In addition to neurological impairment, metabolic changes are an important clinical problem during the progression of HD in human patients. Here we have characterized alterations in a key metabolic tissue, white adipose tissue, in two mouse models of HD. Both the R6/2 Tg and CAG140 KI mouse strains exhibited evidence of adipose tissue dysfunction with disease progression. Despite the fact that the time course of disease in the two models ranges from 12–15 weeks for R6/2 Tg to more than 2 years for CAG140 KI mice, both strains had progressive reductions in body weight, circulating adipokine levels and alterations in gene expression in a set of key adipogenic and lipogenic genes. These findings have implications for understanding the pathogenesis, monitoring the progression and treating the symptoms of HD, as discussed below.
The changes in gene expression in white adipose tissue may contribute to several of the metabolic symptoms that have been noted in late stages of HD in both mice and humans, including weight loss and insulin resistance (22
). Normal white adipose tissue plays an essential role in maintaining metabolic homeostasis, as demonstrated by the fact that genetic deficiency of mature adipose tissue in lipodystrophy leads to insulin resistance, diabetes, increased food intake and hypermetabolic rate (45
). We purposely selected a set of genes that specifically reflects adipocyte differentiation and important functional features of mature adipocytes. Indeed, we found that the gene expression patterns in adipose tissue from HD mice tend to be biphasic; expression of some genes increase at early stages of the disease, but decrease as the disease progresses, in a manner similar to, but less severe than, that observed in mouse models of congenital lipodystrophy (23
). Our gene expression changes were extremely sensitive to disease progression, particularly up to overt spontaneous phenotype development in the fast progressing R6/2 mice. Specifically, there are substantial reductions in the expression of key adipogenic transcription factors such as PPARγ, triglyceride synthesis and storage genes (lipin-1, DGAT, GPDH) and in genes that are characteristic of mature adipocytes (aP2, adipsin). In lipodystrophy, similar alterations in gene expression leads to adipocytes that are less efficient at lipid synthesis and storage, exhibit impaired glucose uptake and have reduced secretion of adipose tissue-derived hormones (22
). Such changes are therefore likely to contribute to the weight loss and insulin resistance that develop in HD. White adipose tissue is uniquely useful for biopsy and it is possible that the changes observed here could be used to track disease progression in HD.
Impaired adipocyte function would also lead to the reduced leptin and adiponectin levels observed in the HD mice. As with adipocyte gene expression, circulating adiponectin and leptin levels are dynamic during the course of the disease, with normal or elevated levels at early time points, and become diminished compared with wild-type mice at later ages. The reduction in adipokine levels could not be accounted for by changes in body weight or fat mass and were most likely related to impaired adipose tissue function and corresponding reduced synthesis of mature adipocyte proteins. The altered function, rather than altered amount, of white adipose could make for more accurate, useful and accessible measurements in the clinical environment. Particularly noteworthy is the 4-fold reduction in leptin levels in CAG140 KI mice from 7 to 22 months of age. Because of their important roles in regulating food intake, energy expenditure and glucose homeostasis, the reduced adipokine levels may contribute to the metabolic dysregulation in HD. Reduced plasma leptin levels have also been reported in human HD patients, although no data exist as to whether the levels diminish within an individual over the course of the disease (46
). These results suggest that circulating levels of leptin and other secreted adipocyte factors normalized to body weight may be a gauge of disease severity and stage, and thus a useful biomarker for monitoring disease progression and/or effects of therapeutic interventions.
Transcriptional dysregulation has been proposed as a key mechanism by which the mutant huntingtin protein exerts its toxic effects in the cell (reviewed in 47
). Transcriptional dysregulation in the brain is an early event in disease pathogenesis and has been shown across different models, including humans and the R6/2 and N171–82Q Tg mouse strains (14
). Altered gene expression levels in peripheral tissues, such as peripheral blood lymphocytes, muscle and pancreatic islets, have also been observed in HD patients and mouse models (14
). Here, we used a set of key genes to determine specifically whether white adipocytes showed dysfunctional gene expression patterns that would alter their function and indeed we found that this was so in two in vivo
models of HD. However, it is not clear whether abnormal gene expression in peripheral tissues is a primary effect of mutant huntingtin expression in those tissues or secondary to systemic changes. Here, we addressed this key question by examining the effect of mutant huntingtin expression in cultured adipocytes. We found that high-level expression of Htt-Q103 in mature adipocytes potently reduced the levels of several genes found to be reduced in vivo
, including PPARγ, lipin-1, aP2 and GPDH. These results strongly indicate that mutant huntingtin within adipocytes has a deleterious effect on gene expression.
Various mechanisms have been proposed for the effects of mutant huntingtin on gene expression in neurons, including interaction of protein fragments/aggregates with components of the core transcription apparatus (53
), with components of the RNA-mediated gene silencing machinery (54
) and with the transcriptional co-activator PGC-1α. PGC-1α has a key role in transcriptional activation of genes in adipose tissue, heart and skeletal muscle and in protection of neurons against oxidative damage (42
). In HD, PGC-1α function is impaired through reduced expression and/or binding to mutant huntingtin (13
), and PGC-1α-deficient mice exhibit spongiform lesions in the striatum, impaired thermogenesis and hyperactivity (32
). Since PGC-1α is known to interact with proteins such as PPARγ2 and lipin-1 (43
) and to co-activate PPARγ2 target genes such as aP2, we investigated whether mutant huntingtin expression in adipocytes affects PGC-1 expression levels and/or activity. We found evidence for reduced PGC-1α and PGC-1β expression levels in adipose tissue of R6/2 Tg mice specifically at a late stage of the disease, and also found that mutant huntingtin interferes with PGC-1α co-activation in cultured adipocytes. These findings identify one potential mechanism by which mutant huntingtin may impair adipocyte gene expression.
HD is a debilitating and fatal disease that exacts its toll through neurodegeneration in the striatum, cerebral cortex and hypothalamus. Our studies demonstrate that expression of mutant huntingtin in white adipose tissue is directly detrimental to tissue function and may influence severity or course of the disease. These observations open the door for novel approaches to study and treat HD, and a detailed study of human patient samples is now warranted. The ability to isolate and culture pre-adipocytes from mouse models provides an additional tool to characterize the molecular pathogenesis of HD. More importantly, the accessibility of small amounts of subcutaneous adipose tissue, and the ability to detect adipose tissue-derived hormones in small volumes of blood, may prove useful in designing and monitoring drug therapies. An important aspect of the current findings is that impaired adipose tissue function is detectable in HD mouse strains at early stages of disease progression. For example, reduced leptin levels were detected in the R6/2 Tg mice at 6 weeks of age, where as hypoactivity, limb clasping, changes in gait and involuntary movements typically do not become evident until 8–12 weeks of age (32
). This raises the possibility that analysis of leptin levels may have diagnostic or prognostic applications, serving as a readily accessible marker to identify patients at risk for poor prognosis or to monitor the progression of disease or response to experimental treatments at early stages of disease progression.
White adipose tissue function is extremely important for glucose homeostasis and energy balance. This suggests that restoring its function in HD patients through peripheral inhibition of mutant huntingtin via RNAi or antisense strategies would have therapeutic value. Furthermore, the activation of adipose tissue PPARγ with thiazolidinedione drugs, which are currently in use for treatment of non-insulin-dependent diabetes, may have beneficial effects on promoting adipocyte maturation, as well as recently reported central (i.e. striatal) benefits (58
). Together, these results support the idea that treatment of peripheral tissues in HD may be valuable in retarding progression of the disease and improving quality of life.