The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis

Tuomas Aleksi Tolvanen

Research output: ThesisDoctoral ThesisCollection of Articles

Abstract

Background: Type 2 diabetes, characterised by insulin resistance, is an everincreasing problem in the world. Diabetic nephropathy is a renal microvascular complication of diabetes that is a common cause of end-stage renal disease worldwide. Low levels of albumin can be detected in the urine at an early stage of diabetic nephropathy, and more severe albuminuria develops as the disease progresses. Insulin resistance is associated with albuminuria in patients with type 1 or type 2 diabetes. Also, the loss of podocytes plays a major role in the pathogenesis of diabetic nephropathy. The mechanisms underlying the development of insulin resistance and apoptosis in podocytes are still not fully understood. This study aimed to investigate the pathophysiological mechanisms leading to insulin resistance and podocyte apoptosis at the molecular level, and concentrated to the roles of CD2-associated protein (CD2AP) and SH2-domain containing 5’ inositol phosphatase 2 (SHIP2) in these processes. Results: We found that the lack of CD2AP in podocytes led to attenuated glucose uptake, without affecting PI3K-AKT-mediated insulin signalling. This led us to investigate the role of CD2AP in glucose transporter trafficking. Indeed, live-cell imaging revealed that internalized Glucose transporter 4 (GLUT4) was trapped in the perinuclear region of podocytes lacking CD2AP. CD2AP co-fractionated with known components of GLUT4 vesicles and interacted with clathrin and clathrin adaptor GGA2. These results suggest that CD2AP plays a role in GLUT4 vesicle trafficking. We further observed that the level and activity of SHIP2, an interaction partner of CD2AP, was increased in the absence of CD2AP in podocytes. Also, production of reactive oxygen species (ROS) and the rate of apoptosis were increased when CD2AP was lacking from podocytes. We hypothesized that inhibition of SHIP2 would decrease the production of ROS and apoptosis, but even though we detected reduced ROS production, inhibition of SHIP2 increased podocyte apoptosis in the absence of CD2AP. In the search for novel SHIP2 inhibitors we found an old anti-diabetic drug, metformin, to bind to and inhibit the activity of SHIP2 in silico, in vitro and in vivo. By inhibiting the activity of SHIP2, metformin increased glucose uptake and protected podocytes from apoptosis initiated by SHIP2 overexpression. SHIP2 exhibited higher activity in the kidney pieces received from nephrectomy patients with type 2 diabetes receiving non-metformin medication compared to nephrectomy patients without diabetes; in comparison, the activity of SHIP2 did not differ between metformin receiving patients with type 2 diabetes and people without diabetes. Patients with type 2 diabetes with metformin medication also showed reduced podocyte loss. Conclusions: In the light of our results we suggest a novel role for CD2AP in the regulation of sorting of internalized GLUT4 and regeneration of insulin responsive GLUT4 vesicle compartment. We also suggest that SHIP2 inhibitors, including metformin, can be used to reduce the oxidative stress of podocytes and to prevent podocyte loss, except in the case of patients suffering from reduced expression of CD2AP.
Original languageEnglish
Awarding Institution
  • University of Helsinki
Supervisors/Advisors
  • Lehtonen, Sanna, Supervisor
Award date18 Dec 2017
Place of PublicationHelsinki
Publisher
Print ISBNs978-951-51-3904-7
Electronic ISBNs978-951-51-3905-4
Publication statusPublished - 18 Dec 2017
MoE publication typeG5 Doctoral dissertation (article)

Fields of Science

  • Adaptor Proteins, Signal Transducing
  • +genetics
  • +metabolism
  • Apoptosis
  • Biological Transport, Active
  • Clathrin-Coated Vesicles
  • Cytoskeletal Proteins
  • Diabetic Nephropathies
  • DNA-Binding Proteins
  • Glucose
  • Insulin Resistance
  • Metformin
  • +pharmacology
  • Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases
  • Podocytes
  • +cytology
  • Reactive Oxygen Species
  • Transcription Factors
  • Transport Vesicles
  • 3111 Biomedicine

Cite this

Tolvanen, T. A. (2017). The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis. Helsinki : [T. A. Tolvanen].
Tolvanen, Tuomas Aleksi. / The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis. Helsinki : [T. A. Tolvanen], 2017. 95 p.
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abstract = "Background: Type 2 diabetes, characterised by insulin resistance, is an everincreasing problem in the world. Diabetic nephropathy is a renal microvascular complication of diabetes that is a common cause of end-stage renal disease worldwide. Low levels of albumin can be detected in the urine at an early stage of diabetic nephropathy, and more severe albuminuria develops as the disease progresses. Insulin resistance is associated with albuminuria in patients with type 1 or type 2 diabetes. Also, the loss of podocytes plays a major role in the pathogenesis of diabetic nephropathy. The mechanisms underlying the development of insulin resistance and apoptosis in podocytes are still not fully understood. This study aimed to investigate the pathophysiological mechanisms leading to insulin resistance and podocyte apoptosis at the molecular level, and concentrated to the roles of CD2-associated protein (CD2AP) and SH2-domain containing 5’ inositol phosphatase 2 (SHIP2) in these processes. Results: We found that the lack of CD2AP in podocytes led to attenuated glucose uptake, without affecting PI3K-AKT-mediated insulin signalling. This led us to investigate the role of CD2AP in glucose transporter trafficking. Indeed, live-cell imaging revealed that internalized Glucose transporter 4 (GLUT4) was trapped in the perinuclear region of podocytes lacking CD2AP. CD2AP co-fractionated with known components of GLUT4 vesicles and interacted with clathrin and clathrin adaptor GGA2. These results suggest that CD2AP plays a role in GLUT4 vesicle trafficking. We further observed that the level and activity of SHIP2, an interaction partner of CD2AP, was increased in the absence of CD2AP in podocytes. Also, production of reactive oxygen species (ROS) and the rate of apoptosis were increased when CD2AP was lacking from podocytes. We hypothesized that inhibition of SHIP2 would decrease the production of ROS and apoptosis, but even though we detected reduced ROS production, inhibition of SHIP2 increased podocyte apoptosis in the absence of CD2AP. In the search for novel SHIP2 inhibitors we found an old anti-diabetic drug, metformin, to bind to and inhibit the activity of SHIP2 in silico, in vitro and in vivo. By inhibiting the activity of SHIP2, metformin increased glucose uptake and protected podocytes from apoptosis initiated by SHIP2 overexpression. SHIP2 exhibited higher activity in the kidney pieces received from nephrectomy patients with type 2 diabetes receiving non-metformin medication compared to nephrectomy patients without diabetes; in comparison, the activity of SHIP2 did not differ between metformin receiving patients with type 2 diabetes and people without diabetes. Patients with type 2 diabetes with metformin medication also showed reduced podocyte loss. Conclusions: In the light of our results we suggest a novel role for CD2AP in the regulation of sorting of internalized GLUT4 and regeneration of insulin responsive GLUT4 vesicle compartment. We also suggest that SHIP2 inhibitors, including metformin, can be used to reduce the oxidative stress of podocytes and to prevent podocyte loss, except in the case of patients suffering from reduced expression of CD2AP.",
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Tolvanen, TA 2017, 'The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis', University of Helsinki, Helsinki .

The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis. / Tolvanen, Tuomas Aleksi.

Helsinki : [T. A. Tolvanen], 2017. 95 p.

Research output: ThesisDoctoral ThesisCollection of Articles

TY - THES

T1 - The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis

AU - Tolvanen, Tuomas Aleksi

N1 - M1 - 95 s. + liitteet

PY - 2017/12/18

Y1 - 2017/12/18

N2 - Background: Type 2 diabetes, characterised by insulin resistance, is an everincreasing problem in the world. Diabetic nephropathy is a renal microvascular complication of diabetes that is a common cause of end-stage renal disease worldwide. Low levels of albumin can be detected in the urine at an early stage of diabetic nephropathy, and more severe albuminuria develops as the disease progresses. Insulin resistance is associated with albuminuria in patients with type 1 or type 2 diabetes. Also, the loss of podocytes plays a major role in the pathogenesis of diabetic nephropathy. The mechanisms underlying the development of insulin resistance and apoptosis in podocytes are still not fully understood. This study aimed to investigate the pathophysiological mechanisms leading to insulin resistance and podocyte apoptosis at the molecular level, and concentrated to the roles of CD2-associated protein (CD2AP) and SH2-domain containing 5’ inositol phosphatase 2 (SHIP2) in these processes. Results: We found that the lack of CD2AP in podocytes led to attenuated glucose uptake, without affecting PI3K-AKT-mediated insulin signalling. This led us to investigate the role of CD2AP in glucose transporter trafficking. Indeed, live-cell imaging revealed that internalized Glucose transporter 4 (GLUT4) was trapped in the perinuclear region of podocytes lacking CD2AP. CD2AP co-fractionated with known components of GLUT4 vesicles and interacted with clathrin and clathrin adaptor GGA2. These results suggest that CD2AP plays a role in GLUT4 vesicle trafficking. We further observed that the level and activity of SHIP2, an interaction partner of CD2AP, was increased in the absence of CD2AP in podocytes. Also, production of reactive oxygen species (ROS) and the rate of apoptosis were increased when CD2AP was lacking from podocytes. We hypothesized that inhibition of SHIP2 would decrease the production of ROS and apoptosis, but even though we detected reduced ROS production, inhibition of SHIP2 increased podocyte apoptosis in the absence of CD2AP. In the search for novel SHIP2 inhibitors we found an old anti-diabetic drug, metformin, to bind to and inhibit the activity of SHIP2 in silico, in vitro and in vivo. By inhibiting the activity of SHIP2, metformin increased glucose uptake and protected podocytes from apoptosis initiated by SHIP2 overexpression. SHIP2 exhibited higher activity in the kidney pieces received from nephrectomy patients with type 2 diabetes receiving non-metformin medication compared to nephrectomy patients without diabetes; in comparison, the activity of SHIP2 did not differ between metformin receiving patients with type 2 diabetes and people without diabetes. Patients with type 2 diabetes with metformin medication also showed reduced podocyte loss. Conclusions: In the light of our results we suggest a novel role for CD2AP in the regulation of sorting of internalized GLUT4 and regeneration of insulin responsive GLUT4 vesicle compartment. We also suggest that SHIP2 inhibitors, including metformin, can be used to reduce the oxidative stress of podocytes and to prevent podocyte loss, except in the case of patients suffering from reduced expression of CD2AP.

AB - Background: Type 2 diabetes, characterised by insulin resistance, is an everincreasing problem in the world. Diabetic nephropathy is a renal microvascular complication of diabetes that is a common cause of end-stage renal disease worldwide. Low levels of albumin can be detected in the urine at an early stage of diabetic nephropathy, and more severe albuminuria develops as the disease progresses. Insulin resistance is associated with albuminuria in patients with type 1 or type 2 diabetes. Also, the loss of podocytes plays a major role in the pathogenesis of diabetic nephropathy. The mechanisms underlying the development of insulin resistance and apoptosis in podocytes are still not fully understood. This study aimed to investigate the pathophysiological mechanisms leading to insulin resistance and podocyte apoptosis at the molecular level, and concentrated to the roles of CD2-associated protein (CD2AP) and SH2-domain containing 5’ inositol phosphatase 2 (SHIP2) in these processes. Results: We found that the lack of CD2AP in podocytes led to attenuated glucose uptake, without affecting PI3K-AKT-mediated insulin signalling. This led us to investigate the role of CD2AP in glucose transporter trafficking. Indeed, live-cell imaging revealed that internalized Glucose transporter 4 (GLUT4) was trapped in the perinuclear region of podocytes lacking CD2AP. CD2AP co-fractionated with known components of GLUT4 vesicles and interacted with clathrin and clathrin adaptor GGA2. These results suggest that CD2AP plays a role in GLUT4 vesicle trafficking. We further observed that the level and activity of SHIP2, an interaction partner of CD2AP, was increased in the absence of CD2AP in podocytes. Also, production of reactive oxygen species (ROS) and the rate of apoptosis were increased when CD2AP was lacking from podocytes. We hypothesized that inhibition of SHIP2 would decrease the production of ROS and apoptosis, but even though we detected reduced ROS production, inhibition of SHIP2 increased podocyte apoptosis in the absence of CD2AP. In the search for novel SHIP2 inhibitors we found an old anti-diabetic drug, metformin, to bind to and inhibit the activity of SHIP2 in silico, in vitro and in vivo. By inhibiting the activity of SHIP2, metformin increased glucose uptake and protected podocytes from apoptosis initiated by SHIP2 overexpression. SHIP2 exhibited higher activity in the kidney pieces received from nephrectomy patients with type 2 diabetes receiving non-metformin medication compared to nephrectomy patients without diabetes; in comparison, the activity of SHIP2 did not differ between metformin receiving patients with type 2 diabetes and people without diabetes. Patients with type 2 diabetes with metformin medication also showed reduced podocyte loss. Conclusions: In the light of our results we suggest a novel role for CD2AP in the regulation of sorting of internalized GLUT4 and regeneration of insulin responsive GLUT4 vesicle compartment. We also suggest that SHIP2 inhibitors, including metformin, can be used to reduce the oxidative stress of podocytes and to prevent podocyte loss, except in the case of patients suffering from reduced expression of CD2AP.

KW - Adaptor Proteins, Signal Transducing

KW - +genetics

KW - +metabolism

KW - Apoptosis

KW - Biological Transport, Active

KW - Clathrin-Coated Vesicles

KW - Cytoskeletal Proteins

KW - Diabetic Nephropathies

KW - DNA-Binding Proteins

KW - Glucose

KW - Insulin Resistance

KW - Metformin

KW - +pharmacology

KW - Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases

KW - Podocytes

KW - +cytology

KW - Reactive Oxygen Species

KW - Transcription Factors

KW - Transport Vesicles

KW - 3111 Biomedicine

M3 - Doctoral Thesis

SN - 978-951-51-3904-7

PB - [T. A. Tolvanen]

CY - Helsinki

ER -

Tolvanen TA. The Roles of CD2AP and SHIP2 in insulin resistance and podocyte apoptosis. Helsinki : [T. A. Tolvanen], 2017. 95 p.