Integrating LCM-Based Spatio-Temporal Transcriptomics Uncovers Conceptus and Endometrial Luminal Epithelium Communication that Coordinates the Conceptus Attachment in Pigs

Wang, Feiyu; Zhao, Shilei; Deng, Dadong; Wang, Weiwei; Xu, Xuewen; Zhao, Shuhong; Yu, Mei

doi:10.3390/ijms22031248

Cited by 14 publications

(17 citation statements)

References 81 publications

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“…Results showed that Chi-miR-451-5p was mainly expressed in the stromal cells of the endometrium, suggesting that stromal cells may play a role in the secretion of chi-miR-451-5p. Additionally, higher chi-miR-451-5p levels were detected in the endometrial luminal epithelium on P16, as communication between the endometrial luminal epithelium and embryo is important for embryo implantation [ 76 ]. Moreover, the dual-luciferase reporter assay proved that chi-miR-451-5p directly targets 3′-UTR of PSMB8 .…”

Section: Discussionmentioning

confidence: 99%

Differential Expression Pattern of Goat Uterine Fluids Extracellular Vesicles miRNAs during Peri-Implantation

Xie

Liu

Zang

et al. 2021

Cells

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Early pregnancy failure occurs when a mature embryo attaches to an unreceptive endometrium. During the formation of a receptive endometrium, extracellular vesicles (EVs) of the uterine fluids (UFs) deliver regulatory molecules such as small RNAs to mediate intrauterine communication between the embryo and the endometrium. However, profiling of small RNAs in goat UFs’ EVs during pregnancy recognition (day 16) has not been carried out. In this study, EVs were isolated from UFs on day 16 of the estrous cycle or gestation. They were isolated by Optiprep™ Density G radient (ODG) and verified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blotting. Immunostaining demonstrated that CD63 was present both in the endometrial epithelium and glandular epithelium, and stain intensity was greater in the pregnant endometrium compared to the non-pregnant endometrium. Small RNA sequencing revealed that UFs’ EVs contained numerous sRNA families and a total of 106 differentially expressed miRNAs (DEMs). Additionally, 1867 target genes of the DEMs were obtained, and miRNA–mRNA interaction networks were constructed. GO and KEGG analysis showed that miRNAs were significantly associated with the formation of a receptive endometrium and embryo implantation. In addition, the fluorescence in situ hybridization assay (FISH) showed that chi-miR-451-5p was mainly expressed in stromal cells of the endometrium and a higher level was detected in the endometrial luminal epithelium in pregnant states. Moreover, the dual-luciferase reporter assay showed that chi-miR-451-5p directly binds to PSMB8 and may play an important role in the formation of a receptive endometrium and embryo implantation. In conclusion, these results reveal that UFs’ EVs contain various small RNAs that may be vital in the formation of a receptive endometrium and embryo implantation.

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Section: Discussionmentioning

confidence: 99%

Differential Expression Pattern of Goat Uterine Fluids Extracellular Vesicles miRNAs during Peri-Implantation

Xie

Liu

Zang

et al. 2021

Cells

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“…Sample collection was carried out according to Wang et al 6 Briefly, Yorkshire gilts were mated to Yorkshire boars at the onset of the second estrus (Day 0) and again 12 h later. On gestational days 12, 15, and 18, the uterus was taken from each gilt and cut into segments (3–4 gilts/gestational day).…”

Section: Methodsmentioning

confidence: 99%

“…Immunofluorescence assays were carried out as described by Wang et al 6 The primary antibodies used were ST6GAL1 (1:50, ab77676, abcam), ST6GAL2 (1:50, AF7747; R&D Systems), E‐cadherin (1:50, ab40772; abcam), β‐catenin (1:50, ab6302; abcam), Vimentin (1:50; sc‐73258), Rac1 (1:30, 66122–1‐Ig; Proteintech) and RhoA (1:50, 10749–1‐Ig; Proteintech). The fluorescent‐dye conjugated secondary antibodies used were Alexa Fluor ® 488 (1:100, ab150077/ab150113; abcam) or Alexa Fluor ® 555 (1:100, A21422/A21428; invitrogen).…”

Section: Methodsmentioning

confidence: 99%

“…The conceptuses freely float in the uterine lumen until gestational days 12–13 and then start to adhere to the uterine (LE) to form an epitheliochorial placenta around gestational day 18. 4 , 5 Previous studies have found that (1) when conceptuses remain free‐floating (before gestational day 12), the uterine lumen is in an open state and a few scattered endometrial folds occur; (2) at the stage of conceptus attachment (around gestational day 15), the endometrial folds that extend greatly to the uterine lumen interlock with each other, thus resulting in closure of the uterine lumen 5 , 6 ; (3) around gestational day 18, the uterine lumen reopens due to the expansion of chorioallantois. 4 Therefore, the endometrial fold extension is, at least partially, the cause of the uterine lumen closure in pigs.…”

Section: Introductionmentioning

confidence: 99%

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Glycomics reveal that ST6GAL1‐mediated sialylation regulates uterine lumen closure during implantation

et al. 2021

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Objectives Implantation failure is a major cause of prenatal mortality. The uterine lumen closure contributes to embryo adhesion to the uterus, but its underlying mechanisms are largely unknown. Our previous study has reported that endometrial fold extension can lead to uterine lumen closure in pigs. The objective of this study was to reveal molecular mechanisms of the uterine lumen closure by characterizing the molecular basis of the endometrial fold extension during implantation in pigs. Materials and methods Uterine and endometrium tissues during implantation were collected in pigs. MALDI‐TOF MS was used to characterize the N‐glycomic profiles. Histochemistry, siRNA transfection, Western blotting, lectin immumoprecipitation, mass spectrometry and assays of wounding healing and cell aggregation were performed to investigate the molecular basis. Results We observed that uterine luminal epithelium (LE) migrated collectively during endometrial fold extension. For the first time, we identified a large number of N‐glycan compositions from endometrium during implantation using MALDI‐TOF MS. Notably, the α2,6‐linked sialic acid and ST6GAL1 were highly expressed in uterine LE when the endometrial folds extended greatly. Subsequently, the role of ST6GAL1‐mediated 2,6‐sialylation in collective epithelial migration was demonstrated. Finally, we found that ST6GAL1‐mediated α2,6‐sialylation of E‐cadherin may participate in collective migration of uterine LE. Conclusions The study reveals a mechanism of uterine lumen closure by identifying that ST6GAL1‐mediated α2,6‐sialylation of cell adhesion molecules contributes to endometrial fold extension through regulating collective migration of uterine LE.

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“…Thus, these studies were unable to accurately capture cell-type-specific gene expression changes. Recently, laser capture microdissection (LCM) has been applied to obtain the global gene expression profiles in luminal epithelial cells, glandular epithelial cells and stromal cells of peri-implantation porcine endometrium ( Zeng et al, 2018 ; Zeng et al, 2019 ; Wang et al, 2021 ). The limitation of LCM-based methods is that cell types such as various immune cells can be hardly isolated.…”

Section: Introductionmentioning

confidence: 99%

Revisiting the Transcriptome Landscape of Pig Embryo Implantation Site at Single-Cell Resolution

Tian

Zhu

et al. 2022

Front. Cell Dev. Biol.

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Litter size is one of the most economically important traits in commercial pig farming. It has been estimated that approximately 30% of porcine embryos are lost during the peri-implantation period. Despite rapid advances over recent years, the molecular mechanism underlying embryo implantation in pigs remains poorly understood. In this study, the conceptus together with a small amount of its surrounding endometrial tissues at the implantation site was collected and subjected to single-cell RNA-seq using the 10x platform. Because embryo and maternal endometrium were genetically different, we successfully dissected embryonic cells from maternal endometrial cells in the data according to single nucleotide polymorphism information captured by single-cell RNA-seq. Undoubtedly, the interaction between trophoblast cells and uterine epithelial cells represents the key mechanism of embryo implantation. Using the CellChat tool, we revealed cell-cell communications between these 2 cell types in terms of secreted signaling, ECM-receptor interaction and cell-cell contact. Additionally, by analyzing the non-pregnant endometrium as control, we were able to identify global gene expression changes associated with embryo implantation in each cell type. Our data provide a valuable resource for deciphering the molecular mechanism of embryo implantation in pigs.

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Integrating LCM-Based Spatio-Temporal Transcriptomics Uncovers Conceptus and Endometrial Luminal Epithelium Communication that Coordinates the Conceptus Attachment in Pigs

Cited by 14 publications

References 81 publications

Differential Expression Pattern of Goat Uterine Fluids Extracellular Vesicles miRNAs during Peri-Implantation

Differential Expression Pattern of Goat Uterine Fluids Extracellular Vesicles miRNAs during Peri-Implantation

Glycomics reveal that ST6GAL1‐mediated sialylation regulates uterine lumen closure during implantation

Revisiting the Transcriptome Landscape of Pig Embryo Implantation Site at Single-Cell Resolution

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