Literature Review: Acylation Stimulating Protein (ASP)

Literature Review: Acylation Stimulating Protein (ASP) 

By: Jen Gavia & Libby Quincey

In the United States, 7.8% of the population is diabetic and this disease continues to raise health care costs and effect individuals nationwide (Zhang, Lee, Cowan, Fabsitz, & Howard, 2011).  Type 2 diabetes increases the risk of obesity and other chronic diseases, such as hypertension, dyslipidemia, and cardiovascular disease (Pimentel, Zemdegs, Theodoro, & Mota, 2009).  Acylation Stimulating Protein (ASP) and insulin are two metabolic hormones that perform regulatory roles in lipid and glucose metabolism.  ASP levels may be an important cellular marker for insulin sensitivity and high concentrations correlate with diabetes (Schrauwen, Hesselink, Jain, & Cianflone, 2005).  In recent studies, endurance exercise and weight loss have shown to reduce ASP levels and promote ASP sensitivity.

In this paper the newly evolving findings on ASP are investigated and related to diabetes and exercise.  In section II, the production and purpose of ASP is reviewed.  Then in section III, ASP deficiency and resistance are explained and related to triglyceride clearance. Section IV compares the relationship of ASP to diabetes and obesity.  In section V, recent studies that involved weight loss surgery and caloric reduction offer insight on its effect on the protein.  In section VI, lipid abnormalities and ASP relationship to lipotoxicity are assessed.  In section VII, the effects on exercise and ASP levels response are investigated among nondiabetic and diabetic participants.  Then in section VIII, the findings of ASP and diabetes and the relationship between ASP and exercise are reviewed and complied.

II. PRODUCTION AND PURPOSE OF ASP  
In adipose tissue, several hormones are synthesized and secreted that have roles in fat metabolism, insulin action, and energy balance (Faraj et al., 2003). ASP is one of those hormones. ASP was isolated and identified less than 25 years ago with much still to learn about this protein (Cianflone et al., 1989). ASP, also referred to as C3adesArg, is a lipogenic adipocytokine (Cianflone, Xia, & Chen, 2003; Faraj et al., 2003; Schrauwen et al., 2005). ASP is found circulating in human plasma at concentrations of 10.3 to 58.1 nM in normal healthy adults based on 13 studies with a total of 376 subjects (Cianflone et al., 2003; Cianflone, 1992). ASP is produced through the alternate complement system (see Figure 1) in adipocytes, which involves three proteins: complement C3, factor B and adipsin (Cianflone et al., 2003). Human adipocytes are able to produce these three proteins late in differentiation and can begin to generate ASP (Maslowska, Sniderman, Germinario & Cianflone, 1997). In mature adipocytes, complement C3 is activated and combines with factor B to form an activated C3, factor B complex. Adipsin splits the complex and C3*Bb convertase is able to cleave inactive complement C3 into C3a and C3b. C3a is cleaved by carboxypeptidase B in the final step to produce ASP (Cianflone et al., 2003). Complement 5a receptor-like receptor (C5L2) is the specific receptor for ASP (Li & Hui, 2009). C5L2 is a newly identified receptor is found in white adipose tissue (WAT) and 3T3-LI cells (Faraj, Sniderman & Cianflone, 2004). Chylomicrons, lipoprotein particles that transport dietary fats within the body, and insulin have been found to influence the production of ASP and C3 (Cianflone et al., 2003; Yang et al., 2006).

Figure 1: The production of ASP through the alternate complement system (Cianflone, Xia, & Chen, 2003).

ASP has the ability to perform complex metabolic and endocrine functions, with its main purpose involving promotion of energy storage in non-diabetic subjects (Ahren, Havel, Pacini & Cianflone, 2003; Havel, 2002). The ASP pathway in the adipocytes interacts in a positive feedback mechanism that increases ASP secretion, increases lipogenesis, and decreases lipolysis, with the net effect being increased non-esterfied fatty acid (NEFA) trapping, or triglyceride storage, within adipocytes (Faraj et al., 2004).

ASP is able to increase triglyceride synthesis and storage in adipocytes in three ways. The first is by directly stimulating lipogensis in adipocytes. Through the ASP-C5L2 pathway, ASP is able to increase triglyceride synthesis by stimulating the activation of diacylglycerol acyltransferase (DGAT) (Li & Hui, 2009). DGAT is the enzyme that catalyzes the final and rate-limiting step in triglyceride synthesis (Cianflone, 1992; Schrauwen et al., 2005). By increasing DGAT, lipogensis is increased in adipocytes through enhanced free fatty acid esterification into stored triglycerides (Cianflone et al., 2003; Faraj et al., 2004; Yang et al., 2006). This process can be seen by an increased postprandial triglyceride clearance (Cianflone et al., 2003; Faraj et al., 2004; Yang et al., 2006). After a meal, fat is broken down and absorbed through the intestine and enters circulation as chylomicrons. Lipoprotein lipase (LPL) interacts with the chylomicrons at the endothelial cell surface to hydrolyze the triglycerides found in the chylomicrons, releasing free fatty acids. The free fatty acids are taken up by the adipose tissue and with the help of DGAT, re-esterified into triglycerides to be stored (Cianflone, 1997; Cianflone, Maslowska & Sniderman, 1999).

The second way ASP increases triglyceride synthesis is by increasing glucose uptake via glucose transporters (Ahren et al., 2003; Havel, 2002; Saleh et al., 1998). ASP is able to influence glucose uptake in adipocytes, fibroblasts, and myotubes (Ozata et al., 2001).  In differentiated adipocytes, once ASP binds to its C5L2 receptor, through a series of activation and phosphorylation, increase the translocation of glucose transporters (GLUT 1, GLUT 3, and GLUT 4) from intracellular vesicles to the plasma membrane (Cianflone et al., 2003; Li & Hui, 2009; Maslowska et al., 1997; Saleh et al., 1998). The glucose transporters allow glucose to enter the cell and glucose is the source of the glycerol molecule that is the backbone of a triglyceride molecule (Maslowska et al., 1997).  The final way ASP can stimulate triglyceride synthesis is by the inhibiting hormone-sensitive lipase (HSL) activity (Ahren et al., 2003; Havel, 2002; Li & Hui, 2009).

The effects of ASP, in terms of glucose transport and fatty acid storage, supplement, but are independent of the actions of insulin, another lipogenic hormone (Cianflone et al., 2003; Faraj et al., 2003; Van Harmelen et al., 1999). Both hormones have a major role in regulating free fatty acid release from adipocytes. In a study by Van Harmelen et al. (1999), it was seen that ASP stimulates free fatty acid re-esterification, while insulin also stimulates fractional free fatty acid re-esterification, but also inhibits lipolysis. It was concluded from the study that the combination of both hormones result in a decrease in net fatty acid release from the adipocytes and almost complete (97%) re-esterification of available fatty acids in the plasma into fat storage (Van Harmelen et al., 1999).

III. ASP DEFICIENCY AND RESISTANCE
Without ASP, the aforementioned lipid metabolism functions would be altered. A dysfunctional ASP pathway may be the result of an absence of ASP (ASP deficiency) or a reduced response to the action of ASP (ASP resistance) (Cianflone et al., 2003). ASP deficiency, although rarely seen in humans, may be caused by a genetic mutation leading to a lack of complement C3 (Cianflone et al., 2003). Absence of any of the proteins involved in the alternate complement system, depicted in Figure 1, could also cause a deficiency in ASP (Cianflone et al., 2003). High plasma ASP levels may be a marker for ASP resistance (Faraji et al., 2004).

However, certain strains of mice (129Sv and C57B1/6) have been genetically altered to be ASP deficient (Cianflone et al., 2003). These strains of mice are called C3 knockout (C3 KO). These mice, which cannot produce ASP because of the lack of C3, are characterized by delayed postprandial triglyceride clearance, increased NEFA, reduced fat mass, and insulin sensitivity (Murray, Havel, Sniderman & Cianflone, 1999a; Murray, Sniderman, Havel & Cianflone, 1999c). Murray, Sniderman & Cianflone (1999b), were first to see that triglyceride clearance slowed down in 129Sv male C3 KO mice, as compared to their wild-type (WT) complements, but once given an intraperitoneal injection of human ASP, triglyceride clearance returned to normal. Murray et al., (1999a) and Murray et al., (1999c) both confirmed this result in their respective studies. Murray et al., (1999a) and Murray et al., (1999c) also demonstrated that male and female C3 KO mice, respectively, had reduced fat mass than the WT mice. Decreased total body weight and direct measurement of adipose tissue depots at inguinal, pectoral, gonadal and perirenal indicated reduced fat mass, as well as lowered plasma levels of leptin, which is an indicator of adiposity (Murray et al., 1999a; Murray et al., 1999c). The leanness of the C3 KO mice might play a role in the apparent insulin sensitivity of these mice, but it was also shown in decreased fasting glucose, insulin, and oral glucose tolerance tests results that suggested a more insulin sensitive state (Murray et al., 1999a; Murray et al., 1999c).

IV. RELATIONSHIP TO DIABETES
Type 2 diabetes mellitus is characterized by the combination of insulin resistance and the decreased functioning of pancreatic B-cells that secrete insulin in response to glucose.  (Li & Hui, 2009).  Insulin sensitivity is reflected by the amount of glucose uptake and utilization by a given quantity of exogenous or endogenous insulin (Lebovitz, 2001). As previously mentioned, insulin stimulates the production of ASP by influencing its precursor protein C3 from adipocytes.  The concentrations of C3 and ASP are inversely related to insulin sensitivity and increased concentrations are evident in type 2 diabetics (Koistinen, et al., 2001).  It is suggested that ASP plasma levels may be an important intracellular marker for insulin sensitivity (Schrauwen et al., 2005).  As shown in Figure 3, ASP functions to enhance glucose transport into cells, therefore, reduced ASP levels reflect improved insulin sensitivity. If there are lower levels of ASP in the plasma it means ASP, and insulin, have done an efficient job of clearing glucose out of the bloodstream and the cells are more sensitive to both ASP and insulin.

Chronically elevated ASP concentrations and metabolic disruptions in glucose and lipid roles are linked to diabetes and plasma lipid abnormalities (Cianflone et al., 2003).  The abnormal ASP-C5L2 pathway of regulatory carbohydrate and lipid metabolism causes the opposite functions to occur. This leads to decreased glucose uptake and elevated plasma free fatty acid (FFA) levels, hyperglycemia, and lipotoxicity.  This results in progressive apoptosis and failure of B-cells, associated with insulin resistance and the progression of diabetes (Li & Hui, 2009).

Figure 3: Insulin promotes ASP secretion and ASP Regulatory Roles influence glucose and fat storage (Havel, 2002).

These metabolic disruptions were analyzed in a study by evaluating the relationship of insulin response and the concentrations of ASP, C3, adiponectin and C-reactive protein (CRP) by Yang et al. (2006).  The Chinese diabetic participants (n=187) were subdivided into: control group (nondiabetic obese and nondiabetic lean) and treatment group (diabetic obese and diabetic lean).  Even though ASP, adiponectin, and C3 are all produced by adipocytes and associated with similar components like glucose, insulin, and lipids, the study found that these hormones did not correlate with each other.  It can be suggested that each perform independent regulatory roles, which contribute to diabetes separately.
Fasting and postprandial ASP levels were consistently higher in obese diabetics when compared to the nondiabetics, but not different from lean diabetic group.  In both diabetic lean and diabetic obese groups, serum C3 concentrations were significantly higher when compared to the nondiabetic control groups (P=0.0001).  C3 was strongly associated with insulin status by correlating with glucose (r=0.283), insulin (r=0.301), and homeostatic model assessment of insulin resistance (HOMA-IR) (r=0.266).  C3 and CRP strongly correlated with HOMA-IR and ASP or adiponectin did not correlate with HOMA-IR.

Figure 4: ASP and BMI in nondiabetic group (Yang et al., 2006).

Figure 5 ASP and BMI in diabetic group (Yang et al., 2006).

It was found that ASP concentrations were significantly higher in obese groups (p=0.0024), 43% in the nondiabetic obese and 33% in the diabetic obese group. ASP primarily correlated with body mass index (BMI) (r=0.241, P=0.001) and lipids.  ASP correlated with the following lipid and lipoprotein factors: NEFA (r=0.143, P<0.05), triglyceride (r=0.180, P=0.015), total cholesterol (r=0.206, P=0.005) and Apo B (r=0.229, P=0.016).  Higher C3 and ASP levels were found in diabetic subjects, despite the weight difference.  This suggests that abnormal ASP levels are correlated with type 2 diabetes, and not only obesity (Yang, et al., 2006).

In a recent study, the regulation of adipose tissue C3 mRNA and insulin sensitivity among lean healthy men (n=9), obese nondiabetic (n=9), and diabetic men (n=12) were compared.  After an overnight fast and blood sampling, participants ingested a high fat meal and afterwards ASP levels were measured at: 0, 1, 2, 3, 4, 6, 8, and 10 hours.  Then on a separate day, the expression of C3mRNA was measured before and after a 240-minute euglycemic hyperinsulinemic clamp (EHC).

It was found that the fasting ASP levels and C3mRNA expression were not significantly different in obese nondiabetic and in type 2 diabetic men, but were slightly higher than in lean healthy men (P<0.005).  This suggests that ASP and C3mRNA levels correlate with adiposity, since the obese men had slightly higher fasting ASP levels and C3mRNA expression as compared to the lean healthy men.  Similar to other research, it can be suggested that high levels of fat mass and body weight correlate with higher concentrations of ASP in the plasma and C3mRNA expression (Cianflone et al., 2003).

After the oral fat load, C3 concentrations did not change significantly in both lean (p=0.14) or type 2 diabetic men (p=0.09).  ASP levels did not increase after an oral fat load in any of the three groups of participants.  When analyzed separately, only nondiabetic men showed fasting plasma ASP levels correlated with the area under the serum triglyceride concentration curve (r= 0.73), fasting (r=0.75) and maximal (r= 0.64) serum triglyceride concentrations during the oral fat tolerance test.  The diabetic men did not correlate with postprandial triglyceride response, which suggests abnormal lipid metabolism.
As partly shown on Figure 6, subcutaneous adipose tissue C3mRNA in nondiabetics were negatively correlated with whole body glucose disposal rate (r=-0.79) and high density lipoprotein (HDL) levels (r=-0.70) and positively correlated with BMI (r=0.72), waist-to-hip ratio (r=0.70), and fasting leptin (r=0.64).  However, in Type 2 diabetic men these correlations were not evident.

Figure 6: Negative correlation between C3 and whole body glucose disposal rate in nondiabetic men in the graph above.  Positive correlation between C3 and BMI in nondiabetic men in the graph below.

When the ASP pathway is properly functioning, it works to enhances insulin sensitivity and increase plasma HDL, as shown in Figure 7.  In this study, these associations were found in the nondiabetic participants and not found in the type 2 diabetics.  Since C3mRNA expression was not correlated with insulin sensitivity in the diabetic men, this suggests abnormalities with glucose metabolism in the body (Koistinen, et al., 2001).

Figure 7: ASP roles in glucose and lipid metabolism (Ben-Ezra, 2006).

According to another study by Weyer, Tataranni, and Pratley (2000) of 33 nondiabetic Pima Indians, the relationship between fasting C3 and ASP concentrations were compared to adiposity, insulin action, and insulinemia.  Plasma glucose was determined by a 75-g oral glucose tolerance test and insulin was determined by an insulin-stimulated glucose disposal (M) by EHC.  ASP was measured by the radioimmunonassy and C3 was analyzed by an immunoturbidimetic assay.

As shown in the Figure 8, fasting ASP and fasting C3 were positively correlated (r=0.43).  Significant positive correlations were found between C3 and BMI (r=0.78), waist-to-thigh ratio (r=0.43), fasting glucose levels (r=0.47), and 2-h insulin concentrations (r=0.58).  Fasting ASP levels were positively correlated with the percent of body fat only (r=0.37) and were not correlated with M, fasting insulin concentration, or 2-h glucose concentrations.  There was no significant correlation between ASP and BMI, waist-to-thigh ratio, fasting glucose, or 2-h insulin concentration.

Figure 8: The relationship of fasting ASP and fasting C3 concentrations (Weyer et al., 2000).

In this study, the fasting ASP was related to adiposity and C3 concentrations, and not to insulin action or insulinemia (Weyer et al., 2000).  This is contrary to previous findings that have found that ASP is associated with insulin sensitivity and numerous lipid factors.  In the study by Koistinen et al. (2001), significant correlation between C3mRNA expression and glucose disposal rate in nondiabetics were found.  According to another study by Yang et al. (2006), ASP correlated with adiposity as well, including BMI (r=0.241, P=0.001), as well as numerous lipid factors.  Overall, the role of ASP with insulin resistance still remains to be determined and further research to investigate this relationship would be beneficial.

V. WEIGHT LOSS
In western societies, obesity has become an epidemic and increased ASP levels are directly associated with obese states (Schrauwen et al., 2005). The proteins C3, ASP, factor B, and adipsin are all elevated in obese and diabetic states, but reduce with weight loss or exercise (Cianflone et al., 2003). As previously mentioned, ASP has a positive relationship to adiposity or body weight, meaning that circulating levels increase with obesity (Faraj et al., 2003). Researchers are also interested in what happens to ASP levels following weight loss and if changes in this hormone, as well as adiponectin, leptin, and ghrelin are predictive of an improved metabolic and cardiovascular risk profile (glucose, insulin, and lipid parameters) (Faraj et al., 2003). Leptin and ASP perform different functions but both hormones affect the storage or oxidation of energy (Xia, Sniderman, & Cianflone, 2002).  In order to test this hypothesis, Faraj et al. (2003) used 50 (39 women and 11 men) morbidly obese subjects (BMI=50.2 ± 8.1 kg/m2) that underwent standardized isolated longitudinal Roux-en-Y gastric bypass surgery. The subjects were divided into two groups based on their weight loss status postoperatively in the 6-month period preceding the final measurement.  If the subjects were still losing weight of more than 10% over the 6-month period, they were deemed weight reducing (negative energy balance) (n=25, 21 women). If they had a weight loss of less than 10% they were put in the weight stable group (neutral energy balance) (n=25, 18 women).

As should be expected, most of the subjects (86%) had elevated plasma ASP levels before the surgery (>23.5±10.8 nM). After the gastric bypass surgery, BMI dramatically decreased with about 16-55% weight lost.  Seven subjects in the weight stable group, as well as four subjects from the weight reducing group reached normal BMI range (20-25 kg/m2). However, most subjects were still considered obese (BMI>30 kg/m2) even post-surgery. ASP did decrease in most subjects after gastric bypass, but since the majority of subjects in the weight stable and weight reducing groups (52% and 64%, respectively) were still considered obese, ASP concentrations were still higher than normal. This can be seen in Figure 9, where the arrows pointing to the left are average ASP concentrations pre-surgery, and the arrows pointing to the left are average post-surgery levels. The rectangular box is the normal reference value used in this study (23.5 ± 10.8 nM). The weight stable group had an average ASP concentration of 31.4±19.8 nM and the weight reducing group averaged 35.5±22.5 nM.

Figure 9: Pre- to postoperative changes in plasma ASP in weight-stable and weight-reducing groups (Faraj et al., 2003)

The correlation analysis showed that for every plasma parameter measured, including ASP, there was a positive correlation between the preoperative value and the postoperative change. Therefore, the higher the preoperative value, the greater the percent change, postoperative. However, baseline plasma preoperative levels of ASP and leptin did not support the hypothesis that preoperative concentrations of these hormones would predict the degree of weight loss after surgery. Adiponectin levels best predicted postoperative weight loss (r = -0.59; P = 0.02). ASP was predictive of some of the improved lipid profiles in weight stable subjects. ASP accounted for 55% (P = 0.009) of the 67% decrease in apolipoprotein B (apoB), an independent risk factor for ischemic heart disease.
This study supported the notion that ASP levels decrease with a decrease in adiposity, albeit the extent of weight loss determines the extent of reduction of ASP concentration. However, further research is needed concerning ASP and other hormones synthesized and secreted by adipose tissue, in regards of extent of weight loss and the effects of weight stability, post weight loss.

VI. LIPID
Lipid storage and ASP are interconnected and genes that affect total cholesterol, low density lipoprotein (LDL), and triglycerides, also influence ASP concentrations (Havel, 2002).  The Randle Cycle, also known as glucose fatty acid cycle, is a homeostatic mechanism in the body that functions to control the metabolism of carbohydrates and lipids (Hue & Taegtmeyer, 2009).  Since ASP functions to regulate lipid and glucose metabolism, the ASP pathways intertwin with lipids.

LPL is synthesized and secreted by adipose and muscle tissue, and it is an important enzyme that hydrolyzes circulating lipoproteins and releases FFA (Li & Hui, 2009).  Insulin is a major stimulator of LPL activity.  ASP increases LPL in adipose tissue, while decreasing LPL in muscle tissue.  However, if abnormalities occur in the ASP pathway, ASP inhibits LPL in the adipose tissue, and increases LPL in the muscle (Faraj et al., 2004).  The regulatory roles are reversed when metabolic disruptions occur with ASP.  As a result, FFA trapping is reduced in adipocytes, and there are higher concentrations of FFA in the plasma (Koistinen et al., 2001).  This may prevent proper functioning of vital organs and ineffective lipid storage, which can lead to lipotoxicity and other complications, including metabolic syndrome.  In lipotoxicity, fatty acids accumulate in tissues such as the liver, skeletal muscle, and pancreatic β cells (St-Pierre, et al., 2008).  When excess fat is shunted to pancreatic β cells, apoptosis occurs, which causes decreased insulin production to occur. High levels of FFA and glucose remain in the blood stream and could lead to insulin resistance (Li & Hui, 2009).

As shown in the study by Faraj et al. (2003), lower ASP levels were the most predictive variable of the plasma apolipoprotein B (apoB) levels after weight loss. ApoB is the primary apolipoprotein of LDL and apoB overproduction results in increased fatty acid flux to the liver. This increased fatty acid flux to the liver may be the driving component to dyslipidemia.  Increased fatty acid flux to adipose tissue results in fat storage and increased ASP secretion.  With caloric restriction, the reduced ASP and apoB levels may reflect diversion of fatty acids for energy demands, reducing both adipose storage and excess hepatic lipoprotein secretion. (Faraj et al., 2003)

VII. RELATIONSHIP TO EXERCISE
Endurance exercise is known to improve fatty acid metabolism and insulin sensitivity.  Therefore, in order to investigate the impact that endurance training has on ASP levels and ASP sensitivity, a two-week exercise-training program of young, untrained, healthy male participants (n=8) was designed.  The participants either performed 2 hours of endurance exercise, or 45 minutes of intermittent exercise, and blood was taken pre and post exercise during the two-week program.  On day 2 and 16, insulin sensitivity was assessed by insulin tolerance test after an overnight fast by EHC.

It was found that the two-week training program resulted in 25% decrease in fasting ASP levels.  According to the findings, it can be suggested that endurance training in nondiabetics reduced ASP levels, implying improved ASP sensitivity.  It was found that baseline ASP levels correlated positively with insulin resistance both before (r=0.86, P<0.01) and after training (r=0.82, P<0.05) and that insulin sensitivity was slightly higher after the two weeks.  Although this study was not able to prove the association between ASP and insulin sensitivity, it can be inferred through ASP’s function of enhancing glucose uptake into adipose tissue and lower ASP levels in the plasma.  This also might be due to the additive effects of exercise on improving insulin sensitivity, which is why a conclusion cannot be made from the present study.

As shown in Figure 10, prior to the training program, ASP levels decreased significantly during the baseline 3 hr submaximal exercise that was used to calculate substrate oxidation..  Then post training program, ASP levels slightly increased during the 3 hr exercise, but not significantly.  This slight increase may be in order to avoid excessive increases of FFA in the plasma, some of which are being used as an energy source, by decreasing lipolysis and stimulating FFA clearance and re-esterification into adipose tissue. Endurance training leads to metabolic adaptations that lead to improved fatty acid re-esterification and fatty acid control and handling.  During and after exercise, ASP is important for fatty acid re-esterfication in adipocytes, by stimulating glucose uptake and promoting the enzyme DGAT (Schrauwen, Hesselink, Jain, & Cianflone, 2005).

Figure 10: The relationship of ASP concentration before and after the training program (Schrauwen et al., 2005).

In a study by Dufaux & Order (1989), ASP levels increased by 29% during a timed run and then returned to baseline values shortly afterwards (Dufaux & Order, 1989).  According to another study, it was found that ASP levels increased acutely by 67% during a timed run then returned to baseline shortly afterwards.  C3 did not produce any exercise-induced changes but baseline values were lower in the athletes compared to the control group (Smith, Chi, Krish, Reynolds, & Cambron, 1990).
In a previous study by St-Pierre et al. (2008), the purpose was to examine the potential changes in plasma ASP during a EHC and to evaluate the metabolic profile of the subjects according to their changes in plasma ASP.  There were 76 nondiabetic overweight and obese postmenopausal women between 48-73 years old that were subdivided in the following three groups: negative ASP responders (NAR), positive ASP responders (PAR), and zero ASP responders (ZAR).  In the study, EHC and plasma ASP were measured at 0, 60, 160, 170, and 180 minutes.  In PAR, ASP levels were significantly higher when compared with fasting conditions.  PAR had higher BMI, visceral fat, fasting insulin levels, lean body mass, and alanine aminotransferase (ALT) than NAR.

 Collectively, it was found that there was a positive association (r=0.29) between fasting ASP levels and VO2 peak, while in the previous study by Schrauwen et al., (2005) an inverse relationship was found.  Elevated ASP response during EHC may be associated with impaired ASP activity resulting in slow triglyceride clearance, ASP resistance, and abnormal lipid and carbohydrate metabolism.  These results may also be influenced by gender (female), hormonal differences (postmenopausal), activity level (sedentary) and BMI (overweight and obese) in this study.  While in the by St-Pierre et al. (2008), the participants were male, moderately trained, and at a normal BMI.

VIII. CONCLUSION
In conclusion, ASP is an important adipocyte produced protein for glucose and lipid metabolism.  Insulin and ASP are two metabolic hormones that perform independent and additive functions.  When abnormalities in the ASP pathway occur then the regulatory roles for metabolism are prevented.  Abnormal lipid storage, increased FFA in the plasma, and impaired glucose uptake link abnormalities of the ASP pathway to type 2 diabetes and other metabolic syndromes like lipotoxicity.  Endurance training can promote insulin sensitivity and reduce ASP levels.  There is still a lot to learn about this hormone and further research should be done regarding ASP due to differences found in studies.

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