Whether obesity accelerates or suppresses autophagy in adipose tissue is still debatable. To clarify dysregulation of autophagy and its role in pathologies of obese adipose tissue, we focused on lysosomal function, protease maturation and activity, both in vivo and in vitro. First, we showed that autophagosome formation was accelerated, but autophagic clearance was impaired in obese adipose tissue. We also found protein and activity levels of CTSL (cathepsin L) were suppressed in obese adipose tissue, while the activity of CTSB (cathepsin B) was significantly enhanced. Moreover, cellular senescence and inflammasomes were activated in obese adipose tissue. In 3T3L1 adipocytes, downregulation of CTSL deteriorated autophagic clearance, upregulated expression of CTSB, promoted cellular senescence and activated inflammasomes. Upregulation of CTSB promoted additional activation of inflammasomes. Therefore, we suggest lysosomal dysfunction observed in obese adipose tissue leads to lower autophagic clearance, resulting in autophagosome accumulation. Simultaneously, lysosomal abnormalities, including deteriorated CTSL function and compensatory activation of CTSB, caused cellular senescence and inflammasome activation. Our findings strongly suggest lysosomal dysfunction is involved in early pathologies of obese adipose tissue.

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The total estrogenic activity of the wastewater from a swine farm in Japan was quantitatively characterized, and the compounds responsible for the estrogenic activity were identified and quantified. The wastewater treatment process consisted of a series of an up-flow anaerobic sludge blanket (UASB) and a trickling filter. Samples were collected at each treatment step, and the total estrogenic activity was determined by use of an in vitro gene expression assay (MVLN; MCF-7 human breast cancer cell stably transfected with the pVit-tk-LUC receptor plasmid). Individual estrogenic compounds were identified and quantified using liquid chromatography-mass spectrometry (LC/MS) and liquid chromatography-tandem mass spectrometry (LC/ MS/MS). To further identify the compounds contributing to the estrogenic activity in the wastewater, the sample extracts were fractionated into 12 fractions (fractions 1-12) by HPLC. The rate of removal of estrogenic activity between the effluent and the influent was greater than 97%. The trickling filter removed the majority of the estrogenic activity. The removal rates of specific estrogenic compounds ranged from 44 to 99%. Estrogenic activity was detected mainly in the fractions containing estrone (El), 17beta-estradiol (betaE2), 17alpha-estradiol (alpha E2), estriol (E3), bisphenol A (alphaPA), and equol (EQ0). The ratios of betaE2-EQc (betaE2 equivalents derived from chemical analysis) to betaE2-EQB (betaE2 equivalent derived from bioassay) in the 12 fractions collectively were contributed by El (17-30%), betaE2 (23-30%), acE2 (<1%), E3 (1-2%), BPA (<1%), and EQO (2-3%) in the influent and El (16-37%), PE2 (<1-7%), alphaE2 (<1%), E3 (<1-3%), BPA (<1%), and EQO (<1%) in the effluent. The compounds responsible for most of the estrogenic activity measured in the bioassay were natural estrogens such as El and betaE2.