We have previously shown that CCN2, also known as connective tissue growth factor, potently inhibits differentiation of pre-adipocyes into adipocytes in vitro. The current study aimed to determine if CCN2 may divert the white adipose cell differentiation programme into a beige fat profile, by uncoupling protein-1 (UCP-1) induction.
NIH3T3-L1 cells were treated with rhCCN2 at the pre-adipocyte stage, and also in mature differentiated adipocytes induced by standard methods at day 10 of the differentiation programme. Changes in UCP-1 mRNA by qPCR and protein by Western immunoblot, and related factors affecting thermogenesis and energy turnover, such as PGC-1α and heat shock proteins (HSPs), were examined. Subsequently, in a high fat fed (65% calories as fat) mouse model at 20 weeks of feeding, relationships of CCN2 mRNA with UCP-1 mRNA species were studied in fat tissue depots.
Results showed that in NIH3T3-L1 cells, UCP-1 mRNA and protein were induced by rhCCN2 addition, which was most potent and rapid in differentiated adipocytes (5.5 fold mRNA and 5 fold protein induction at 24 h; each P<0.05 vs no CCN2 addition), rather than pre-adipocytes (1.2 and 1.6 fold induction at 24 and 48 h respectively for mRNA and 2.8 fold for protein at 48 h; each P<0.05 vs no CCN2 addition). In contrast, rhCCN2 prevented mRNA induction otherwise seen in differentiated adipocytes of the energy regulating PGC-1α transcriptional coactivator. Amongst HSPs in adipocytes, HSP-70 mRNA was specifically induced 2.7 fold (P<0.05) by rhCCN2, but HSP-60 and HSP-90 were unaffected. Subsequently, in high fat fed obese mice, CCN2 mRNA levels correlated strongly and positively with UCP-1 mRNA (but not PGC-1α) in both central (epididymal) and subcutaneous fat tissue (r=0.94, P=0.003; and r =0.76, P=0.01, respectively).
In summary, this novel data shows that CCN2 may link high fat feeding to induction of beige fat, which is an hypothesis now requiring direct in vivo testing.