Renal Lipid Metabolism and Lipotoxicity in Diabetes

Renal lipotoxicity in diabetes

Journal Pre-proof

Renal lipid metabolism and lipotoxicity in diabetes Laongdao Thongnak MS , Anchalee Pongchaidecha PhD , Anusorn Lungkaphin PhD PII: DOI: Reference:

S0002-9629(19)30397-0 https://doi.org/10.1016/j.amjms.2019.11.004 AMJMS 950

To appear in:

The American Journal of the Medical Sciences

Received date: Accepted date:

6 June 2019 20 November 2019

Please cite this article as: Laongdao Thongnak MS , Anchalee Pongchaidecha PhD , Anusorn Lungkaphin PhD , Renal lipid metabolism and lipotoxicity in diabetes, The American Journal of the Medical Sciences (2019), doi: https://doi.org/10.1016/j.amjms.2019.11.004

This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Inc. on behalf of Southern Society for Clinical Investigation.

Renal lipid metabolism and lipotoxicity in diabetes Laongdao Thongnak MS1, Anchalee Pongchaidecha PhD1, Anusorn Lungkaphin PhD1,2,* 1

Department of Physiology, Faculty of Medicine, Chiang Mai University, Chiang Mai,

Thailand 2

Functional Food Research Center for Well-being, Chiang Mai University, Chiang Mai

University, Chiang Mai, Thailand *Corresponding author: Anusorn Lungkaphin, Department of Physiology, Faculty of Medicine, Chiang Mai University, Chiang Mai, 50200, Thailand Email: [email protected]; [email protected] Tel: +66.53-945362-4, Fax: +66.53-945365

Short title: Renal lipotoxicity in diabetes

The authors have no conflicts of interest to disclose.

Acknowledgements: This work was supported by Thailand Research Fund (RSA6080015 AL); the Royal Golden Jubilee PhD program (PhD/0063/2560 LT and AL); the Faculty of Medicine Research Fund, Chiang Mai University (AL and AP); Graduate Research Scholarship Chiang Mai University (LT) and the Functional Food Research Center for Well-being, Chiang Mai University (AL).

Abbreviation ABCA1

Adenosine triphosphate binding cassette transporter A1

ACC

Acetyl-CoA carboxylase

ACE

Angiotensin converting enzyme

ACE2

Angiotensin converting enzyme 2

ACO

Acyl-coenzyme A oxidase

ACR

Albumin to creatinine ratio

ACS

Acyl-CoA synthetase

AdipoR1

Adiponectin receptor 1

AdipoR2

Adiponectin receptor 2

ADRP

Adipose differentiation-related protein

AGEs

Advanced glycation end products

Akt

Protein kinase B

AMP

Adenosine monophosphate

AMPK

5’ adenosine monophosphate-activated protein kinase

Apo AI

Apolipoprotein AI

ATP

Adenosine triphosphate

AT1R

Angiotensin II type 1 receptor

AT2R

Angiotensin II type 2 receptor

Bcl-2

B-cell lymphoma 2

BSA

Bovine serum albumin

BUN

Blood urea nitrogen

BW

Body weight

CE/TC

Cholesterol ester/total cholesterol

ChREBP

Carbohydrate responsive element-binding protein

CPT1

Carnitine palmitoyltransferase 1

Cr.clearance

Creatinine clearance

CRP

C-reactive protein

DGAT1

Diacylglycerol O-acyltransferase 1

DGAT2

Diacylglycerol O-acyltransferase 2

ER

Endoplasmic reticulum

ERK1/2

Extracellular signal-regulated protein kinases 1 and 2

ERRα

Estrogen related receptor alpha

ETC

Electron transport chain

FA

Fatty acids

Fatp4

Fatty acid transport protein 4

FAT

Fatty acid translocase

FAS

Fatty acid synthase

FBG

Fasting blood glucose

FFA

Free fatty acids

FGF21

Fibroblast growth factor 21

FoxO1

Forkhead box O1

FoxO3a

Forkhead box O3

FXR

Farnexoid X receptor

GMB

Glomerular basement membrane

G6Pc

Glucose-6-phosphatase catalytic-subunit

HbA1c

Hemoglobin A1c

HDL

High-density lipoproteins

HFD

High fat diet

HMG-CoA

3-hydroxy-3-methyl-glutaryl-CoA

HGECs

Human glomerular epithelial cells

HOMA-β

Homeostatic model assessment of β-cell function

HOMA-IR

Homeostatic model assessment of insulin resistance

HO-1

Heme oxygenase-1

HRGECs

Human renal glomerular epithelial cells

IL-1

Interleukin-1

IL-6

Interleukin-6

IL-8

Interleukin-8

Keap1

Kelch-like ECH-associated protein 1

KW

Kidney weight

KW/BW

Kidney weight/body weight

KW/TL

Kidney weight/tibia length

LDL

Low-density lipoprotein

L-PK

Liver pyruvate kinase

LPO

Lipid peroxidation

LXR-α

Liver X receptor alpha

LXR-β

Liver X receptor beta

MCD

Malonyl-CoA decarboxylase

MCP-1

Monocyte chemotactic protein 1

MDA

Malondialdehyde

MUFAs

Monounsaturated fatty acids

NAD, NADH

Nicotinamide adenine dinucleotide

NEFA

Non-esterified fatty acids

NF-κB

Nuclear factor-kappa B

NRF-1

Nuclear respiratory factor-1

Nrf2

Nuclear factor erythroid 2 (NF-E2)-related factor 2

p-ACC

Phosphorylated acetyl-CoA carboxylase

PAI-1

Plasminogen activator inhibitor-1

PCR

Protein to creatinine ratio

PEPCK

Phosphoenolpyruvate carboxykinase

p-ERK

Phosphorylate extracellular signal-regulated protein kinases

PGC-1α

Peroxisome proliferator-activated receptor gamma coactivator-1 alpha

PI3K

Phosphoinositide 3-kinase

PMN

Polymorphonuclear leukocytes

PPARα

Peroxisome proliferator-activated receptor alpha

PPAR

Peroxisome proliferator-activated receptor gamma

ROS

Reactive oxygen species

SCD1

Stearoyl-CoA desaturase-1

SCD2

Stearoyl-CoA desaturase-2

SIRT1

NAD-dependent deacetylase sirtuin-1

SOD1

Superoxide dismutase 1

SOD2

Superoxide dismutase 2

SREBP

Sterol regulatory element-binding protein

STZ

Streptozotocin

TAG

Triacylglycerol

TBARs

Thiobarbituric acid reactive

TCA

Tricarboxylic acid cycle

TG

Triglyceride

TGF-β

Transforming growth factor beta

TNF-α

Tumor necrosis factor alpha

TUNEL

Terminal deoxynucleotidyl transferase dUTP nick end labeling

T1D

Type 1 diabetes

T2D

Type 2 diabetes

UACR

Urinary albumin/creatinine ratio

UAE

Urine albumin excretion

VEGF

Vascular endothelial growth factor

ZDF

Zucker diabetic fatty

1-MNA

1-methylnicotinamide

3-NT

3-Nitrotyrosine

4-HNE

4-Hydroxynonenal

Abstract The pathogenesis of diabetic kidney disease is a complex process caused by both glucotoxicity and lipotoxicity, due to lipid accumulation. In cases of diabetic animals, lipid deposition is found in both tubular and glomerular portions of the kidneys, which are the major sites of diabetic nephropathy lesions. The aim of this review was to provide insights into the mechanisms that lead to the development of renal lipid accumulation and the effects of renal lipotoxicity in the diabetic condition. An increased number of lipogenic genes and a decreased number of lipid oxidation genes are also detected in diabetic kidneys, both of which lead to lipid accumulation. The induction of oxidative stress, inflammation, fibrosis, and apoptosis caused by lipid accumulation and lipid metabolites is called lipotoxicity. Renal lipotoxicity due to derangement in lipid metabolism may be a pathogenic mechanism leading to diabetic nephropathy and renal dysfunction.

Keywords: Renal lipid accumulation; renal lipotoxicity; diabetic condition; renal function

Introduction Diabetes has reached epidemic proportions worldwide and is a leading risk factor for kidney disease. Between one third and one half of diabetic patients develop kidney disease.1 Diabetic nephropathy (DN) is a microangiopathy complication which occurs in 20% of diabetic patients that can develop into end-stage renal disease, resulting in the high mortality rate in diabetic patients. Metabolic and hemodynamic changes are correlative factors in the pathogenesis of DN (Figure 1). Prolonged hyperglycemia triggers multiple and complex molecular mechanisms to promote diabetic nephropathy, including: (1) Advanced glycation end products (AGEs) formation, which are glycated proteins or lipids that act as the transdifferentiation of epithelial cells to aggravate tubulointerstitial fibrosis development.2 (2) Protein kinase C (PKC) activation with subsequently upregulated growth factor expression may lead to glomerular basement membrane thickening. (3) Hexosamine and polyol pathway activation promotes ROS production though the increase in oxidative stress and suppression of anti-oxidant capacity.3 (4) Hyperglycemia promotes oxidative stress, the secretion of proinflammatory cytokines, and other pathways, which consequently induce cellular death.4 In addition, hyperglycemia induces decreased nitric oxide (NO) production through the suppression of the Akt-eNOS pathway5 and ultimately increased angiotensin II sensitivity, resulting in increased intracapillary pressure on the efferent side of glomerulus.6 According to hemodynamic changes of glomerulus hyperperfusion and hyperfiltration, kidney damage as demonstrated by leakage of albumin or protein was observed in the early stage of diabetic kidney disease patients.7, 8 The role of dyslipidemia in the pathogenesis of diabetes has been more recently studied.9

Dyslipidemia, which is characterized by the elevation of plasma triglyceride, low-density lipoprotein (LDL), and very-low density lipoprotein (VLDL) along with the decreased highdensity lipoprotein (HDL) levels, is commonly seen in diabetic patients.10, 11 Insulin is an anabolic hormone that suppresses triglyceride lipolysis to free fatty acids via inhibition of hormonesensitive lipase (HSL) in adipose tissue.12 Additionally, insulin plays a crucial role in regulating serum VLDL concentration by activating lipoprotein lipase (LPL) to suppress hepatic VLDL synthesis and stimulate VLDL clearance.9 In both patients with type 1 diabetes with poor glycemic control and patients with type 2 diabetes with hyperinsulinemia, the inhibition of triglyceride lipolysis is diminished, resulting in increased free fatty acid delivery to the liver. One major source of triglyceride in the liver, hepatic de novo lipogenesis, may also indirectly exacerbate insulin resistance by 5’ adenosine monophosphate-activated protein kinase (AMPK) suppression.13 Moreover, in diabetic patients, an increased VLDL-TG secretion through stimulation of microsomal triglyceride transfer protein MTP and impaired VLDL-TG removal mainly caused by reduced activity of LPL leading to serum lipid abnormalities can be found.10, 14 These alterations account for the pathophysiology of diabetic dyslipidemia. The downregulation of genes for lipolysis and fatty acid β-oxidation along with an increase in lipid synthesis in diabetes associated with triglyceride accumulation in kidneys have both been reported.15, 16 The imbalance in lipid homeostasis may contribute to renal lipid accumulation and lipotoxicity in cases of diabetes, which subsequently leads to kidney dysfunction and diabetic nephropathy. In clinical studies, lipid accumulation and lipid droplets in glomerular and tubular portions were found in kidney biopsy samples from patients with diabetic nephropathy and diabetic kidney disease.17-19 Lipotoxicity, through excess adipokines from renal ectopic fat

deposits and toxic metabolites of fatty acids, activates various signaling pathways, including those which result in oxidative stress, inflammation, fibrosis, and apoptosis, leading to renal cell damage and dysfunction. In addition, lipotoxicity can also enhance insulin resistance via the activation of the protein kinase pathway and vice versa,20 which increases the pathogenicity and severity of diabetes. Despite the significant impact of renal lipid metabolism, the pathophysiological consequences of renal lipid accumulation have only partially been elucidated. This review focuses on the association between renal lipotoxicity and the diabetic condition and the mechanisms involved. Renal Lipid Metabolism in the Non-Diabetic Condition Lipids are the major source of energy in the renal cortex, which has a high energy demand to maintain filtration, secretion, and absorptive functions.21 As shown in Figure 2, fatty acids can enter the renal proximal tubule cells bi-directionally by means of a high-affinity fatty acid translocase (FAT)/CD36,22 from the basolateral membrane, and by endocytosis from the apical membrane.23, 24 Endocytosis of the fatty acids occurs in albumin-bound form, which is subsequently hydrolyzed into albumin and free fatty acids in the proximal tubular cells.25 Albumin is returned to the circulation by transcytosis, while fatty acids accumulate in the fatty acid pool inside the renal cell. Fatty acids that enter the cell are converted to fatty acyl-CoA by acyl-CoA synthetase (ACS). Fatty acyl-CoA is transported into mitochondriavia carnitine palmitoyltransferase 1 (CPT1), the transport being the rate-limiting step for fatty acid oxidation.26 Fatty acyl-CoA is converted into acetyl-CoA by β-oxidation and adenosine triphosphate (ATP) is subsequently

generated by the tricarboxylic acid cycle (TCA) or Kreb’s cycle and the electron transport chain (ETC).27 Excess acetyl-CoA can be converted to malonyl-CoA by acetyl-CoA carboxylase (ACC).28 An increase in malonyl-CoA not only inhibits CPT1 function, leading to a reduction in the mitochondrial influx of fatty acyl-CoA, but also activates fatty acid synthase (FAS) activity to convert fatty acyl-CoA into palmitate, which enters the fatty acid pool and is stored inside the cell as triglyceride.29, 30 Regulation of Renal Lipid Metabolism The homeostasis of lipid metabolism is regulated by both fatty acid oxidation or fatty acid utilization and fatty acid synthesis. The level of membrane transport protein expression, such as CD36/FAT and CPT-1, administer the level of fatty acid influx into the cell and mitochondria, respectively.31 These proteins are regulated by peroxisome proliferator-activated receptor alpha (PPARα) transcription factor.32-35 Moreover, peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α), a transcription factor of mitochondrial biogenesis, and estrogen-related receptor alpha (ERRα) work together with PPARα to regulate the transcription of the fatty acid oxidation genes, such as acyl-CoA dehydrogenase. In addition, the level of malonyl-CoA, a potent mitochondrial fatty acid uptake inhibitor, also regulates fatty acid homeostasis secondary to the inhibition of CPT-1. Two enzymes relate to the level of malonyl-CoA: ACC and malonyl-CoA decarboxylase (MCD) enzymes. ACC enzymes catalyze the carboxylation of acetyl-CoA to produce malonyl-CoA, while MCD converts malonyl-CoA to acetyl-CoA. Several transcription factors can regulate ACC expression, including AMPK,36 sterol regulatory element-binding protein (SREBP), carbohydrate responsive element-binding protein

(ChREBP), and nuclear respiratory factor-1 (NRF-1). Phosphorylation or an inactive form of ACC can activate MCD activity, resulting in the activation of fatty acid β-oxidation. Also, the products of the β-oxidation, such as the increase in NADH/NAD+ or acetyl-CoA/CoA ratios, have been shown to have a negative feedback on their reaction.37 As mentioned above, there are several factors regulating lipid metabolism and lipid homeostasis that lead to the balance of fat in the body. ATP is used in the regulation of arteriole vasoconstriction and activity of the epithelial Na+ channel and is a major source of energy for maintaining renal function.38 The marked decreases in mitochondria and extracellular ATP production in kidney of diabetic mice are directly associated with renal glomerular and tubular dysfunction.39, 40 Under conditions that increased the AMP/ATP ratio or resulted in a low level of ATP production, β-oxidation which provides a more energetic yield of ATP, arises predominantly due to activation of the AMPK pathway.41 AMPK is an enzyme that plays a crucial role in cellular energy homeostasis through activating utilization and inhibiting synthesis activity. AMPK, by inhibiting ACC activity, decreases malonyl-CoA level, increases CPT1 activity and thus increases fatty acid oxidation.42-44 AMPK also promotes cellular fatty acid utilization by inhibiting either mTORs-SREBP1 or HMGCoA reductase, which relate to lipid synthesis and cholesterol synthesis45, and also deactivates phosphoenolpyruvate carboxykinase (PEPCK) enzyme that controls the gluconeogenic pathway.46 Moreover, AMPK has been found to activate PPARα to stimulate fatty acid oxidation via increased PGC-1α activity (Figure 3).47, 48 Regardless of these studies, the factors that regulate the mechanism of lipid metabolism have still not been completely elucidated.

Renal Lipid Metabolism in the Diabetic Condition In diabetic kidneys, the enhancement of fatty acid synthesis together with the suppression of fatty acid oxidation are the major cause of renal lipid accumulation. The impairment of lipid metabolism with underlying mechanisms of renal lipotoxicity are presented in this review.

In Vitro Studies Table 1 shows that high glucose conditions stimulate fatty acid synthesis and downregulate fatty acid oxidation in renal cells. Increases in de novo lipogenesis by the activation of SREBP and ChREBP with the decreases in fatty acid oxidation genes, such as PPARα, PGC-1α, and ERR1-α were observed in mesangial cells incubated under high glucose conditions.47, 49 The elevation of acetyl-CoA, an intermediate energetic substance, can stimulate the translocation of SREBP and ChREBP transcription factors from cytoplasm into the nucleus50, 51

and then activate downstream protein expression. SREBP-1 can stimulate ACC and FAS

activities, while ChREBP can activate liver pyruvate kinase (L-PK) to convert acetyl-CoA to triglycerides, which then accumulate in the cells.27, 52 In addition, SREBP-2 can activate HMGCoA reductase and HMG-CoA synthase to stimulate de novo cholesterol synthesis.53 Triglyceride and cholesterol accumulation in glomerular and tubule-interstitial cells were also observed with the increased expression of SREBP-1, ChREBP, and their related lipogenic genes, such as ACC and FAS, in high-fat diet (HFD)-fed mice.54, 55 These findings emphasized that SREBP and ChREBP are the major transcription factors to regulate fatty acid synthesis in the diabetic condition.

Down-regulation of fatty acid oxidation genes, such as PPARα, PGC-1, and CPT-1, that resulted in a decrease in fatty acid β-oxidation has been shown in the hyperglycemic condition.47, 49, 56 Because of the diminished AMPK activity by hyperglycemia, PPARα, PGC-1, and CPT-1 expression, which are downstream proteins of the AMPK pathway that regulate fatty acid oxidation, were decreased. Finally, impaired fatty acid oxidation accelerated renal lipid accumulation and increased fatty acid synthesis in the diabetic condition. In addition, in human renal glomerular epithelial cells (HRGECs) incubated under high glucose and cholesterol conditions, ATP-binding cassette transporter A1 (ABCA1) cholesterol transporter was decreased, correlating with an increase in renal cholesterol content.18 The decreasing level of ABCA1 expression was also observed in diabetes and nephropathy mouse models.18, 57, 58 ABCA1 is involved in the cellular lipid removal pathway mediating cholesterol efflux to lipid-poor apolipoprotein AI (apo AI) and nascent high-density lipoproteins (HDL).59 Thus, the decrease in ABCA1 may have the effect of reducing the HDL level in the diabetic condition associated with dyslipidemia. These findings indicate that impaired cholesterol efflux transporters may be a part of the development of dyslipidemia and subsequent lipid accumulation. In Vivo Studies Tables 2 and 3 present the in vivo studies in diabetic animal models. Type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM) conditions impair the homeostasis of fat metabolism by accelerating fatty acid synthesis as shown by the upregulated expression of SREBP-1, SREBP-2, ChREBP,47, 49, 56-58, 60-64 and adipose differentiation-related protein (ADRP) or adipophilin,64-66 a specific marker of lipid accumulation,67 which is a lipid droplet-associated

protein expressed during adipocyte cell differentiation.68 Moreover, the activity of phosphorylated acetyl-CoA carboxylase (p-ACC) was decreased due to increased malonyl-CoA, which can inhibit CPT-1 function and increase FAS activity as well. Impaired fatty acid transport into mitochondria via CPT-1, as well as the increase in FAS activity, leads to increased triglyceride synthesis and lipid storage in renal cells as shown by the increases identified by positive Oil red O staining.18, 60, 61, 63, 65, 66, 69, 70 In addition, increasing levels of fatty acids in circulation in a dyslipidemic condition may deliver more albumin-bound fatty acids to the apical membrane, promoting fatty acid transport into renal cells. The level of stearoyl-CoA desaturase (SCD), an enzyme that catalyzes the formation of monounsaturated fatty acids (MUFAs), specifically oleate and palmitoleate from saturated fatty acids such as stearoyl-CoA and palmitoyl-CoA, is an area of controversy in diabetic models. The SCD1 level was found to be elevated together with an increase in fatty acid synthesis in T1DM Akita mice57 and T1DM db/db mice.63 Contrary to this, in HFD-induced diabetic mice, there may be an SCD1 deficiency,71 which can directly cause endoplasmic reticulum (ER) stress, leading to apoptosis of proximal tubular epithelial cells. The excessive monounsaturated fatty acids may be converted to polyunsaturated fatty acids without stimulating lipid synthesis. Furthermore, the reasons for the controversy around the SCD1 level could be attributed to the differing etiology in each diabetic model. Both T1DM and T2DM diabetic models show decreased fatty acid oxidation as evidenced by down-regulation of fatty acid oxidation genes, such as PPARα, PGC-1α, and ERR1α.47, 49, 56, 61, 62, 69 This effect may be a result of the decreased NAD+-dependent deacetylase sirtuin 1 (SIRT1) activation due to impaired insulin levels and signaling.47, 61, 69 In physiological

conditions, SIRT1 stimulates deacetylation of PGC1α and FOXO1 which in turn activates fatty acid oxidation.72 Consequently, a decrease in SIRT1 may reduce fatty acid oxidation through SIRT1-PGC1α signaling and lead to lipid accumulation in the diabetic condition. In addition, a long-term increase in fatty acid β-oxidation with impaired mitochondrial function and hyperglycemia can also activate ROS production and contribute to oxidative stress. Oxidative stress can in turn negatively affect the electron transport chain complexes in mitochondria, leading to a decrease in ATP production due to incomplete β-oxidation.73 This may be a cause of decreased fatty acid oxidation in patients with diabetes. Therefore, impaired homeostasis of fat metabolism by accelerating fatty acid synthesis with diminished fatty acid oxidation initiates renal lipid accumulation in both T1DM and T2DM conditions. Renal Lipotoxicity in the Diabetic Condition The dyslipidemic condition delivers more albumin-bound fatty acids transport into the renal cells, leading to overloading of fatty acids in mitochondria. In addition to activation of NADPH oxidase activity, long-term increasing fatty acid level activates mitochondrial dysfunction with incomplete β-oxidation which generates ROS production and subsequently induces oxidative stress in renal tissues. In addition, hyperglycemia, hyperinsulinemia, and low levels of adiponectin in the diabetic condition decrease AMPK activation,74 which is a regulator of energy balance, and subsequently decrease fatty acid oxidation. The increased AMP/ATP ratio caused by mitochondrial dysfunction and diminished fatty acid oxidation directly affects renal proximal tubular epithelial cells, which involve high levels of ATP consumption and mainly depend on the β-oxidation. Surplus fatty acids due to impaired β-oxidation are restored in the form of triglyceride inside the renal cells by activating the fatty acid synthesis pathway. In the

diabetic mice model, SREBP, ChREBP and lipogenic signaling proteins such as FAS and ACC are upregulated in the renal cells. Both decreases in fatty acid oxidation and increases in fatty acid synthesis due to hyperglycemia and dyslipidemia result in renal lipid accumulation in diabetes. Increased renal lipid accumulation, via a process termed lipotoxicity, can stimulate several signaling pathways leading to oxidative stress, inflammation, ER stress, fibrosis and apoptosis,75 which all lead to a deterioration in renal function.76 Oxidative stress in the diabetic kidney promotes peroxidation of lipids that accumulated in renal cells. The level of renal 4hydroxynonenal (4-HNE), a marker of lipid peroxidation, was significantly increased in streptozotocin-induced diabetic models,70 HFD-induced diabetes60 and also in the Zucker fatty diabetic model.66 Moreover, 3-nitrotyrosine (3-NT), 8-hydroxyguanosine (8-OHdG), 8isoprostane and malondialdehyde (MDA), which are lipid peroxidation molecules, were significantly augmented in the diabetic condition.47, 49, 56, 58, 60, 61, 70, 77 Moreover, decreasing in anti-oxidant effects of adipokines, such as adiponectin in diabetes also promotes oxidative damage.78 Thus, decreasing levels of antioxidant enzymes along with the increase in lipid peroxidation molecules are associated with renal lipid accumulation induced oxidative stress in the kidneys of diabetic models. Excess lipid and lipid metabolite molecules can induce the infiltration of inflammatory cells into renal cells as shown by macrophage infiltration and an increased release of inflammatory cytokines, such as interleukin (IL)-6, and tumor necrosis factor (TNF)-α.18, 57, 62, 66, 70, 77 These findings were consistent with those of Kim et al58 and Wang et al63, who showed improper activation of the nuclear factor (NF)-κB pathway resultied in upregulation of proinflammatory cytokines in diabetic mice.

Adiponectin, an adipocytokine that regulates fat metabolism by activating the AMPKPPARα pathway through adiponectin receptors (AdipoR1 and AdipoR2), has been reported to correlate with renal dysfunction.79 A previous study demonstrated an anti-inflammatory effect of adiponectin through the inhibition of the TNF-α and NF-κB pathways.80 Adiponectin also exerts antioxidant effects by inhibition of ROS production induced by hyperglycemia and oxidized LDL in the diabetic condition.81, 82 It has been reported that AdipoR1 is mainly expressed in the glomerular and proximal tubular cells; thereby a decrease in adiponectin receptor expression leads to impaired adiponectin signaling. Decreasing levels of serum adiponectin were found in the T2DM model,58, 61, 62, 64 which was related to renal dysfunction.83 Moreover, decreasing in antioxidant and anti-inflammatory effects of adiponectin promote oxidative stress and inflammatory damage in the diabetic condition.78 However, it has been shown that T1DM patients had significantly higher serum adiponectin levels than nondiabetics.84 The inconsistency of adiponectin level in diabetic patients might be due to the differences in etiology of the disease. Inflammatory cytokines released from inflammatory cells in fat deposition can stimulate not only inflammation but also profibrotic and growth factors such as CTGF, TGF-β1 and collagen type IV which contribute to the development of fibrosis.47, 49, 56-58, 60, 61, 63, 65, 66, 70, 77, 85 Oxidative stress, inflammation, and fibrosis can directly activate the apoptotic pathway. In addition, metabolites of fatty acid such as fatty acyl CoA, ceramide, and DAG can activate the caspase cascade and induce apoptosis as evidenced by the increase in apoptotic nuclei in TUNEL assay and an increased Bax/Bcl-2 ratio.47, 49, 56, 61, 62, 70, 71, 77, 85 In Zucker Diabetic Fatty (ZDF) rats, elevated levels of ceramide as well as increasing apoptosis were observed in

pancreatic islet cells,86 supporting the possibility that dyslipidemia could induce insulin resistance and enhance the severity of hyperglycemia by impairing β-cells in the pancreas.87 In addition, the overproduction of metabolites of fatty acids and reactive oxygen species (ROS) from the TCA cycle can directly affect serine phosphorylation of insulin receptor substrates (IRS) by PKC, thus augmenting insulin resistance88. Therefore, impaired renal lipid metabolism and lipotoxicity have been found to lead to renal dysfunction in diabetic kidney disease. Similar patterns of renal lipid accumulation associated with kidney dysfunction and the development of histological lesions were also observed in clinical studies in patients with T2DM nephropathy.17, 18, 65 Histological staining of renal tissue under the conditions of diabeticinduced lipid accumulation revealed renal injury, glomerular hypertrophy, increased mesangial cell expansion, matrix accumulation, and tubulointerstitial fibrosis.17, 18, 47, 49, 56-58, 60-65, 69, 77, 85 The attenuation of podocyte number and podocyte markers, including synaptopodin, nephrin, and podocin, was also found in the diabetic condition.17, 57, 63, 65 Increases in the kidney weight/body weight (KW/BW) ratio, urinary albumin (albuminuria) or urinary albumin/creatinine ratio (UACR), serum blood urea nitrogen (BUN), and serum creatinine in diabetic models also indicated both renal glomerular and tubular dysfunction resulting from lipid accumulation and toxicity. However, serum creatinine and BUN levels were not significantly elevated in some studies,17, 47, 49, 58, 61, 66, 69 which might be explained by the variations in procedures used in the studies including different strains of animals, unmatched reagents, differing formulae of the food, and incomparable duration of the diabetic condition in these studies.

From these results, it might be concluded that apart from glucotoxicity, renal lipid accumulation-induced cellular damage by activated oxidative stress, inflammation, fibrosis and apoptotic pathways or renal lipotoxicity in the diabetic condition could be a contributing factor for renal dysfunction and progressive diabetic complications, especially diabetic nephropathy. However, there may be other intracellular signaling pathways involved in renal lipid metabolism and lipotoxicity which have not currently been fully established and require further clarification. Conclusions Glycolysis is decreased because of the impairment of insulin signaling and/or insulin deficiency in the diabetic condition, leading to a depletion of cellular ATP.89 The AMPK pathway is subsequently activated to increase ATP production by promoting fatty acid β-oxidation. Nevertheless, a high rate or long-term occurrence of fatty acid oxidation with a deterioration of mitochondrial biosynthesis can generate ROS as a result of incomplete β-oxidation. Concomitant with the oxidative stress from hyperglycemia, fatty acid oxidation is decreased and intermediate substances of fat metabolism lead to accumulated fat products in the form of triglycerides. Reduced activation of energy metabolism genes, such as PPARα, and PGC-1α, due to decreased AMPK signaling, is followed by the down-regulation of fatty acid oxidation genes concurrent with the activation of the genes that are involved in triglyceride synthesis. These alterations result in increased triglyceride synthesis and subsequent accumulation in proximal tubular cells.90 In addition, reduced expression of the cholesterol transporter, ABCA1, leads to a reduction in HDL and the recycling of excess cholesterol in the circulation.91 These factors may be the cause of or at least a contributor to renal lipid accumulation in the diabetic condition. Both the increase in lipogenesis and the decrease in lipolysis result in lipid accumulation in

renal cells, which leads to the production of toxic metabolites. This production in turn induces inflammatory cell infiltration, the production of inflammatory cytokines and activation of fibrotic and apoptotic pathways known aslipotoxicity.92 To summarize, renal lipotoxicity due to derangement in lipid metabolism may be a pathogenic mechanism leading to diabetic nephropathy and renal dysfunction.

Acknowledgements This work was supported by Thailand Research Fund (RSA6080015 AL); the Royal Golden Jubilee PhD program (PhD/0063/2560 LT and AL); the Faculty of Medicine Research Fund, Chiang Mai University (AL and AP); Graduate Research Scholarship Chiang Mai University (LT) and the Functional Food Research Center for Well-being, Chiang Mai University (AL).

References 1.

Bakris GL. Recognition, pathogenesis, and treatment of different stages of nephropathy in patients with type 2 diabetes mellitus. Mayo Clin Proc. 2011;86(5):444-56.

2.

Baragetti I, El Essawy B, Fiorina P. Targeting Immunity in End-Stage Renal Disease. Am J Nephrol. 2017;45(4):310-9.

3.

Aronson D. Hyperglycemia and the pathobiology of diabetic complications. Adv Cardiol. 2008;45:1-16.

4.

Volpe CMO, Villar-Delfino PH, Dos Anjos PMF, et al. Cellular death, reactive oxygen species (ROS) and diabetic complications. Cell Death Dis. 2018;9(2):119.

5.

Taguchi K, Matsumoto T, Kamata K, et al. Akt/eNOS pathway activation in endotheliumdependent relaxation is preserved in aortas from female, but not from male, type 2 diabetic mice. Pharmacol Res. 2012;65(1):56-65.

6.

Papadopoulou-Marketou N, Chrousos GP, Kanaka-Gantenbein C. Diabetic nephropathy in type 1 diabetes: a review of early natural history, pathogenesis, and diagnosis. Diabetes Metab Res Rev. 2017;33(2).

7.

Perkins BA, Ficociello LH, Roshan B, et al. In patients with type 1 diabetes and new-onset microalbuminuria the development of advanced chronic kidney disease may not require progression to proteinuria. Kidney Int. 2010;77(1):57-64.

8.

Persson F, Rossing P. Diagnosis of diabetic kidney disease: state of the art and future perspective. Kidney Int Suppl (2011). 2018;8(1):2-7.

9.

Hirano T. Pathophysiology of Diabetic Dyslipidemia. J Atheroscler Thromb. 2018;25(9):771-82.

10.

Reaven GM, Greenfield MS. Diabetic hypertriglyceridemia: evidence for three clinical syndromes. Diabetes. 1981;30(Suppl 2):66-75.

11.

Taskinen MR. Diabetic dyslipidaemia: from basic research to clinical practice. Diabetologia. 2003;46(6):733-49.

12.

Lewis GF, Carpentier A, Adeli K, et al. Disordered fat storage and mobilization in the pathogenesis of insulin resistance and type 2 diabetes. Endocr Rev. 2002;23(2):201-29.

13.

Verges B. Abnormal hepatic apolipoprotein B metabolism in type 2 diabetes. Atherosclerosis. 2010;211(2):353-60.

14.

Pykalisto OJ, Smith PH, Brunzell JD. Determinants of human adipose tissue lipoprotein lipase. Effect of diabetes and obesity on basal- and diet-induced activity. J Clin Invest. 1975;56(5):1108-17.

15.

Saudek CD, Eder HA. Lipid metabolism in diabetes mellitus. Am J Med. 1979;66(5):84352.

16.

Wang C, Li L, Liu S, et al. GLP-1 receptor agonist ameliorates obesity-induced chronic kidney injury via restoring renal metabolism homeostasis. PLoS One. 2018;13(3):e0193473.

17.

Herman-Edelstein M, Scherzer P, Tobar A, et al. Altered renal lipid metabolism and renal lipid accumulation in human diabetic nephropathy. J Lipid Res. 2014;55(3):561-72.

18.

Yin QH, Zhang R, Li L, et al. Exendin-4 Ameliorates Lipotoxicity-induced Glomerular Endothelial Cell Injury by Improving ABC Transporter A1-mediated Cholesterol Efflux in Diabetic apoE Knockout Mice. J Biol Chem. 2016;291(51):26487-501.

19.

Yang W, Luo Y, Yang S, et al. Ectopic lipid accumulation: potential role in tubular injury and inflammation in diabetic kidney disease. Clin Sci (Lond). 2018;132(22):2407-22.

20.

Yazici D, Sezer H. Insulin Resistance, Obesity and Lipotoxicity. Adv Exp Med Biol. 2017;960:277-304.

21.

Epstein FH. Oxygen and renal metabolism. Kidney Int. 1997;51(2):381-5.

22.

Pepino MY, Kuda O, Samovski D, et al. Structure-function of CD36 and importance of fatty acid signal transduction in fat metabolism. Annu Rev Nutr. 2014;34:281-303.

23.

Yokoi H, Yanagita M. Targeting the fatty acid transport protein CD36, a class B scavenger receptor, in the treatment of renal disease. Kidney Int. 2016;89(4):740-2.

24.

Moestrup SK, Nielsen LB. The role of the kidney in lipid metabolism. Curr Opin Lipidol. 2005;16(3):301-6.

25.

Brunskill NJ. Albumin and proximal tubular cells--beyond endocytosis. Nephrol Dial Transplant. 2000;15(11):1732-4.

26.

Stadler K, Goldberg IJ, Susztak K. The evolving understanding of the contribution of lipid metabolism to diabetic kidney disease. Curr Diab Rep. 2015;15(7):40.

27.

Shimano H. Sterol regulatory element-binding proteins (SREBPs): transcriptional regulators of lipid synthetic genes. Prog Lipid Res. 2001;40(6):439-52.

28.

Wakil SJ, Abu-Elheiga LA. Fatty acid metabolism: target for metabolic syndrome. J Lipid Res. 2009;50 Suppl:S138-43.

29.

Clapham JC, Arch JR. Thermogenic and metabolic antiobesity drugs: rationale and opportunities. Diabetes Obes Metab. 2007;9(3):259-75.

30.

Browning JD, Horton JD. Molecular mediators of hepatic steatosis and liver injury. J Clin Invest. 2004;114(2):147-52.

31.

Yang X, Okamura DM, Lu X, et al. CD36 in chronic kidney disease: novel insights and therapeutic opportunities. Nat Rev Nephrol. 2017;13(12):769-81.

32.

Motojima K, Passilly P, Peters JM, et al. Expression of putative fatty acid transporter genes are regulated by peroxisome proliferator-activated receptor alpha and gamma activators in a tissue- and inducer-specific manner. J Biol Chem. 1998;273(27):16710-4.

33.

Poirier H, Niot I, Monnot MC, et al. Differential involvement of peroxisome-proliferatoractivated receptors alpha and delta in fibrate and fatty-acid-mediated inductions of the

gene encoding liver fatty-acid-binding protein in the liver and the small intestine. Biochem J. 2001;355(Pt 2):481-8. 34.

Rakhshandehroo M, Hooiveld G, Muller M, et al. Comparative analysis of gene regulation by the transcription factor PPARalpha between mouse and human. PLoS One. 2009;4(8):e6796.

35.

Burri L, Thoresen GH, Berge RK. The Role of PPARalpha Activation in Liver and Muscle. PPAR Res. 2010;2010.

36.

Valentine RJ, Coughlan KA, Ruderman NB, et al. Insulin inhibits AMPK activity and phosphorylates AMPK Ser(4)(8)(5)/(4)(9)(1) through Akt in hepatocytes, myotubes and incubated rat skeletal muscle. Arch Biochem Biophys. 2014;562:62-9.

37.

Kerbey AL, Radcliffe PM, Randle PJ. Diabetes and the control of pyruvate dehydrogenase in rat heart mitochondria by concentration ratios of adenosine triphosphate/adenosine diphosphate, of reduced/oxidized nicotinamide-adenine dinucleotide and of acetylcoenzyme A/coenzyme A. Biochem J. 1977;164(3):509-19.

38.

Solini A, Usuelli V, Fiorina P. The dark side of extracellular ATP in kidney diseases. J Am Soc Nephrol. 2015;26(5):1007-16.

39.

Tang LX, Wang B, Wu ZK. Aerobic Exercise Training Alleviates Renal Injury by Interfering with Mitochondrial Function in Type-1 Diabetic Mice. Med Sci Monit. 2018;24:9081-9.

40.

Yuan S, Liu X, Zhu X, et al. The Role of TLR4 on PGC-1alpha-Mediated Oxidative Stress in Tubular Cell in Diabetic Kidney Disease. Oxid Med Cell Longev. 2018;2018:6296802.

41.

Kim J, Yang G, Kim Y, et al. AMPK activators: mechanisms of action and physiological activities. Exp Mol Med. 2016;48:e224.

42.

Kahn BB, Alquier T, Carling D, et al. AMP-activated protein kinase: ancient energy gauge provides clues to modern understanding of metabolism. Cell Metab. 2005;1(1):15-25.

43.

Dolinsky VW, Dyck JR. Role of AMP-activated protein kinase in healthy and diseased hearts. Am J Physiol Heart Circ Physiol. 2006;291(6):H2557-69.

44.

Kim D, Lee JE, Jung YJ, et al. Metformin decreases high-fat diet-induced renal injury by regulating the expression of adipokines and the renal AMP-activated protein kinase/acetyl-CoA carboxylase pathway in mice. Int J Mol Med. 2013;32(6):1293-302.

45.

Yap F, Craddock L, Yang J. Mechanism of AMPK suppression of LXR-dependent Srebp-1c transcription. Int J Biol Sci. 2011;7(5):645-50.

46.

Hardie DG. AMPK: a target for drugs and natural products with effects on both diabetes and cancer. Diabetes. 2013;62(7):2164-72.

47.

Kim MY, Lim JH, Youn HH, et al. Resveratrol prevents renal lipotoxicity and inhibits mesangial cell glucotoxicity in a manner dependent on the AMPK-SIRT1-PGC1alpha axis in db/db mice. Diabetologia. 2013;56(1):204-17.

48.

Yang H, Feng A, Lin S, et al. Fibroblast growth factor-21 prevents diabetic cardiomyopathy via AMPK-mediated antioxidation and lipid-lowering effects in the heart. Cell Death Dis. 2018;9(2):227.

49.

Hong YA, Lim JH, Kim MY, et al. Fenofibrate improves renal lipotoxicity through activation of AMPK-PGC-1alpha in db/db mice. PLoS One. 2014;9(5):e96147.

50.

Rui L. Energy metabolism in the liver. Compr Physiol. 2014;4(1):177-97.

51.

Kersten S. Mechanisms of nutritional and hormonal regulation of lipogenesis. EMBO Rep. 2001;2(4):282-6.

52.

Grove KJ, Voziyan PA, Spraggins JM, et al. Diabetic nephropathy induces alterations in the glomerular and tubule lipid profiles. J Lipid Res. 2014;55(7):1375-85.

53.

Miserez AR, Muller PY, Barella L, et al. Sterol-regulatory element-binding protein (SREBP)-2 contributes to polygenic hypercholesterolaemia. Atherosclerosis. 2002;164(1):15-26.

54.

Jiang T, Wang Z, Proctor G, et al. Diet-induced obesity in C57BL/6J mice causes increased renal lipid accumulation and glomerulosclerosis via a sterol regulatory element-binding protein-1c-dependent pathway. J Biol Chem. 2005;280(37):32317-25.

55.

Bobulescu IA. Renal lipid metabolism and lipotoxicity. Curr Opin Nephrol Hypertens. 2010;19(4):393-402.

56.

Koh ES, Lim JH, Kim MY, et al. Anthocyanin-rich Seoritae extract ameliorates renal lipotoxicity via activation of AMP-activated protein kinase in diabetic mice. J Transl Med. 2015;13:203.

57.

Proctor G, Jiang T, Iwahashi M, et al. Regulation of renal fatty acid and cholesterol metabolism, inflammation, and fibrosis in Akita and OVE26 mice with type 1 diabetes. Diabetes. 2006;55(9):2502-9.

58.

Kim HW, Lee JE, Cha JJ, et al. Fibroblast growth factor 21 improves insulin resistance and ameliorates renal injury in db/db mice. Endocrinology. 2013;154(9):3366-76.

59.

Tsun JG, Yung S, Chau MK, et al. Cellular cholesterol transport proteins in diabetic nephropathy. PLoS One. 2014;9(9):e105787.

60.

Tanaka Y, Kume S, Araki S, et al. Fenofibrate, a PPARalpha agonist, has renoprotective effects in mice by enhancing renal lipolysis. Kidney Int. 2011;79(8):871-82.

61.

Park HS, Lim JH, Kim MY, et al. Resveratrol increases AdipoR1 and AdipoR2 expression in type 2 diabetic nephropathy. J Transl Med. 2016;14(1):176.

62.

Park CH, Noh JS, Fujii H, et al. Oligonol, a low-molecular-weight polyphenol derived from lychee fruit, attenuates gluco-lipotoxicity-mediated renal disorder in type 2 diabetic db/db mice. Drug Discov Ther. 2015;9(1):13-22.

63.

Wang XX, Levi J, Luo Y, et al. SGLT2 Protein Expression Is Increased in Human Diabetic Nephropathy: SGLT2 protein inhibition decreased renal lipid accumulation, inflammation, and the development of nephropathy in diabetic mice. J Biol Chem. 2017;292(13):5335-48.

64.

Kim BH, Lee ES, Choi R, et al. Protective Effects of Curcumin on Renal Oxidative Stress and Lipid Metabolism in a Rat Model of Type 2 Diabetic Nephropathy. Yonsei Med J. 2016;57(3):664-73.

65.

Falkevall A, Mehlem A, Palombo I, et al. Reducing VEGF-B Signaling Ameliorates Renal Lipotoxicity and Protects against Diabetic Kidney Disease. Cell Metab. 2017;25(3):71326.

66.

Dominguez J, Wu P, Packer CS, et al. Lipotoxic and inflammatory phenotypes in rats with uncontrolled metabolic syndrome and nephropathy. Am J Physiol Renal Physiol. 2007;293(3):F670-9.

67.

Muthusamy K, Halbert G, Roberts F. Immunohistochemical staining for adipophilin, perilipin and TIP47. J Clin Pathol. 2006;59(11):1166-70.

68.

Imamura M, Inoguchi T, Ikuyama S, et al. ADRP stimulates lipid accumulation and lipid droplet formation in murine fibroblasts. Am J Physiol Endocrinol Metab. 2002;283(4):E775-83.

69.

Mori J, Patel VB, Ramprasath T, et al. Angiotensin 1-7 mediates renoprotection against diabetic nephropathy by reducing oxidative stress, inflammation, and lipotoxicity. Am J Physiol Renal Physiol. 2014;306(8):F812-21.

70.

Zhang C, Shao M, Yang H, et al. Attenuation of hyperlipidemia- and diabetes-induced early-stage apoptosis and late-stage renal dysfunction via administration of fibroblast growth factor-21 is associated with suppression of renal inflammation. PLoS One. 2013;8(12):e82275.

71.

Iwai T, Kume S, Chin-Kanasaki M, et al. Stearoyl-CoA Desaturase-1 Protects Cells against Lipotoxicity-Mediated Apoptosis in Proximal Tubular Cells. Int J Mol Sci. 2016;17(11).

72.

Li L, Yoshitomi H, Wei Y, et al. Tang-Nai-Kang alleviates pre-diabetes and metabolic disorders and induces a gene expression switch toward fatty acid oxidation in SHR.CgLeprcp/NDmcr rats. PLoS One. 2015;10(4):e0122024.

73.

Sas KM, Kayampilly P, Byun J, et al. Tissue-specific metabolic reprogramming drives nutrient flux in diabetic complications. JCI Insight. 2016;1(15):e86976.

74.

Jeon SM. Regulation and function of AMPK in physiology and diseases. Exp Mol Med. 2016;48(7):e245.

75.

Zambo V, Simon-Szabo L, Szelenyi P, et al. Lipotoxicity in the liver. World J Hepatol. 2013;5(10):550-7.

76.

Izquierdo-Lahuerta A, Martinez-Garcia C, Medina-Gomez G. Lipotoxicity as a trigger factor of renal disease. J Nephrol. 2016;29(5):603-10.

77.

Cheng Y, Zhang J, Guo W, et al. Up-regulation of Nrf2 is involved in FGF21-mediated fenofibrate protection against type 1 diabetic nephropathy. Free Radic Biol Med. 2016;93:94-109.

78.

Esfahani M, Movahedian A, Baranchi M, et al. Adiponectin: an adipokine with protective features against metabolic syndrome. Iran J Basic Med Sci. 2015;18(5):430-42.

79.

Choi SR, Lim JH, Kim MY, et al. Adiponectin receptor agonist AdipoRon decreased ceramide, and lipotoxicity, and ameliorated diabetic nephropathy. Metabolism. 2018;85:348-60.

80.

Robinson K, Prins J, Venkatesh B. Clinical review: adiponectin biology and its role in inflammation and critical illness. Crit Care. 2011;15(2):221.

81.

Ouedraogo R, Wu X, Xu SQ, et al. Adiponectin suppression of high-glucose-induced reactive oxygen species in vascular endothelial cells: evidence for involvement of a cAMP signaling pathway. Diabetes. 2006;55(6):1840-6.

82.

Plant S, Shand B, Elder P, et al. Adiponectin attenuates endothelial dysfunction induced by oxidised low-density lipoproteins. Diab Vasc Dis Res. 2008;5(2):102-8.

83.

Kim Y, Lim JH, Kim MY, et al. The Adiponectin Receptor Agonist AdipoRon Ameliorates Diabetic Nephropathy in a Model of Type 2 Diabetes. J Am Soc Nephrol. 2018;29(4):1108-27.

84.

Bjornstad P, Pyle L, Kinney GL, et al. Adiponectin is associated with early diabetic kidney disease in adults with type 1 diabetes: A Coronary Artery Calcification in Type 1 Diabetes (CACTI) Study. J Diabetes Complications. 2017;31(2):369-74.

85.

Tanaka Y, Kume S, Araki H, et al. 1-Methylnicotinamide ameliorates lipotoxicity-induced oxidative stress and cell death in kidney proximal tubular cells. Free Radic Biol Med. 2015;89:831-41.

86.

Cai Y, Lydic TA, Turkette T, et al. Impact of alogliptin and pioglitazone on lipid metabolism in islets of prediabetic and diabetic Zucker Diabetic Fatty rats. Biochem Pharmacol. 2015;95(1):46-57.

87.

Acosta-Montano P, Garcia-Gonzalez V. Effects of Dietary Fatty Acids in Pancreatic Beta Cell Metabolism, Implications in Homeostasis. Nutrients. 2018;10(4).

88.

Kim JA, Wei Y, Sowers JR. Role of mitochondrial dysfunction in insulin resistance. Circ Res. 2008;102(4):401-14.

89.

Okabe K, Yaku K, Tobe K, et al. Implications of altered NAD metabolism in metabolic disorders. J Biomed Sci. 2019;26(1):34.

90.

Wang Q, Liu S, Zhai A, et al. AMPK-Mediated Regulation of Lipid Metabolism by Phosphorylation. Biol Pharm Bull. 2018;41(7):985-93.

91.

Babashamsi MM, Koukhaloo SZ, Halalkhor S, et al. ABCA1 and metabolic syndrome; a review of the ABCA1 role in HDL-VLDL production, insulin-glucose homeostasis, inflammation and obesity. Diabetes Metab Syndr. 2019;13(2):1529-34.

92.

Engin AB. What Is Lipotoxicity? Adv Exp Med Biol. 2017;960:197-220.

93.

Cano-Europa E, Ortiz-Butron R, Camargo EM, et al. A canola oil-supplemented diet prevents type I diabetes-caused lipotoxicity and renal dysfunction in a rat model. J Med Food. 2016;19(11):1041-7.

Figure 1. Schematic overview of interaction of metabolic and hemodynamic factors in the pathogenesis of diabetic nephropathy. Abbreviations: AGEs, advanced glycation end products; GSH, reduced glutathione; PKC, protein kinase C; RAAS, renin angiotensin aldosterone system; TGF-β, transforming growth factor beta; VEGF, vascular endothelial growth factor.

Figure 2. Lipid metabolism in renal proximal tubule cells. Fatty acids (FA) that enter renal proximal tubule cells both by endocytosis and transport via CD36 are converted to fatty acylCoA by acyl-CoA synthetase (ACS). Fatty acyl-CoA is transported into mitochondria via carnitine palmitoyltransferase 1 (CPT1). β-oxidation converts fatty acyl-CoA to acetyl-CoA and subsequently ATP is generated by the tricarboxylic acid (TCA) or Kreb’s cycle and the electron transport chain. Excess acetyl-CoA can be converted to malonyl-CoA by acetyl-CoA carboxylase (ACC). An increase in malonyl-CoA not only inhibits CPT1 function leading to further reduction in β-oxidation, but also activates fatty acid synthase (FAS) activity to convert fatty acyl-CoA into palmitate which feeds into the fatty acid pool and then stored inside the cell as triglycerides. In diabetes condition (presented by red arrow) which lead to an increase in fatty acid uptake into renal cells and the activation of the oxidation of fatty acids. However, a long-term increase in βoxidation with impairment of mitochondria function can in turn lead to incomplete β-oxidation leading to a decrease in ATP production. Along with the decrease in AMPK activity which can suppress fatty acid synthesis, excess acetyl-CoA can be converted to fatty acyl-CoA that in turn enters the fatty acid pool and increases the triglycerides stored inside the cell, lipid accumulation.

Figure 3. The role of AMPK in the regulation of lipid metabolism. Under ATP depletion, AMPK increases fatty acid oxidation through inhibiting ACC activity leading to decreased malonyl-CoA level. Moreover, AMPK has been found to activate PPARα to stimulate fatty acid oxidation via increased PGC-1α activity. AMPK also promotes cellular fatty acid utilization by inhibiting both mTORs-SREBP1 and HMG-CoA reductase which related to lipid synthesis and cholesterol synthesis and also deactivates phosphoenolpyruvate carboxykinase (PEPCK) enzyme that controls gluconeogenic pathway.

Table 1. Renal lipotoxicity in in vitro studies.

Metabolic parameter N/A

Results Renal lipid accumulation N/A

Mesangial cell + 30 mmol/l D-glucose

N/A

N/A

HGECs + 40 mmol/l D-glucose

N/A

N/A

Diabetic Model Mesangial cell + 30 mmol/l D-glucose

HRGECs+ 25 mmol/L D-glucose +400 µg/ml Cholesterol

↑Total Cholesterol ↑Free Cholesterol ↑Cholesterol Ester ↑CE/TC (%)

↑ Renal Cholesterol content

Renal lipid metabolism FA Oxidation: ↓PPARα ↓SIRT1 ↓PGC-1α ↓ERR-1α ↓CPT-1A FA synthesis: ↑SREBP1 FA Oxidation: ↓PPARα ↓PGC-1α ↓ERR-1α FA synthesis: ↑SREBP ↑ChREBP FA oxidation: ↓PPARα ↓PPAR ↓PGC-1α ↓ERR-1α ↓p-ACC Cholesterol efflux: ↓ABCA1

Other findings

Interpretation

Ref.

Hyperglycemia, through decreased FA oxidation and increased FA synthesis, induced renal cell lipotoxicity associated with increased oxidative stress and apoptosis.

47

↓p-/Total Thr AMPK ↑PI3K 473 ↑p-Ser Akt 253 ↑p-/Total Ser FoxO3a Oxidative stress: ↑isoprostane Apoptosis:↑TUNEL ↓p-AMPK Oxidative stress: ↓SOD1, ↓SOD2 Apoptosis: ↓Bcl-2/Bax

Hyperglycemia, through decreased FA oxidation and increased FA synthesis, induced renal cell lipotoxicity associated with increased oxidative stress and apoptosis.

49

Hyperglycemia, through decreased FA oxidation, induced renal cell lipotoxicity associated with increased oxidative stress and apoptosis.

56

↓cAMP activity ↓p-PI3K ↓p-Akt ↑p-ERK1/2 Inflammation: ↑TNF-α, ↑IL-6

Decrease in cholesterol efflux caused hyperlipidemic conditions, intracellular lipid accumulation and inflammation.

18

↓p-/Total AMPK ↑PI3K ↑p-/total Akt ↑p-/total FoxO3a Oxidative stress: ↓SOD1, ↓SOD2 Apoptosis: ↑TUNEL 173

Abbreviations: FA, Fatty acids; PPARα, Peroxisome proliferator-activated receptor alpha; PPAR, Peroxisome proliferator-activated receptor gamma; PGC-1α, Peroxisome proliferator-activated receptor gamma coactivator 1-alpha; ERR-1α, Estrogen-related receptor 1 alpha; SREBP, Sterol regulatory element-binding proteins; ChREBP, Carbohydrate-responsive element-binding protein; AMPK, 5' AMP-activated protein kinase; PI3K, Phosphoinositide 3-kinase; Akt, Protein Kinase B; FoxO3a, Forkhead box O3; TUNEL, Terminal deoxynucleotidyltransferase dUTP nick end labeling; SIRT1, NAD-dependent deacetylase sirtuin-1; CPT-1A, Carnitine palmitoyltransferase 1A; SOD1, Superoxide dismutase 1; SOD2, Superoxide dismutase 2; ACC, Acetyl-CoA carboxylase; Bcl-2, B-cell lymphoma 2; ABCA1, ATP-binding cassette transporter 1;ERK1/2, Extracellular signalregulated protein kinases 1 and 2; TNF-α, Tumor necrosis factor alpha; IL-6, Interleukin 6; HGECs, Human glomerular epithelial cells; HRGECs, Human renal glomerular epithelial cells

Table 2. Renal lipotoxicity in in vivo studies in T1DM.

Diabetic Model C57BL/6J -/ApoE mice + HFD + 4 doses of STZ (55 mg/kg BW)

Metabolic parameter ↑FBG ↑HbA1c ↑Cholesterol ↑LDL ↓HDL ↔TG

Kidney function ↑KW/BW ↑Albuminuria ↑UAE ↔BUN ↑Cr. clearance

Results Renal lipid accumulation Oil Red O, CD31 staining: ↑ lipid accumulation ↑Cholesterol content in renal cortex

Akita mice

↑TG ↑Cholesterol

↑UACR Podocyte marker : ↓Synaptopodin ↓Podocin

N/A

db/db mice with insulin deficient diabetes

↑FBG ↑HbA1c ↓Serum insulin

↔KW/BW ↑Urinary albumin Podocyte marker : ↓Synaptopodin ↓Nephrin ↓Podocyte number

Oil red O: ↑Neutral lipid accumulation in glomerular and tubular cell

C57BL/6 mice +5 days of STZ (50 mg/kg BW)

↑FBG ↑TG ↑NEFA

↑Kidney hypertrophy ↑Albuminuria ↑Urine volume ↑UAE Podocyte marker : ↓Synaptopodin

Oil Red O: ↑Neutral lipid in glomerulus

Renal lipid metabolism Cholesterol efflux: ↓ABCA1

FA synthesis: ↑SREBP-1 ↑SCD ↓FXR Cholesterol synthesis: ↑SREBP-2 ↓ABCA1 ↓LXR-α, LXR-β FA synthesis: ↑ChREBP ↑LPK ↑SCD1 ↑DGAT

N/A

Other findings Inflammation: ↑C-reactive protein ↑TNF-α, ↑IL-6 Filipin staining: ↑Cholesterol accumulation H&E,PAS staining: ↑Glomerular hypertrophy, ↑GBM thickness, ↑Mesangial expansion Inflammation: ↑VEGF, ↑TNF-α Fibrosis: ↑TGF-β, ↑PAI-1 PAS staining: ↑Mesangial expansion, ↑Matrix accumulation, ↑Tubulointerstitial fibrosis Inflammation : ↑p65NF-κB ↔MCP-1↓ACE,ACE2 ↓AT1R,AT2R ↓Angiotensinogen PAS staining: ↑Mesangial expansion Immunofluorescence: ↑Fibronectin ↑Type IV collagen Glucose metabolism: ↔PEPCK,G6Pc ↑VEGF-R1 PAS staining: ↑Mesangial expansion

Interpretation

Ref.

Hyperglycemia and dyslipidemia led to increased lipid accumulation in renal cortex resulting in renal inflammation and dysfunction.

18

Diabetic condition induced an increase in FA and cholesterol synthesis and a decrease in cholesterol efflux together with renal inflammation and fibrosis.

57

In T1D, increased FA synthesis and lipotoxicity resulted in renal dysfunction and inflammation without affecting renal gluconeogenic genes.

63

Hyperglycemia led to increased glomerular lipid accumulation resulting in renal dysfunction.

65

Diabetic Model FGF21-KO mice + STZ (200 mg/kg BW)

Metabolic parameter ↑TG

Kidney function ↑KW/TL ↑PCR ↑ACR ↑BUN

Results Renal lipid accumulation Oil Red O : ↑Lipid accumulation ↑Renal TG content

Renal lipid metabolism N/A

C57BL/6J mice + STZ (150 mg/kg BW)

↓BW ↑FBG ↑TG ↑Cholesterol

↑KW/TL ↑UACR

N/A

N/A

Sprague-Dawley + STZ (50 mg/kg BW)

↓BW, ↓Mesenteric fat weight ↓Retroperito neal fat weight ↓Epididymal fat pad weight ↑FFA ↑Cholesterol ↑TG ↑FBG

↑Urine flow ↑Osolarity clearance ↓Cr. clearance

N/A

N/A

Other findings Oxidative stress: ↑3-NT, ↑4-HNE Inflammation: ↑ICAM-1 ↑TNF-α, ↑PAI-1 Fibrosis: ↑CTGF Apoptosis: ↑TUNEL Oxidative stress: ↑3-NT, ↑4-HNE, ↑Renal MDA ↑Nrf-2, ↑NQO-1, ↑HO-1 Inflammation: ↑TNF-α, ↑PAI-1 Fibrosis: ↑CTGF, TGF-β1 H&E staining: ↑Glomerular enlargement ↑GBM thickening ↑Mesangial expansion PAS staining: ↑Renal glycogen accumulation Masson’s trichrome stain: ↑Fibrosis Apoptosis: ↑TUNEL, ↑Bax/Bcl-2 ↑Cleaved caspase 3 Oxidative stress: ↑ROS quantification ↑Lipid peroxidation

Interpretation

Ref.

Kidneys of diabetic mice showed lipid accumulation with oxidative stress, inflammation, fibrosis and apoptosis.

70

STZ-induced T1DN showed renal dysfunction and morphological changes with increased oxidative stress, inflammation, fibrosis, and apoptosis.

77

STZ-induced T1D caused dyslipidemia resulting in renal dysfunction.

93

Abbreviations: HFD, High fat diet; STZ, Streptozotocin; FA, Fatty acids; FFA, Free fatty acids; NEFA , Non-esterified fatty acids; BW, Body weight; FBG, Fasting blood glucose; BUN, Blood urea nitrogen; Cr.clearance, Creatinine clearance; UAE, Urine albumin excretion; UACR, Urine albumin/creatinine ratio; PCR, Protein to creatinine ratio; HbA1c, Hemoglobin A1c; KW/BW, Kidney weight/Body weight; KW/TL, Kidney weight/Tibia length; PPARα, Peroxisome proliferator-activated receptor alpha; PPAR, Peroxisome proliferator-activated receptor gamma; PGC-1α, Peroxisome proliferator-activated receptor gamma coactivator 1-alpha; ERR-1α, Estrogen-related receptor 1 alpha; SREBP, Sterol regulatory element-binding proteins;ChREBP, Carbohydrate-responsive element-binding protein; AMPK, 5' AMP-activated protein kinase; PI3K, Phosphoinositide 3-kinase; Akt, Protein Kinase B; FoxO1, Forkhead box O1; FoxO3a, Forkhead box O3; TUNEL, Terminal deoxynucleotideacyltransferase dUTP nick end labeling; SIRT1, NAD-dependent deacetylase sirtuin-1; CPT-1A, Carnitine palmitoyltransferase 1A; SOD1, Superoxide dismutase 1; SOD2, Superoxide dismutase 2; ACC, Acetyl-CoA carboxylase; Bcl-2, B-cell lymphoma 2; AdipoR1, Adiponectin receptor 1; AdipoR2, Adiponectin receptor 2; ApoAI, Apolipoprotein A1; CE/TC, Cholesterol ester/Total cholesterol; ABCA1, ATP-binding cassette transporter 1; ERK1/2, Extracellular signal-regulated protein kinases 1 and 2; TNF-α, Tumor necrosis factor alpha; IL-6, Interleukin 6; LPK, L-pyruvate kinase; SCD1, Stearoyl-CoA desaturase 1; DGAT, Diacylglycerolacyltransferases; NF-κB, Nuclear factor-kappa B;MCP-1, Monocyte chemotactic protein 1; ACE, Angiotensin converting enzyme; ACE2, Angiotensin converting enzyme 2; AT1R, Angiotensin II type 1 receptor; AT2R, Angiotensin II type 2 receptor; TG, Triglyceride; PEPCK, Phosphoenolpyruvate carboxykinase; G6Pc, Glucose-6-phosphatase catalytic-subunit; FXR, Farnexoid X receptor; LXR-α, Liver X receptor alpha; LXR-β, Liver X receptor beta; VEGF, Vascular endothelial growth factor; TGF-β, Transforming growth factor beta; PAI-1, Plasminogen activator inhibitor-1; 3-NT, 3-Nitrotyrosine; 4-HNE, 4-Hydroxynonenal; GMB, Glomerular basement membrane; ROS, Reactive oxygen species.

Table 3. Renal lipotoxicity in in vivo studies in T2DM. Results Diabetic Model C57BLKS/J mice

Metabolic parameter ↑BW ↑FBG ↑HbA1c

Kidney function ↑KW ↑Albumin ↑Cr.clearance ↔Creatinine

Renal lipid accumulation ↔Renal total cholesterol ↑Renal NEFA ↑Renal TG

Renal lipid metabolism FA Oxidation: ↓SIRT1 ↓PGC-1α ↓PPARα ↓ERR-1α FA synthesis: ↑SREBP1

C57BLKS/J mice

↑BW ↑FBG ↑HbA1c ↑Cholesterol ↑TG ↑FFA

↑KW ↑Albuminuria ↑Urine volume ↑Cr.clearance ↔Creatinine

↔Intrarenal total cholesterol ↔Intrarenal TG ↑Intrarenal FFA

FA Oxidation: ↓PPARα ↓PGC-1α ↓ERR-1α ↓pACC FA synthesis: ↑SREBP1 ↑ChREBP-1

C57BLKS/J db/db mice

↑BW ↑FBG ↑HbA1c ↑NEFA ↑TG ↑Cholesterol

↑KW ↑Albuminuria ↑Urine volume

↑Renal FFA ↑Renal TG ↑Renal Cholesterol

FA Oxidation: ↓PPARα ↓PPAR ↓pACC/ACC FA synthesis: ↑SREBP

Other findings

Interpretation

Ref.

↓p-/total AMPK ↑PI3K, PI3K activity 473 ↑p-Akt Ser ↑p-/total FoxO3a Oxidative stress: ↓SOD1, SOD2 ↑Serum 8-OHdG ↑Urine 8-OHdG ↑Urine isoprostane Fibrosis: ↑TGF-β1 ↑Collagen Type IV PAS staining: ↑Mesangial area Apoptosis: ↑TUNEL, ↑Bax/Bcl-2 ↑Cleaved caspase 3 173 ↓p-/Total The AMPK ↑PI3K, PI3K activity 473 ↑p-Ser Akt 253 ↑p-/Total Ser FoxO3a 256 ↔p-/Total Ser FoxO1 Oxidative stress: ↓SOD1, ↔SOD2 ↑Urine 8-isoprostane Fibrosis:↑TGF-β1 ↑Collagen Type IV,↑F4/80 PAS staining : ↑Fractional mesangial area Apoptosis: ↓Bcl-2/Bax, ↑TUNEL ↓pAMPK/AMPK Oxidative stress: ↑Urine isoprostane Fibrosis: ↑TGF-β1 ↑Collagen type IV Apoptosis: ↓Bcl2/Bax, ↑TUNEL PAS staining:↑Mesangial area

T2DM induced renal lipotoxicity through decreasing FA oxidation and increasing FA synthesis which induced oxidative stress, fibrosis, and apoptosis.

47

T2DM induced renal lipotoxicity through decreasing FA oxidation and increasing FA synthesis which increased oxidative stress, fibrosis, and apoptosis.

49

Diabetic mice showed increased FA synthesis and decreased FA oxidation which impaired kidney function via increased oxidative stress, fibrosis, and apoptosis.

56

Diabetic Model

C57BL/6 mice+ HFD induced diabetes

Diabetic db/db mice

C57BLKS/J mice

Results Metabolic parameter ↑BW ↑FBG ↑Cholesterol ↑FFA ↑HbA1c ↑Serum insulin ↔TG

↑BW ↑FBG ↑HbA1c ↑FGF21 ↑Serum insulin ↑HOMA-IR ↑Insulin resistance ↑Cholesterol ↑TG ↑LDL ↔HDL ↓Adiponectin ↑BW ↑FBG ↑HbA1c ↓Adiponectin

Renal lipid accumulation Oil Red O: ↑Neutral lipid accumulation ↑Renal TG content

Renal lipid metabolism FA oxidation: ↑ACC ↓ACO ↓CPT-1 ↔PPARα ↔PPAR FA synthesis: ↑SREBP-1

↔KW/BW ↔Creatinine ↑Urine volume ↑UAE

↑Cholesterol content ↑TG content ↑LPO content

Cholesterol synthesis: ↓ABCA ↑SREBP1c ↑p-ERK

↔KW ↔BUN ↔Creatinine ↑Urine volume ↑Albuminuria

↑Renal NEFA ↑Renal TG ↔Renal total cholesterol Oil Red O: ↑Neutral lipid accumulation

FA Oxidation: ↓SIRT1 ↓PPARα ↓PGC-1α ↓ERR-1α ↓p-/total ACC FA synthesis: ↑SREBP-1c

Kidney function ↑UAE

Other findings Oxidative stress: ↑4-HNE ↑Renal MDA ↓Catalase ↓SOD Fibrosis: ↑Fibronectin ↑Collagen type I,IV ↑PAI-1 PAS staining: ↑Mesangial area ↑ Glomerular volume Oxidative stress: ↑Serum 8-iso prostane ↑Urine 8-iso prostane Inflammation: ↑p65NF-ĸB Fibrosis: ↑Collagen type IV ↑TGF-β1 ↑Fibronectin, ↑Laminin PAS staining: ↑Glomerular tuft hypertrophy ↑Mesangial expansion 256

↑p-FoxO1(Ser ) 253 ↑p-FoxO3a(Ser ) ↓AdipoR1, R2 172 ↓p-AMPK(Thr ) Oxidative stress: ↑Urine 8-OHdG ↑Renal 8-OHdG ↑Urine isoprostane Inflammation: ↓p-/total eNOS Fibrosis: ↑TGF-β, ↑Collagen type IV PAS staining: ↑Mesangial area Apoptosis: ↑TUNEL ↓Bcl-2/Bax

Interpretation

Ref.

HFD-induced alteration of lipid metabolism led to renal lipid accumulation, oxidative stress and fibrosis.

60

Increased synthesis and accumulation of cholesterol in the kidney of T2DM mice impaired renal function, induced oxidative stress, inflammation and fibrosis.

58

Increased FA synthesis and decreased FA oxidation in diabetes caused lipid accumulation in the kidney and development of renal dysfunction via oxidative stress, inflammation, fibrosis, and apoptosis.

61

Results Diabetic Model C57BLKS/6 mice (db/db)

Otsuka-LongEvansTokushima Fatty (OLETF)

HFD induced diabetes

C57BKS/ db Lepr mice

Metabolic parameter ↑BW ↑Glucose ↑Insulin ↑Leptin ↑TG ↑Cholesterol ↑NEFA ↓Adiponectin

Kidney function ↑BUN ↑Creatinine

↑FBG ↑Cholesterol ↑TG ↑FFA ↓Serum insulin ↓HOMA-β ↓Adiponectin ↔HOMA-IR ↔Epididymal fat weight ↑Postprandial blood glucose ↑Glucose tolerance

↑KW/BW ↑UACR ↑Urinary albumin

↑FBG ↑Ketone bodies ↑TG ↑LDL ↑NEFA ↓HDL ↔Glucose tolerance

↑ACR ↑Cr.clearance Podocyte marker: ↓Podocin ↓Synaptopodin

↑ACR Podocyte marker : ↓Synaptopodin

Renal lipid accumulation N/A

↔Renal TG

N/A

Oil Red O: ↑Neutral lipid ↑Lipid droplet

Renal lipid metabolism FA Oxidation: ↓PPARα FA synthesis: ↑SREBP1 ↑SREBP2

FA synthesis: ↑SREBP-1 ↑SREBP-2 ↑ADRP FA oxidation: ↓pACC/ACC

↑Adipophilin

↑Adipophilin ↑Fatp4

Other findings Oxidative stress: ↑TBARs ↔GSH ,↑GSSG phox ↑p22 , ↑NOX-4 ↑RAGE ↑JNK, p-JNK, c-Jun H&E staining: ↑Glomerular enlargement ↑Renal injury Inflammation: ↑TNF-α Apoptosis: ↑Bax ↑Cytochrome C ↑Caspase 3 ↔p-AMPK/AMPK Oxidative stress: ↓Urine SOD ↓Nrf2/keap1, ↓HO-1 ↑Urine MDA H&E staining: ↑Glomerular hypertrophy EM:↑GBM thickness ↑Open slit pore ↑Glomerular volume PAS staining: ↑Mesangial expansion

↑VEGF-B ↑Collagen type IV PAS staining: ↑Mesangial expansion TEM: ↑GBM thickeness ↓Podocyte slit density

Interpretation

Ref.

T2DM mice showed increased FA synthesis and decreased FA oxidation which impaired kidney function via increased oxidative stress, inflammation, and apoptosis.

62

Renal lipid accumulation altered renal lipid metabolism leading to renal dysfunction.

64

HFD increased renal lipid droplets and caused mesangial expansion and renal dysfunction Diabetic mice showed dyslipidemia that accelerated lipid accumulation in the glomeruli and kidney dysfunction.

65

65

Diabetic Model Zucker fatty diabetic rats

C57BL/6Jlepr/lepr (db/db) mice

C57BL/6 mice + HFD

Results Metabolic parameter ↑BW ↑FBG ↑Cholesterol ↑TG Systemic inflammation: ↑PMN ↑Monocyte ↑CRP ↑IL-1 ↓Lymphocyte ↔Adiponectin ↔MCP-1 ↔IL-8 ↑BW ↑Random blood glucose ↑TAG

↑BW ↑FBG

Kidney function ↑KW ↑KW/BW ↑Albuminuria ↑Cr.clearance ↔ Creatinine

↑KW ↑KW/TL ↔Creatinine ↑Urinary albumin

↑Urine albumin ↔KW/BW

Renal lipid accumulation Oil Red O: ↑Renal tubules laden with lipid ↑Renal TG accumulation

Renal lipid metabolism FA synthesis: ↑ADRP FA Oxidation: ↔FAO activity ↔LCAD activity ↑pACC

Oil Red O: ↑Renal lipid accumulation ↑Perirenal adipose tissue ↑Renal TG

FA Oxidation: ↓SIRT1 ↓PPARα

N/A

FA synthesis: ↓SCD1, ↔SCD2 ↓ADRP ↔FAS ↔DGAT1, ↔DGAT2 FA oxidation: ↓ACO ↔CPT1 ↔ACC

Other findings

Interpretation

Ref.

↑pAMPK ↔AMPK Oxidative stress: ↑4-HNE Renal inflammation: ↑Macrophage ↑mRNA of IL-1β ↑IL-6 ↑TGF-β1 Receptor ↔mRNA of TGF-β1 ↔TNF-α ↔TNF-α Receptor

Increased FA synthesis in diabetic nephropathy resulted in renal lipid accumulation, systemic and renal inflammation and renal dysfunction.

66

↑p-/total Erk1/2 ↑ACE2 ↑Renal ACE2 activity Oxidative stress: ↑NADPH oxidase ↑DHE staining ↑Renal nitrotyrosine level PASstaining: ↑Mesangial expansion Masson’s trichrome,H&E staining: ↑Macrophage in perirenal adipose tissue Apoptosis : ↑TUNEL ↑Cleaved caspase 3

Db/db mice with T2DN had decreased FA oxidation resulting in renal lipid accumulation, renal dysfunction, oxidative stress and inflammation.

69

HFD consumption reduced SCD1 activity which led to decrease in lipid droplet synthesis and apoptosis.

71

Diabetic Model C57BL/6J mice+ FFA-BSA (0.36g/30gBW)

Results Metabolic parameter N/A

Kidney function N/A

Renal lipid accumulation N/A

Renal lipid metabolism NAD metabolism : ↑1-MNA ↔NAD

Other findings

Interpretation

Ref.

Inflammation : ↑MCP-1 ↑PAI-1 ↑4-HNE Fibrosis:↑Fibronectin H&E staining: ↑Destruction of nuclei and cytoplasm ↑Eosinophilic ↑Necrotic debris with in tubule lumen Apoptosis: ↑TUNEL ↑Cleaved caspase3

NAD metabolism in high-fat condition involved in the induction of renal morphological changes, inflammation, fibrosis and apoptosis.

85

Abbreviations: HFD, High fat diet; FFA, Free fatty acids; BSA, Bovine serum albumin; BW, Body weight; FBG, Fasting blood glucose; FA, Fatty acids; FFA, Free fatty acids; HbA1c, Hemoglobin A1c; TG, Triglycerides; HOMA-β, Homeostatic model assessment of β-cell function; HOMA-IR, Homeostatic model assessment of insulin resistance;FGF21, Fibroblast growth factor 21; LDL, Low density lipoprotein; HDL, High density lipoprotein; NEFA, Non-essential fatty acids; TAG, Triacylglycerol; PMN, Polymorphonuclear leukocytes; CRP, Creactive protein; IL-1, Interleukin-1; IL-8, Interleukin-8; MCP-1, Monocyte chemotactic protein 1; UAE, Urine albumin excretion; KW, Kidney weight; KW/BW, Kidney weight/Body weight ratio; KW/TL, Kidney weight/Tibia length; UACR, Urine albumin to creatinine ratio; ACR, Albumin to creatinine ratio; BUN, Blood urea nitrogen; Cr.clearance, Creatinine clearance; LPO, Lipid peroxidation; ACC, Acetyl-CoA carboxylase; ACO, Acyl-coenzyme A oxidase; CPT-1, Carnitine palmitoyltransferase 1; PPARα, Peroxisome proliferatoractivated receptor alpha; PPAR, Peroxisome proliferator-activated receptor gamma; SREBP-1, Sterol regulatory element-binding protein 1; SREBP-2, Sterol regulatory elementbinding protein 2; ADRP, Adipose differentiation related protein; ABCA, ATP-binding cassette transporter A; p-ERK, Phosphorylate extracellular signal-regulated protein kinases; PGC-1α, Peroxisome proliferator-activated receptor gamma coactivator 1-alpha; FoxO1, Forkhead box O1; FoxO3a, Forkhead box O3; ERR-1α, Estrogen-related receptor 1 alpha; ChREBP-1, Carbohydrate-responsive element-binding protein 1; SIRT1, NAD-dependent deacetylase sirtuin-1; SCD1, Stearoyl-CoA desaturase-1; SCD2,Stearoyl-CoA desaturase-2; DGAT1, Diglyceride O-acyltransferase 1; DGAT2, Diglyceride O-acyltransferase 2; 1-MNA, 1-methylnicotinamide; NAD, Nicotinamide adenine dinucleotide; Fatp4, Fatty acid transport protein 4; 4-HNE, 4-Hydroxynonenal; MDA, Malondialdehyde; SOD, Superoxide dismutase; PAI-1, Plasminogen activator inhibitor-1; AMPK, 5' AMP-activated protein kinase; Nrf2, nuclear factor erythroid 2 (NF-E2)-related factor 2; Keap1, Kelch-like ECH-associated protein 1; HO-1, Heme oxygenase 1; NF-ĸB, Nuclear factor kappa-B; TGF-β1, Transforming growth factor beta 1; TBARs, Thiobarbituric acid reactive.