Masuno, H, T Kidani, K Sekiya, K Sakayama, T Shiosaka, H Yamamoto and K Honda. 2002. Bisphenol A in combination with insulin can accelerate the conversion of 3T3-L1 fibroblasts to adipocytes.  Journal of Lipid Research 3:676-684.

other work relevant to BPA and weight homeostasis

Masuno et al. report that low levels of bisphenol A (BPA) combined with insulin can affect the formation of adipocytes (fat cells). This finding provides a challenge to current interpretations of the causes of the obesity epidemic now underway in the US and many other countries around the world. It raises the possibility that contaminant disruption of weight regulation may be a contributing factor.

Obesity is about body fat, which is stored in the body in fat cells called adipocytes. At the cellular level, obesity is caused either because of an increase in the number of fat cells (called fat cell hyperplasia) or an enlargement of fat cells with each cell carrying greater amounts of fat (fat cell hypertrophy), or both.

This paper suggests that exposure to BPA may increase body fat via both fat cell hyperplasia and hypertrophy: In these tissue culture experiments it both triggers the conversion of cells to adipocytes and increases the quantity of stored fat.

While much work remains to be done to determine whether this physiological impact of BPA is relevant to weight regulation in people, what is certain is that human exposure to BPA is widespread through its use in dental procedures, in food storage and in polycarbonate plastic. For example, polycarbonate baby bottles heated by microwave leach BPA into milk fed to infants. The possibility that BPA may be contributing to the world-wide epidemic of obesity makes aggressive study of Masuno et al's hypothesis imperative.

What did they do? Masuno et al. carried out their experiments in a cell tissue culture line of mouse cells (called 3T3-L1 cells) which have become a model system for exploring some of the factors that affect the conversion rate to adipocytes (fat cells). This conversion rate is now understood to be a key step in developing obesity. The more rapid and extensive the conversion, the more likely is obesity.

Over the past 25 years, work with 3T3-L1 cells and similar cell lines has allowed a detailed characterization of the steps leading to adipocyte formation, called adipogenesis. During adipogenesis, pre-adipocyte cells go through a series of stages that involve characteristic changes in shape and in the distribution and amount of fat within them. Characteristic genes are also expressed at different stages, and the presence of these genes can be used as biochemical markers for adipogenesis.

In their experiments Masuno et al. combined different levels of insulin and bisphenol A in a series of tests to tease apart the impact on cellular differentiation of the compounds individually and together. Their methods take advantage of prior work that has characterized the morphological and biochemical markers that accompany adipocyte formation. They used a parallel series of experiments with compounds known to stimulate cell differentiation as positive controls.

They were particularly interested in the impact of exposure on the activity levels of two enzymes that had been established by earlier studies to be indicators of conversion of pre-adipocytes to adipocytes, LPL (lipoprotein lipase) and GDPH (glycerol-3-phosphate dehydrogenase), and on the accumulation of fat (triacylglycerol, or TG) within the cells. Typically, LPL appears before GDPH activity, which is acommpanied by TG build-up.

What did they find? Masuno et al. first established a dose-response relationship between BPA and LPL activity, in the presence of insulin. Remember that the presence of LPL is a marker for reaching the final stages of adipogenesis.

The level of LPL activity in 3T3-L1 cells rises with increasing BPA exposure in the presence of insulin.

They then performed an additional series of experiments using 20 µg/ml of BPA, comparing cell parameters for cells with different regimes of BPA and insulin treatment. They confirmed that BPA triggered the conversion of pre-adipocytes into adipocytes.

This experiment ran for 11 days. During the first 2 days, cells were either exposed, or not exposed (control group) to BPA. During the latter 9 days, they were exposed only to insulin. Wasuno et al. refer to these two different phases as the "trigger" (BPA) and the "treatment" (insulin). The BPA-triggered cells showed a 60% increase in LPL activity (compared to controls), a 500% increase in GPDH, and a 150% increase in fat content. Microscopic examination of the cells showed that the lipid droplets within the cells had coalesced and become larger, consistent with the what happens during adipogenesis. So the BPA exposure triggered the formation of fat cells.

They repeated a variation on this experiment, and determined that BPA not only triggers the conversion, it stimulates the conversion process once triggering has occurred. They did this by leaving BPA in the trigger and adding BPA to the treatment. Thus during the first 2 days, all cells were exposed to BPA (comparable to the experimental group in the prior experiment). During the latter 9, one group was exposed only to insulin (control) the other to both BPA and insulin. The combination of insulin and BPA caused a 1,300% increase in fat levels. GPDH activity was 3.3 times higher than cultures treated only with insulin, and the percentage of lipid-positive cells in the cultures was 83%. These results are much higher than the effect of simply using BPA only as a trigger (e.g., 1,300% vs. 150% increase in fat). Hence Masuno et al. concluded that BPA in the treatment phase stimulated the conversion process more effectively than insulin alone.

Masuno et al. report that these chemical and morphological impacts are very similar to positive control experiments with compounds known to stimulate adipogenesis (dexamethasone [DEX] and 1-methyl-3-isobutylxanthine [MIX]). This is consistent with their interpretation that BPA also stimulates adipogenesis.

Masuno et al. looked more closely at the behavior of the 3T3-L1 cells over time in response to a BPA trigger and BPA-insulin treatment, comparing them to the positive controls (DEX, MIX and insulin in the trigger and insulin in the treatment). DNA content of the BPA-insulin group increased, although not as much as the positive controls. Changes in fat content and LPL activity were similar between the BPA-insulin group and positive controls, again consistent with the interpretation that BPA is stimulating adipogenesis.

What does it mean? Heretofore, the prevailing wisdom about the causes of the world-wide obesity epidemic have focused almost exclusively on life style issues: consumption of junk food, decreases in exercise, etc. These remain plausible interpretations, and almost certainly are contributing factors. But this research report by Masuno et al. opens up an entirely new front: the disruption of weight regulation (weight homeostasis) by hormone-disrupting contaminants. In this case, BPA triggers and then stimulates two of the key biological mechanisms underlying obesity: it increases the number of adipocytes (fat cell hyperplasia), and enhances their fat storage (fat cell hypertrophy).

Many steps lie between these initial observations and confirmation that contaminant disruption of weight homeostasis contributes to obesity in people, although there are a few hints elsewhere in the scientific literature that Masuno's mechanism may be relevant. For example, Howdeshell et al. had noted in an earlier paper that BPA-treated mice matured heavier than controls, but they had no insight as to mechanism.

Given the enormous health consequences of obesity and the diseases to which it is linked, like diabetes, pursuing this line of research now becomes imperative. What is intriguing is that as further research confirms the plausible involvement of endocrine disrupting compounds in interfering with weight homeostasis, then there are simple and attainable interventions that can introduced on a precautionary basis: reducing exposures.