Kidney Week

Abstract: FR-PO395

SMPDL3b Modulates Insulin Signaling in Lipid Raft Domains and Interferes with Diabetic Kidney Disease

Session Information

• Diabetic Kidney Disease: Basic - II
  October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Diabetic Kidney Disease

• 601 Diabetic Kidney Disease: Basic

Authors

• Mitrofanova, Alla, University of Miami School of Medicine, Miami, Florida, United States

Alla Mitrofanova,

• Mallela, Shamroop Kumar, University of Miami School of Medicine, Miami, Florida, United States

Shamroop Kumar Mallela,

• Ducasa, Gloria Michelle, University of Miami School of Medicine, Miami, Florida, United States

Gloria Michelle Ducasa,

• Yoo, Tae-Hyun, Yonsei University College of Medicine, Seoul, Korea (the Republic of)

Tae-Hyun Yoo,

• Molina David, Judith T., University of Miami, Miami, Florida, United States

Judith T. Molina David,

• Ge, Mengyuan, University of Miami, Miami, Florida, United States

Mengyuan Ge,

• Kim, Jin Ju, University of Miami, Miami, Florida, United States

Jin Ju Kim,

• Pedigo, Christopher E., Yale University, New Haven, Connecticut, United States

Christopher E. Pedigo,

• Faul, Christian, The University of Alabama at Birmingham, Birmingham, Alabama, United States

Christian Faul,

• Ishimoto, Yu, Tokyo University, Hongo, ToKyo, Japan

Yu Ishimoto,

• Inagi, Reiko, The University of Tokyo Graduate School of Medicine, Bunkyo-ku, TOKYO, Japan

Reiko Inagi,

• Merscher, Sandra M., University of Miami, Miami, Florida, United States

Sandra M. Merscher,

• Fornoni, Alessia, University of Miami, Miami, Florida, United States

Alessia Fornoni,

Background

SMPDL3b is a recently identified phosphodiesterase localized in lipid raft domains that regulates lipid composition and plasma membrane fluidity. As SMPDL3b is upregulated in diabetes while insulin receptor signaling is impaired, our study was aimed at testing the hypothesis that SMPDL3b affects the generation of sphingolipids involved in the regulation of insulin receptor signaling.

Methods

For in vitro studies, control and SMPDL3b overexpressing human podocytes were used. Cells were treated with insulin (0.1 and 1 nM, 30 min) or ceramide-1-phosphate (C1P; 100 uM, 1h) and analyzed by Western blotting or PCR. Lipidomic analysis was performed using LC-MS analysis and TLC plates. Co-immunoprecipitation experiments were performed using HEK293 cells. For in vivo studies, podocyte-specific Smpdl3b deficient diabetic db/db mice were produced using Cre-LoxP technology. Starting at 4 weeks of age, vital parameters (weight, glycemia, urine) were measured bi-weekly. For C1P replacement therapy, diabetic mice were IP injected with 30 mg/kg C1P daily for 28 days. Mice from all experiments were sacrificed for in-depth phenotypical analysis. All animal studies were performed in accordance with the NIH IACUC Guide. For statistical analysis One-Way ANOVA followed by Bonferroni’s posttest or Student t-test were used.

Results

SMPDL3b binds to both IR isoforms (IRA and IRB) and, when in excess, competes with the binding of IRB to caveolin-1 and alters pro-survival insulin signaling in podocytes. In vivo, we demonstrated that kidneys of diabetic db/db mice are characterized by SMPDL3b excess and C1P deficiency, whereas podocyte-specific Smpdl3b deficient diabetic db/db mice show a normal C1P content and are protected from the development of diabetic kidney disease (DKD). Exogenous administration of C1P is sufficient to restore proper IR signaling in vitro and to protect from DKD in vivo.

Conclusion

Taken together, we identified new sphingolipid modulator of insulin signaling and demonstrated that replacement of deficient active sphingolipid species such as C1P may represent a novel approach to treat diabetic complications such as DKD.

Funding

• NIDDK Support