Diabetes. 2015 Apr;64(4):1284-98.

Connective tissue growth factor modulates adult β-cell maturity and proliferation to promote β-cell regeneration in mice.

 

Riley KG1, Pasek RC2, Maulis MF2, Peek J3, Thorel F4, Brigstock DR5, Herrera PL4, Gannon M6.
  • 1Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN.
  • 2Department of Medicine, Vanderbilt University, Nashville, TN.
  • 3The School for Science and Math at Vanderbilt, Vanderbilt University, Nashville, TN.
  • 4Department of Genetic Medicine and Development, University of Geneva, Geneva, Switzerland.
  • 5Center for Cell and Vascular Biology, Children’s Research Institute, The Ohio State University, Columbus, OH.
  • 6Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN Department of Medicine, Vanderbilt University, Nashville, TN Department of Veterans Affairs, Tennessee Valley Health Authority, Nashville, TN Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN maureen.gannon@vanderbilt.edu.

 

Abstract

Stimulation of endogenous β-cell expansion could facilitate regeneration in patients with diabetes. In mice, connective tissue growth factor (CTGF) is expressed in embryonic β-cells and in adult β-cells during periods of expansion. We discovered that in embryos, CTGF is necessary for β-cell proliferation, and increased CTGF in β-cells promotes proliferation of immature (MafA-) insulin+ cells. CTGF over-expression, under non-stimulatory conditions, does not increase adult β-cell proliferation. Here, we tested the ability of CTGF to promote β-cell proliferation and regeneration after partial β-cell destruction. β-cell mass reaches 50% recovery after 4 weeks of CTGF treatment, primarily via increased β-cell proliferation, which is enhanced as early as 2 days of treatment. CTGF treatment increases the number of immature β-cells, but promotes proliferation of both mature and immature β-cells. A shortened β-cell replication refractory period is also observed. CTGF treatment up-regulates positive cell cycle regulators and factors involved in β-cell proliferation, including HGF, serotonin synthesis, and integrin β1. Ex vivo treatment of whole islets with recombinant human CTGF induces β-cell replication and gene expression changes consistent with those observed in vivo, demonstrating that CTGF acts directly on islets to promote β-cell replication. Thus, CTGF can induce replication of adult mouse β-cells given a permissive micro-environment.

PMID: 25392241

 

Supplement

The major focus of my laboratory is to identify and understand the molecular and cell biological mechanisms regulating the generation, survival, expansion, and regeneration of insulin-producing β cells. Both Type 1 diabetes (T1D) and Type 2 diabetes (T2D) are characterized by reduced functional β cell mass. Thus, strategies to enhance and maintain functional β cell mass in vivo would provide therapy for both forms of diabetes. ~9% percent of the total US population has T2D, the prevalence of which increases with age, in part due to a decreased ability of β cells to respond to proliferative cues as they get older. Our long-term goal is to identify novel therapeutic targets to preserve β cell mass and promote adult β cell proliferation to enhance functional β cell mass in people with diabetes.

Our group focuses on β cell replication and has identified novel factors that promote adult β cell mass expansion. We determined that the transcription factor FoxM1 is a critical regulator of postnatal β cell replication (1), and discovered that this protein is required for increased β cell proliferation in response to various stimuli, including pregnancy (2). FoxM1 mutant females develop gestational diabetes due to failed β cell mass expansion in maternal islets. Foxm1 expression is induced in islets in response to high glucose, high fat diet (HFD), β cell injury, and growth factors such as placental lactogen and Connective Tissue Growth Factor (CTGF). Our studies have identified CTGF as a potent β cell mitogen. CTGF is expressed in embryonic β cells and is essential for embryonic β cell proliferation (3,4). In vivo, CTGF induction following partial β cell destruction induces Foxm1 expression, promotes β cell proliferation and restores β cell mass (5). Treatment of adult mouse islets ex vivo with recombinant CTGF induces β cell proliferation and also induces expression of Foxm1 as well as the CTGF receptors integrinα5 and integrinβ1 (5). Activation of FoxM1 in vivo in β cells from middle-aged mice can rejuvenate their replicative potential and induce increases in β cell mass (6). Thus, we have identified relevant pathways to target for induction of proliferation of adult human β cells as a potential means for driving functional β cell mass expansion to alleviate diabetes. However, the molecular mechanism whereby CTGF induces Foxm1 expression, adult β cell proliferation, and β cell mass regeneration is currently unknown.

 

MG Fig1

Fig 1. Many physiological and pathological stimuli induce β cell proliferation, including hormones, growth factors, metabolites, and β cell death. All of these stimuli have been shown to induce transcription of the Foxm1 The FoxM1 transcription factor regulates many genes involved in cell replication and is required for β cell proliferation and β cell mass expansion in response to each of these stimuli. In the absence of FoxM1, β cell mass does not increase and diabetes ensues.

 

Activation of endogenous β cell mass expansion could facilitate regeneration in individuals with diabetes. In our mouse model, we found that induction of CTGF in remaining β-cells after 50% β cell ablation resulted in an increase in recruitment of both macrophages and T-cells to islets (7). Up-regulation of several macrophage and T-cell chemoattractant genes was also observed in these islets. Gene expression analyses revealed an increase in markers associated with a more pro-inflammatory M1 macrophage polarization phenotype and a decrease in markers of M2-type macrophages. To test whether the recruited macrophages were required for the effects of CTGF on β cell proliferation, we depleted macrophages systemically using clodronate-loaded liposomes. Depletion of macrophages (without changes in T-cell number) completely blocked CTGF-mediated β cell proliferation. These data reveal that macrophages are critical for CTGF-mediated adult β cell proliferation in the setting of partial β cell loss and are likely secreting a factor(s) that either enhances β cell responsiveness to CTGF or directly induces β cell proliferation. This is the first study to link a specific β cell proliferative factor with immune-mediated β cell proliferation in a β cell injury model and suggests that the immune system plays a positive role in β cell regeneration under certain circumstances. Being able to harness immune-derived β cell proliferative factors may therefore enhance β cell mass regeneration in individuals with diabetes.

 

MG Fig2

Fig 2. Following partial β cell loss, secretion of CTGF from β cells induces Foxm1 gene transcription and macrophage (mf) recruitment. The combination of CTGF plus mf induces β cell proliferation and regenerates β cell mass. It is currently unknown through which signaling pathway CTGF mediates its effects on adult β cells. The mf-derived factor that promotes β cell proliferation is also unidentified at this time.

 

References

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  2. Zhang, H., J. Zhang, C.F. Pope, L. Crawford, R. Vasavada, S.M. Jagasia, and M. Gannon. (2010). Gestational diabetes mellitus resulting from impaired cell compensation in the absence of FoxM1, a novel downstream effector of placental lactogen. Diabetes 59, 143-152. PMID: 19833884
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  5. Riley, K.G., R.C. Pasek, M.F. Maulis, J. Peek, F. Thorel, D.R. Brigstock, P.L. Herrera, and M. Gannon. (2015). CTGF modulates adult  cell maturity and proliferation to promote  cell regeneration in mice. Diabetes. 64(4), 1284-1298. PMID: 25392241
  6. Golson, M.L., J.C. Dunn, M.F. Maulis, P.K. Dadi, Anna B. Osipovich, M.A. Magnuson, D. A, Jacobson, and M. Gannon. (2015). Activation of FoxM1 revitalizes the replicative potential of aged -cells in male mice and enhances insulin secretion. Diabetes. 64(11), 3829-3838. PMID: 26251404
  7. Riley, K.G., M.F. Maulis, P.L.Kendall, A.H. Hasty, and M. Gannon. (2015). CTGF promotes -cell regeneration via immune cell modulation. Molecular Metabolism. 4, 584-591. PMID: 26266091

 

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