{'fraction_non_alphanumeric': 0.02586206896551724, 'fraction_numerical': 0.011853448275862068, 'mean_word_length': 5.337883959044369, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Technical note: Improving the efficiency of generating bovine extraembryonic endoderm cells.

The formation of extra embryonic endoderm (XEN) occurs early in embryonic development. The cell types that develop from the XEN remain poorly studied in ruminant species because of the lack of suitable cell culture model systems. The goal of this work was to establish a protocol for producing XEN cell cultures from bovine blastocysts. Previous work identified fibroblast growth factor 2 (FGF2) as a facilitator of bovine XEN development. Further refinements in culture conditions studied here included exposure to 20% fetal bovine serum and FGF2 replenishment. These modifications yielded an endoderm outgrowth formation incidence of 81.6 ± 5.5% compared to 33.3% ± 5.5% in bovine serum albumen-supplemented (BSA) controls. These cells resembled XEN when examined morphologically and contained XEN transcripts (GATA4, GATA6) as well as transcripts present in visceral (BNIP1, VEGFA) and parietal (CXCR4, THBD, HHEX) XEN. Two XEN cell lines were maintained for prolonged culture. Both lines continued to proliferate for approximately 6 wk before becoming senescent. These cultures maintained an XEN-like state and continued to express GATA4 and GATA6 until senescence. An increase in abundance of visceral and parietal XEN transcripts was observed with continued culture, suggesting these cells either undergo spontaneous differentiation or retain the ability to form various XEN cell types. Stocks of cultured cells exposed to a freeze-thaw procedure possessed similar phenotypic and genotypic behaviors as non-frozen cells. To conclude, a procedure for efficient production of primary bovine XEN cell cultures was developed. This new protocol may assist researchers in exploring this overlooked cell type for its roles in nutrient supply during embryogenesis.