Metabolic syndrome is a group of states that take place together, increasing the risk of type 2 diabetes mellitus and steatohepatitis. These states consist of high blood glucose and pressure, extra fat around the waist, and dyslipidemia (1). It has a high prevalence in developed countries (about 22.4 % in adult individuals) (2). There is a body of growing evidence suggesting that obesity is the most important factor in the development of metabolic syndrome (3-5).
The pathogenesis of metabolic syndrome is still controversial; 3 pathways are under the spot light including insulin signaling, hepatic lipogenesis and fatty acid β-oxidation (6-8). Because of high calorie diet and a sedentary life style, especially in developed countries, researchers believe that by adding supplements containing agents whose can affect the 3 mentioned pathways, lipid metabolism can be improved. Recent studies demonstrate that fat accumulation leads to inflammation. Inflammatory mediators, in cells which have receptors for insulin hormone, will cause insulin resistance. In this state; muscles, adipose tissue, and liver fail to respond well to insulin and cannot easily take up glucose from blood circulation (9-11). Hepatocytes and adipocytes are two important groups of cells which can develop insulin resistance; which is regarded as the most effective factor in the pathogenesis of hyperglycemia and hepatic steatosis (12). It seems that adipocytes, hepatocytes and pancreatic β-islet cells are vertices of a triangle in the progression of the metabolic disorders which lead to insulin resistance and its consequences (13-15).
Peroxisome proliferator-activated receptors (PPARs) are a group of ligand inducible transcription factors that play an important role in expression of proteins which are involved in lipid metabolism. Studies indicate that PPARs activation can improve insulin resistance. Based on previous studies, PPARs agonists seem to be useful and effective in prevention of development of insulin resistance (16-19).
Isoflavonoids are a cluster of flavonoid phenolic compounds. They are well known as phytoestrogens because of biological effects through estrogen receptors. In addition to estrogenic effects, some other biological effects of isoflavonoids have been identified recently such as activation of PPARs (16).
Genistein is one of the better-known soy derived isoflavonoids that has attracted the attention of scientists because of its beneficial effects on insulin resistance (20). The effects of Genistein in balancing carbohydrate and lipid metabolisms have been proved. Biochanin A and Formononetin are methylated forms of Genistein and demethylation of these compounds usually takes place in the intestine by acetogenic bacteria (Figure 1) (17,21-23).
Although diverse metabolic effects of Genistein have been clearly indicated, this narrative review will mostly focus and highlight the effect of Genistein in insulin resistance development in both in vitro and in vivo models from different studies.

Figure 1. Chemical formula of Genistein and Biochanin A (17).

 Abbreviations' list

ACC: Acetyl CoA Carboxylase
AMPK: Adenosine Monophosphate-activated Protein Kinase
ACOX: Acyl-coenzyme A Oxidase 
AP-1: Activating Protein 1
AS160: Akt Substrate of 160 kDa
CPT1: Carnitine Palmitoyltransferase I 
CRE: CREB Response Element
CREB: cAMP Response Element-binding Protein
Epac1: Exchange factor directly activated by cAMP 1
ERK: Extracellular Signal-regulated kinase
FAS: Fatty Acid Synthase
FOXO1: Forkhead Box O1
G6pase: Glucose 6-phosphatase
Grb2: Growth Factor Receptor-bound protein 2
GSK3: Glycogen Synthase Kinase 3
IRS: Insulin Receptor Substrate
JAK2: Janus Kinase 2
JNK: c-Jun N-terminal Kinase
LXR: Liver X Receptor
MAPK: Mitogen Activated Protein Kinase
MEK: MAPK/ERK Kinase
p-CREB: Phosphorylated cAMP Response Element-binding Protein
PDPK1: Phosphoinositide-dependent Protein Kinase 1
PEPCK: Phosphoenolpyruvate Carboxykinase
PGC1α: PPARγ Coactivator 1 alpha
PI3K: Phosphatidylinositol 3 Kinase
p-IRS: Phosphorylated Insulin Receptor Substrate
PKB: Protein Kinase B
PLC: Phospholipase C
p-LXR: Phosphorylated Liver X Receptor
PPARα: Peroxisome Proliferator-activated Receptor α
PPRE: PPAR Response Element
p70S6k1: p70 S6 Kinase 1
Raf-1: Rapidly Accelerated Fibrosarcoma
Ras: Rat Sarcoma
SCD1: Stearoyl-coA Desaturase
SOS: Son of Sevenless
SREBP-1c: Sterol Regulatory Element-binding Protein 1-c
TNF(α): Tumor Necrosis Factor (α)
UCP2: Uncoupling Protein 2
 
Metabolic Effects of Genistein on Liver
Liver is the main center of carbohydrate and lipid metabolism in human organism; in other words, the liver regulates the homeostasis of glucose and lipids in blood circulation. After meal and glucose absorption, insulin, is secreted from β-islet cells of the pancreas into blood circulation (13). Insulin binds to its receptor in hepatocytes and autophosphorylation of insulin receptor will occur in tyrosine residues. Insulin receptor is a tyrosine kinase protein, which phosphorylates insulin receptor substrate1,2 (IRS 1, 2) in tyrosine residues and initiates the insulin signaling cascade. P-IRS binds to PI3k and helps the PI3k to phosphorylate the PIP₂ to PIP₃. Akt (PKB) activates when it binds to PIP₃ and will have been phosphorylated by PDPK1. Activated Akt phosphorylates the serine and threonine residues in GSK3; in this way, inactivates the GSK3. Due to inactivation of GSK3, GS continues the synthesis of glycogen; resulting in glucose gradient from blood to liver (24-26). Additionally, activated Akt phosphory-lates the FOXO1 transcription factor. FOXO1 inactivates during phosphorylation and in this way, mRNA expression of enzymes which have important roles in gluconeogenesis such as PEPCK and G6pase, will decrease. Actually, hepatocytes uptake the blood glucose and downregulate the gluconeogenesis pathway for homeostasis of blood glucose (27).
Obesity is the most important risk factor of hyperglycemia. Since the liver is the main center of lipogenesis, in obese persons, lipid accumulation in hepatocytes usually occurs. Peroxidation of fatty acids produces free radicals which lead to oxidative stress. Inflammation initiates in response to oxidative stress and inflammatory mediators accumulate in the liver. Inflammatory mediators activate different kinases; p70S6k1 is one of the important enzymes. p70s6k phosphorylates the IRS in serine residues. If IRS phosphorylates in the serine residues, it will degrade in proteasomes; so, insulin signaling cascade will not initiate and insulin resistance will occur (24,25). Genistein is an AMPK activator and PPARα agonist (28). AMPK is a kinase which activates in response to AMP and phosphorylates the CREB transcription factor. P-CREB binds to CRE in the promoter of PPARα gene and increases the mRNA expression of PPARα transcription factor (29). Proteins, which are involved in fatty acid β-oxidation such as CPT1, ACOX and UCP2, have PPRE in promoter region of own genes, therefore the expression of these proteins will increase in response to increased PPARα protein expression (Table 1) (30-32). Furthermore, AMPK phosphorylates and inactivates the LXR nuclear receptor in threonine residues; in this way, AMPK decreases the mRNA expression of SREBP-1c transcription factor. SREBP-1c is a transcription factor which upregulates the mRNA expression of enzymes involved in lipogenesis such as FAS, ACC and SCD1. That is to say, decreased protein expression of SREBP-1c leads to decreased expression of proteins involved in lipogenesis resulting in decreased lipogenesis (Table 1) (33,34). Also, it should be noted that AMPK phosphorylates and inactivates the p70S6K1, the enzyme which secretes from inflammatory mediators and inhibits the initiation of insulin signaling cascade. P70S6K1 is an effective factor in mRNA expression of SREBP-1c. It has been otherwise stated that p70S6K1 phosphorylates the LXR in serine residues and then p-LXR upregulates the SREBP-1c mRNA expression (34,35).
As a result, Genistein can increase fatty acid β-oxidation, reduce lipogenesis and improve insulin resistance in hepatocytes.
Metabolic Effects of Genistein on Adipose Tissue
Another group of cells which have receptors for insulin are adipocytes. Insulin binds to its receptor on adipocytes and PI3K/Akt signaling cascade will initiate. In adipocytes, activated p-Akt inactivates the AS160 protein during phosphorylation in tyrosine residues. Activated AS160, changes the Rab-GTP to Rab-GDP. On the other hand, GLUT4 translocation to the cell membrane is dependent on Rab-GTP. Therefore, by increased number of Rab-GTP due to inactivated AS160, the number of GLUT4 on the cell membrane of adipocytes will increase. Also, it can be said that by increasing the number of GLUT4 on cell membranes, glucose uptake by adipocytes will increase and as a result, the glucose concentration in the blood circulation will decrease (36,37).
As previously mentioned, obesity is the most important risk factor for insulin resistance. Lipid accumulation in adipocytes will cause inflammation and secretion of inflammatory mediators. TNFα is one of the inflammatory mediators which activates the p70S6K1. This enzyme, like in hepatocytes, phosphorylates the IRS in serine residues and in this way inhibits the insulin signaling cascade. The result of insulin signaling inhibition in adipocytes is the decreased number of GLUT4 on cell membrane. By decreasing the number of GLUT4 on adipocytes' membrane, the rate of glucose uptake will decrease and because of this, the glucose concentration in the blood circulation will increase, which means insulin resistance (34,35).
In addition to PI3K/Akt cascade, insulin initiates the other signaling cascade which is called MAPK/Ras signaling cascade. Similar to PI3k/Akt pathway, tyrosine phosphorylation of IRS is the first step of the MAPK/Ras pathway. In the next step, p-IRS interacts with Grb2; the protein which recruits SoS to change the Ras-GDP to Ras-GTP. A complex is made by binding the Ras-GTP to Raf-1 and phosphorylates the MEK protein family. Likewise, p-MEK phosphorylates the MAPK protein family such as ERK1/2/5, p38MAPK and JNK. Finally, p-MAPKs, through phosphorylation of various transcription factors in serine and tyrosine residues at the nucleus, regulate the expression of various genes. It should be noted that, in addition to insulin, Ras/MAPK pathway activates in response to growth factors, environmental stress and pro-inflammatory mediators (38,39).
TNFα is one of the pro-inflammatory mediators which can phosphorylate and activate JNK through MAPK/Ras pathway. P-JNK, phosphorylates c-jun in cytoplasm and then, c-jun binds to c-fos to make Ap-1 complex. Ap-1 is a transcription factor which translocates to the nucleus and downregulates the expression of adiponectin mRNA in adipocytes so synthesis and secretion of adiponectin will decrease subsequently (40). Adiponectin is a peptide hormone which is secreted from adipocytes into the blood circulation (41-43). Adiponectin binds to its receptors on skeletal muscle cells and hepatocytes; facilitates the insulin signaling cascade through the phosphorylation of PkB in an alternative pathway. As a result, by decreased expression of adiponectin mRNA and decreased concentration of adiponectin in the bloodstream subsequently, insulin sensitivity in skeletal muscle cells and hepatocytes will decrease (44,45). On the other hand, p-JNK phosphorylates the IRS in serine and threonine residues. As stated above, p-IRS in serine and threonine residues cannot initiate the insulin signaling cascade; it consequently will cause insulin resistance (34).
Genistein, in adipocytes, prevents phosphorylation of JNK, in other words the mRNA expression of adiponectin will not inhibite. That is to say, Genistein is an agonist of PPARγ transcription factor which is one the most important transcription factors in upregulating the expression of adiponectin mRNA. As a result, Genistein prevents from downregulation of adiponectin mRNA expression and facilitates the upregulation of adiponectin mRNA expression simultaneously (Table 1) (46).

Metabolic Effects of Genistein on Skeletal Muscle
The other group of cells which have a receptor for insulin are skeletal muscle cells. That is to say, the insulin-receptor complex stimulates the cellular uptake of glucose. Obesity and hyperlipidemia lead to lipid deposition in skeletal muscle cells (47,48). Insulin resistance, as previously mentioned, is the consequence of lipid accumulation. In other words, in insulin resistant state, insulin-dependent glucose uptake is markedly decreased in skeletal muscle cells (49).
Leptin is a peptide hormone dominantly secreted by adipocytes and has receptors on skeletal muscle cells. When leptin binds to its specific receptor, it phosphorylates the JAK. In the next step, p-JAK phosphorylates and activates AMPK. It should be added that AMPK is the key protein in fatty acid β-oxidation signaling pathway (50-53). Activated AMPK (p-AMPK), phosphorylates and inactivates ACC. During the inactivation of ACC, the concentration of Malonyl-CoA will decrease. Malonyl-CoA is regarded as a fatty acid β-oxidation inhibitor agent. As a result, by decreased concentration of Malonyl-CoA, fatty acid β-oxidation will increase. On the other hands; p-AMPK translocates to the nucleus and activates the PPAR-α and PGC1α transcription factors. By the PPARα and PGC1α activation, the mRNA expression of genes involved in fatty acid β-oxidation like CPT-1 and UCP2 will increase (54-61). In conclusion, leptin signaling pathway prevents lipid accumulation in skeletal muscle cells.
The development of obesity and lipid accumulation in skeletal muscle cells result in leptin resistance and lipid deposition in skeletal muscle cells will severely intensify (62-64). In other words, leptin resistance is associated with insulin resistance.
Genistein stimulates fatty acid β-oxidation in skeletal muscle cells in a way independent of leptin receptor. Based on recent studies, Genistein activates the AC in skeletal muscle cells and the concentration of cAMP increase. In the next step, increased cAMP activates Epac1, which in turn activates the PLC (65-68). Activated PLC phosphorylates the JAK2 and as a consequence, the phosphorylation of AMPK occurs. As mentioned previously, activated AMPK stimulates the fatty acid β-oxidation and in this way, prevents lipid accumulation. In conclusion, the cAMP-JAK-AMPK pathway plays an important role in regulating the fatty acid β-oxidation. Through stimulating the mentioned pathway, Genistein can be a beneficial agent in prevention of lipid accumulation in skeletal muscle cells.

Effects of Genistein on Pancreas
After meal and glucose absorption, the increased concentration of blood glucose leads to glucose uptake by pancreatic β cells, which is the main stimulator of insulin secretion. Insulin facilitates the glucose uptake by adipocytes and skeletal myocytes and in this way prevents hyperglycemia. It should be noted that insulin is the most important agent in the pathophysiology of hyperglycemia. In other words, in hyperglycemic patients, there is an insufficient secretion of insulin due to complete or partial destruction of pancreatic β-islet cells. It seems feasible that preventing from destruction of the pancreatic β-islet cells can be a therapeutic target in hyperglycemic patients (69).
Due to recent studies on β-islet cells, Genistein initiates the Ac/cAMP/pKA/p-ERK1,2 cascade which leads to proliferation of β cells. Also, β-islet cells proliferation is accompanied with increased secretion of insulin (Table 1) (70-73).
Genistein is an estrogen receptor agonist but its effect on β-islet cells proliferation is independent of estrogen receptors. In other words, by blocking the estrogen receptors, the effect of Genistein on β-islet cell proliferation remains (74).
Genistein stimulates the GIIS in an alternative pathway. On the basis of previous studies, Genistein stimulates GIIS through the cAMP/pKA pathway; which leads to increase intracellular ca²⁺. This pathway is controversial; proteins which are specific in this pathway are still unknown. Blocking the intracellular protein synthesis lead to decreased secretion of insulin (Table 1) (74,75).

Table 1. Summary of researches' results about metabolic effects of Genistein