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Gene expression profiles of Wnt4 high and Wnt4 low populations of murine islets at postnatal day 1 (P1).

ABSTRACT: To find the differences between β cells expressing Wnt4 and those that do not express it (or express it at low levels) at neonatal stage, murine islets at postnatal day 1 (P1) were collected. Based on a reporter expressing Cre recombinase from the Wnt4 locus, we observed that Wnt4 expression starts in a small subset of β cells at embryonic day (E) 18.5. At neonatal stages, around 50% of β cells express Wnt4 at P0 and most of cells express it by P5. We collected Wnt4 high and Wnt4 low islet cells by FACS and compared their gene expression profile. Wnt4 high islet cell population showed a significant increase of β-cell maturation and function related genes, including transcription factors such as Pdx1 and Nkx6.1, insulin 2, glucokinase (Gck), and the glucose transporter Slc2a2. On the contrary, Wnt4 low islet cell population showed significant increasing of proliferation markers such as Pcna, Mki67 and the positive cell cycle regulators Ccnd1, Ccnd3, Ccna2, Cdk1, Cdk2 and Cdk6. Taken together, these results show that Wnt4 expression is higher in more mature but less proliferative β cells.

ORGANISM(S): Mus musculus

PROVIDER: GSE210237 | GEO | 2022/08/02

REPOSITORIES: GEO

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Project description:To study the physiological role of WNT4 in the postnatal ovary, a mouse strain bearing a floxed Wnt4 allele was created and mated to the Amhr2tm3(cre)Bhr strain to target deletion of Wnt4 to granulosa cells. Wnt4flox/-;Amhr2tm3(cre)Bhr/+ mice had significantly reduced ovary weights and produced smaller litters (P<0.05). Serial follicle counting demonstrated that, while Wnt4flox/-;Amhr2tm3(cre)Bhr/+ mice were born with a normal ovarian reserve and maintained normal numbers of small follicles until puberty, they had only 25.2% of the normal number of healthy antral follicles. Some Wnt4flox/-;Amhr2tm3(cre)Bhr/+ mice had no antral follicles or corpora lutea and underwent premature follicle depletion. RTPCR analyses of Wnt4flox/-;Amhr2tm3(cre)Bhr/+ granulosa cells and cultured granulosa cells that overexpress WNT4 demonstrated that WNT4 regulates the expression of Star, Cyp11a1 and Cyp19, steroidogenic genes previously identified as downstream targets of the WNT signaling effector CTNNB1. WNT4- and CTNNB1-overexpressing cultured granulosa cells were analyzed by microarray for alterations in gene expression, which showed that WNT4 also regulates a series of genes involved in late follicle development and the cellular stress response via the WNT/CTNNB1 signaling pathway. Together, these data indicate that WNT4 is required for normal antral follicle development, and may act by regulating granulosa cell functions including steroidogenesis. Experiment Overall Design: Granulosa cells were obtained from 20-26 day-old mice of various genotypes 48h after eCG treatment, using the needle puncture method as previously described. Cells were then infected with adenoviruses to express eGFP or Cre, or overexpress WNT4 in serum-free medium, and subsequently harvested for RNA extraction as described below. Preliminary experiments demonstrated that an infection efficiency of >80% could be obtained at an MOI of ~50 (as determined by analysis of fluorescent signal in Ad-eGFP-infected cells) and that the Ad-Cre and Ad-WNT4 viruses produced efficient recombination of the floxed Ctnnb1 alleles and robust WNT4 overexpression, respectively. Microarray analyses were done using triplicate RNA samples from each adenoviral treatment described above, and using mouse expression set 430 microarrays (Affymetrix, Santa Clara, CA).

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Gene expression profiles of Wnt4 high and Wnt4 low populations of murine islets at postnatal day 1 (P1).

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