Characterization of the male ApcMin/+ mouse as a hypogonadism model related to cancer cachexia

Abstract

Cancer cachexia, the unintentional loss of lean body mass, is associated with decreased quality of life and poor patient survival. Hypogonadism, involving a reduction in circulating testosterone, is associated with the cachectic condition. At this time there is a very limited understanding of the role of hypogonadism in cancer cachexia progression. This gap in our knowledge is related to a lack of functional hypogonadal models associated with cancer cachexia. The Apc(Min/+) mouse is an established colorectal cancer model that develops an IL-6 dependent cachexia which is physiologically related to human disease due to the gradual progression of tumor development and cachexia. The purpose of this study was to assess the utility of the Apc(Min/+) mouse for the examination of hypogonadism during cancer cachexia and to investigate if IL-6 has a role in this process. We report that Apc(Min/+) mice that are weight stable have comparable testosterone levels and gonad size compared to wild type mice. Cachectic Apc(Min/+) mice exhibit a reduction in circulating testosterone and gonad size, which has a significant association with the degree of muscle mass and functional strength loss. Circulating testosterone levels were also significantly associated with the suppression of myofibrillar protein synthesis. Skeletal muscle and testes androgen receptor expression were decreased with severe cachexia. Although testes STAT3 phosphorylation increased with severe cachexia, systemic IL-6 over-expression for 2 weeks was not sufficient to reduce either testes weight or circulating testosterone. Inhibition of systemic IL-6 signaling by an IL-6 receptor antibody to Apc(Min/+) mice that had already initiated weight loss was sufficient to attenuate a reduction in testes size and circulating testosterone. In summary, the Apc(Min/+) mouse becomes hypogonadal with the progression of cachexia severity and elevated circulating IL-6 levels may have a role in the development of hypogonadism during cancer cachexia.

Keywords: Cancer cachexia; Hypogonadism; Muscle; Testosterone.

Fig. 1.. The reduction in circulating testosterone correlates with several parameters of cachexia in the Apc^Min/+ mouse.

(A) Circulating testosterone throughout the progression of cachexia. (B) Upper representative Western blot of the androgen receptor in the gastrocnemius. Lower quantified androgen receptor expression throughout the progression of cachexia in the gastrocnemius muscle. (C) Correlation between circulating testosterone and gastrocnemius weight (gastroc). (D) Correlation between circulating testosterone and myofibrillar protein synthesis. Values are means ± SE. *Signifies difference from wild-type mice. # Signifies difference from mice with 6–19% body weight loss.

Fig. 2.. Gonadal atrophy during cachexia.

(A) Testes weight throughout the progression of cachexia. Correlation between testes weight and (B) testosterone, (C) gastrocnemius muscle (gastroc) weight and (D) grip strength. Values are means ± SE. *Signifies difference from wild-type mice. # Signifies difference from mice with 6–19% body weight loss.

Fig. 3.

(A) Representative Western blot of the androgen receptor, Bax, phosphorylated and total STAT3 in the testes. Quantified (B) androgen receptor, (C) Bax and (D) Ratio of phospho and total STAT3 expression throughout the progression of cachexia. Values are means ± SE. *Signifies difference from wild-type mice. # Signifies difference from mice with 6–19% body weight loss.

Fig. 4.. Acute IL-6 over-expression does not induce hypogonadism.

(A) Circulating testosterone and (B) testes weight in mice over-expressing systemic IL-6 or control vector. Values are means ± SE.

Fig. 5.. Inhibition of IL-6 signaling attenuates the development of hypogonadism in the Apc^Min/+ mouse.

(A) Circulating testosterone and (B) testes weight in mice treated with an IL-6 receptor antibody (IL-6Rec Ab) or PBS control. Values are means ± SE. *Signifies difference from PBS treated wild-type mice. & Signifies difference from PBS treated Apc^Min/+ mice.

Fig. 6.. Summary of inflammatory, systemic and tissue specific changes during the progression of cachexia in Apc^Min/+ mouse.

The initiation of cachexia is associated with an increase in circulating IL-6, reduction in muscle IGF-1 expression, muscle protein synthesis and increased ubiquitin dependent proteolysis. These changes were independent of alterations in gonadal size or function. During severe cachexia, the onset of hypogonadism corresponds with a further increase in circulating IL-6 and the rise in endotoxemia. Muscle protein synthesis and IGF-1 expression are suppressed further while AMPK activation is increased. As for protein degradation, severe cachexia brings a further increase in ubiquitin-dependent proteolytic activity and the activation of autophagy-related proteolysis. Arrows indicate direction of change. Darker arrows indicate a further increase or decrease during the transition to severe cachexia.