Many epidemiologic studies have shown associations between increased CRP levels and increased risk for cardiovascular disease, diabetes, and multiple components of the metabolic syndrome including obesity, insulin resistance, dyslipidemia, and increased blood pressure1,4,5
. However, epidemiologic studies cannot resolve issues of pathogenesis and considerable controversy exists as to whether increased CRP functions as a disease marker, a disease mediator, or both. To directly investigate the effects of increased levels of human CRP on disease pathogenesis, it is necessary to determine the impact of actively perturbing CRP levels in the whole organism.
Infusions of highly purified CRP can be used to manipulate CRP levels in humans and in animals, however, such studies are typically performed on a relatively short term basis and are inconvenient for studying the pathogenic effects of chronic exposure to increased levels of CRP. Delivery of human CRP to animals by virally mediated gene transfer can facilitate chronic studies but can be complicated by immune reactions to the administration of heterologous forms of CRP. In contrast, transgenic techniques can be used to manipulate CRP levels over an extended period of time in vivo without many of the limitations associated with CRP injections or delivery of CRP by viral vectors.
Given the potential benefits of transgenic approaches for investigating the biologic effects of increased levels of CRP, many investigators have used transgenic mice expressing human CRP to study its role in atherosclerosis and cardiovascular function14–19
. However, relatively little work has been done in these models with respect to pathogenesis of features of the metabolic syndrome and risk factors for diabetes. Owing to the fact that CRP is not an acute phase reactant in mice and is synthesized in very low amounts in mice, some investigators have also raised questions about the utility of mouse models for studying the pathogenic effects of human CRP20–24
. Recently, several groups have advocated use of the rat as an alternative model for studying the biologic actions of human CRP24–26
. Because human CRP can activate rat complement and can be pro-inflammatory in rats, we investigated the blood pressure and metabolic effects of transgenically expressed human CRP in the SHR model that is genetically predisposed to developing multiple features of the metabolic syndrome.
In the current studies, we have found that in the SHR model, transgenic expression of human CRP in liver promotes insulin resistance, hypertriglyceridemia, as well as increased blood pressure. The serum level of human CRP in the transgenic rats was similar to the serum level of endogenous rat CRP in the transgene negative controls. Accordingly, in the transgenic SHR, the overall CRP level (the level of rat and human CRP combined) was increased by less than two fold compared to the normal CRP level observed in the SHR controls. This relative increase in total CRP level in the transgenic SHR is similar in magnitude to the relative increase in CRP observed in humans with metabolic syndrome. Thus, in the SHR model, serum levels of human CRP that are close to the endogenous levels of CRP normally found in rats can be associated with effects on the main hemodynamic and biochemical features of the metabolic syndrome. However, it should also be noted that the absolute increase in CRP in our rat model was very large compared to the absolute increase of CRP observed in humans with the metabolic syndrome. Therefore, in the SHR transgenic rats, as with any animal model, caution should be taken when extrapolating the results to humans.
Insulin resistance is considered to be a central component of the metabolic syndrome and in the current studies, we have found that transgenic expression of human CRP can promote disturbances in insulin and glucose metabolism including hyperinsulinemia and impaired insulin stimulated glucose incorporation into skeletal muscle glycogen. These findings are consistent with recent in vitro studies in which exposure of L6 myocytes to human CRP was found to decrease both insulin stimulated glucose uptake and glucose incorporation into glycogen27
. The in vitro studies by D’Alessandris et al. indicate that human CRP affects insulin stimulated phosphorylation of IRS-1 and suggest that human CRP causes disturbances in glucose metabolism by impairing insulin signaling pathways that regulate cell glucose transport27
In the current studies, transgenic expression of human CRP was also associated with reduced urinary excretion of cGMP suggesting that increased levels of CRP may be attenuating NO production. It has been reported that CRP might reduce the activity of endothelial nitric oxide synthase (eNOS) by affecting the stability of eNOS mRNA either directly or through the action of IL6 or TNFα28
. In addition, it has been shown that CRP can activate NADPH oxidase and cause increases in reactive oxygen species that may decrease NO production by inducing eNOS uncoupling26,29
. Consistent with the current findings, Guan et al.30
recently reported that virus mediated expression of human CRP in Wistar rats also induces increases in blood pressure (measured by direct puncture of carotid artery under anesthesia) and decreases in urinary cGMP. In addition, Vongpatanasin et al.31
have reported that transgenic expression of rabbit CRP in mice augments pressor responses to angiotensin II and promotes hypertension by inducing a decline in bioavailable NO and secondary downregulation of angiotensin II type 2 receptors in the vasculature. In contrast, alterations in angiotensin II type 1 receptor activity did not appear to play a role in the effects of CRP on blood pressure31
. Taken together, these observations support the possibility that increases in human CRP might be aggravating hypertension and features of the metabolic syndrome in part by reducing NO bioavailability. In future studies, it will be of interest to confirm whether the effects of human CRP on blood pressure in transgenic SHR rat are more likely mediated by downregulation of NO activity and AT2
receptors than by activation of AT1
Increased oxidative stress represents a well known mechanism that may mediate some of the adverse effects of increased CRP levels on risk for hypertension, diabetes, and cardiovascular disease12,13,26
. It has long been suspected that increased oxidative stress in liver and skeletal muscle might play an important role in risk for diabetes and that increased oxidative stress in the kidney may be an important determinant of hypertension. In the current studies, we found that transgenic expression of human CRP causes increased oxidative tissue damage in both liver and kidney. These observations suggest that increased CRP levels maybe influencing risk for the metabolic syndrome by inducing oxidative damage in a variety of tissues. As previously shown by Hein et al., activation of NADPH oxidase is one of the principal mechanisms whereby human CRP stimulates production of reactive oxygen species26
. The current studies suggest that impaired activity of glutathione peroxidase and reductions in glutathione levels could also be contributing to the effects of human CRP on oxidative stress and tissue damage.
It should be emphasized that the current studies on the metabolic effects of human CRP were performed in the SHR model which is a strain known to be genetically susceptible to developing multiple features of the metabolic syndrome. It is possible that other hypertensive models or strains might differ from the SHR with respect to their susceptibility to the adverse metabolic effects of human CRP. Having established that human CRP can aggravate features of the metabolic syndrome in the most widely used animal model of essential hypertension, future studies can now be planned to determine whether other strains or models exist that are more or less susceptible than the SHR to the adverse metabolic effects of human CRP. If so, this could open the door to linkage mapping of genetic factors that influence metabolic responses to increased levels of human CRP.