, and role of arginase during early mouse pregnancy are still unknown. In our study, ARG1 immunofluorescence is mainly detected in uterine epithelium and gradually decreases from days 1 to 5 of pregnancy. From days 1 to 4 of pregnancy, there is no detectable ARG2 immunofluorescence in the mouse uterus. On day 5 of pregnancy, ARG2 signals are strongly seen in the primary decidua surrounding the implanting blastocyst at the implantation site, but not at the inter-implantation site. There is a temporary increase for ARG2 levels under mouse in vitro decidualization, suggesting ARG2 may be involved in the initiation of mouse decidualization. Prl8a2, a marker of mouse in vitro decidualization, is significantly decreased after ARG levels are suppressed. However, Arg2 overexpression obviously increases Prl8a2 levels. Mouse in vitro decidualization is downregulated by arginine and ornithine but stimulated by a low dose of urea. Urea has a beneficial effect on uterine receptivity and antioxidative enzymes. Our results indicate that ARG2 plays an important role during mouse decidualization by balancing the levels of arginine, ornithine, and urea.