The hepatokine fetuin-A disrupts functional maturation of pancreatic beta cells

Felicia Gerst,Norbert Stefan,Rita Nano,Ann-Kathrin Fritz,Hans-Ulrich Häring,Silvio Nadalin,Elisabeth Kemter,Eckhard Wolf,Andreas Dahl,Alfred Königsrainer,Estela Lorza-Gil,Falko Fend,Gabriele Kaiser,Andreas L Birkenfeld,Martin Heni,Susanne Ullrich,Robert Wagner,Marie Gauder,Lorenzo Piemonti

doi:10.1007/s00125-021-05435-1

Abstract

Aims/hypothesisNeonatal beta cells carry out a programme of postnatal functional maturation to achieve full glucose responsiveness. A partial loss of the mature phenotype of adult beta cells may contribute to a reduction of functional beta cell mass and accelerate the onset of type 2 diabetes. We previously found that fetuin-A, a hepatokine increasingly secreted by the fatty liver and a determinant of type 2 diabetes, inhibits glucose-stimulated insulin secretion (GSIS) of human islets. Since fetuin-A is a ubiquitous fetal glycoprotein that declines peripartum, we examined here whether fetuin-A interferes with the functional maturity of beta cells.MethodsThe effects of fetuin-A were assessed during in vitro maturation of porcine neonatal islet cell clusters (NICCs) and in adult human islets. Expression alterations were examined via microarray, RNA sequencing and reverse transcription quantitative real-time PCR (qRT-PCR), proteins were analysed by western blotting and immunostaining, and insulin secretion was quantified in static incubations.ResultsNICC maturation was accompanied by the gain of glucose-responsive insulin secretion (twofold stimulation), backed up by mRNA upregulation of genes governing beta cell identity and function, such as NEUROD1, UCN3, ABCC8 and CASR (Log2 fold change [Log2FC] > 1.6). An active TGFβ receptor (TGFBR)–SMAD2/3 pathway facilitates NICC maturation, since the TGFBR inhibitor SB431542 counteracted the upregulation of aforementioned genes and de-repressed ALDOB, a gene disallowed in mature beta cells. In fetuin-A-treated NICCs, upregulation of beta cell markers and the onset of glucose responsiveness were suppressed. Concomitantly, SMAD2/3 phosphorylation was inhibited. Transcriptome analysis confirmed inhibitory effects of fetuin-A and SB431542 on TGFβ-1- and SMAD2/3-regulated transcription. However, contrary to SB431542 and regardless of cMYC upregulation, fetuin-A inhibited beta cell proliferation (0.27 ± 0.08% vs 1.0 ± 0.1% Ki67-positive cells in control NICCs). This effect was sustained by reduced expression (Log2FC ≤ −2.4) of FOXM1, CENPA, CDK1 or TOP2A. In agreement, the number of insulin-positive cells was lower in fetuin-A-treated NICCs than in control NICCs (14.4 ± 1.2% and 22.3 ± 1.1%, respectively). In adult human islets fetuin-A abolished glucose responsiveness, i.e. 1.7- and 1.1-fold change over 2.8 mmol/l glucose in control- and fetuin-A-cultured islets, respectively. In addition, fetuin-A reduced SMAD2/3 phosphorylation and suppressed expression of proliferative genes. Of note, in non-diabetic humans, plasma fetuin-A was negatively correlated (p = 0.013) with islet beta cell area.Conclusions/interpretationOur results suggest that the perinatal decline of fetuin-A relieves TGFBR signalling in islets, a process that facilitates functional maturation of neonatal beta cells. Functional maturity remains revocable in later life, and the occurrence of a metabolically unhealthy milieu, such as liver steatosis and elevated plasma fetuin-A, can impair both function and adaptive proliferation of beta cells.Data availabilityThe RNAseq datasets and computer code produced in this study are available in the Gene Expression Omnibus (GEO): GSE144950; https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE144950Graphical abstract

Highlights

The functional characteristic of mature beta cells is glucosestimulated insulin secretion (GSIS), defined as cell’s ability to secrete appropriate amounts of insulin in response to a stimulatory concentration of glucose [1, 2]
To assess whether fetuin-A impacts on functional maturation of beta cells, neonatal islet cell clusters (NICCs) were maturated in human serum albumin (HSA)- or fetuin-Acontaining medium
An active TGFBR–SMAD2/3 pathway is essential for beta cell maturation, as endorsed by the observation that SB431542 increased, while TGFβ-1 decreased, the mRNA and protein levels of aldolase B (ALDOB), a marker of functionally immature as well as of human diabetic beta cells (Fig. 2j–n) [27, 28]. These findings indicate that selective expression of beta cell markers during NICC maturation requires an active TGFBR–SMAD2/3 pathway

Summary

Introduction

The functional characteristic of mature beta cells is glucosestimulated insulin secretion (GSIS), defined as cell’s ability to secrete appropriate amounts of insulin in response to a stimulatory concentration of glucose [1, 2]. To achieve such a functional phenotype, neonatal beta cells undergo a tightly regulated process of maturation. GSIS declines with ageing or due to metabolic stress, resulting in insulin deficiency, which is the ultimate cause of diabetes onset. Growing evidence suggests that metabolic stress and insulin resistance curtail both endocrine identity and adaptive proliferation of beta cells [9]

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Conclusion