Anti 11 β hsd 2

Approximately 100 mg of rat tissues was homogenized in 1 mL of RIPA buffer and 10 μL of phenylmethylsulfonyl fluoride (PMSF). Total protein was extracted by centrifugation at 15000 rpm for 30 min at 4°C and quantified by enhanced BCA protein assay kit. Protein expression was measured by Western blot analysis. Briefly, protein was resolved on 10% SDS-polyacrylamide gel and transferred to polyvinylidene difluoride microporous membranes (Millipore, Bedford, MA, USA) that were probed with anti-11β HSD I (Abcam, Cambridge, UK, dilution 1 : 800), anti-11 β HSD II (Abcam, Cambridge, UK, dilution 1 : 1000), or GAPDH (Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA, dilution 1 : 500) antibodies. Immunodetection was performed using the ECL detection system according to the manufacturer's instructions. Quantitative densitometric analyses of Western blot images were achieved using ImageJ 1.44p software.

Luteolin (purity > 98%, batch number: XC071225) was purchased from Xi'an Xiaocao Botanical Development Co., Ltd. (Xi'an, China). Sodium carboxymethyl cellulose (CMC-Na), prednisone, prednisolone, and dexamethasone (internal standard for HPLC analysis) were purchased from Sigma-Aldrich Corporation (St. Louis, MO, USA). Methanol of HPLC grade was purchased from TEDIA Company Inc. (Beijing, China). The following items were purchased from the cited commercial sources: anti-11β-HSDI (Abcam, Cambridge, UK, dilution 1 : 800), anti-11β-HSD II (Abcam, Cambridge, UK, dilution 1 : 1000), GAPDH (Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA, dilution 1 : 500) antibodies, goat anti-rabbit IgG (HþL) HRP (BS13278), GAPDH (AP0063), and goat anti-mouse IgG (HþL) HRP (BS12478). All other reagents were of analytical grade.