title: 4-Hydroxyisoleucine improves hepatic insulin resistance by restoring glycogen synthesis in vitro.
abstract: OBJECTIVE
  Hydroxyisoleucine (4-HIL), derived from fenugreek seeds, improves insulin resistance in 3T3-L1 adipocytes and L6 myotubes. However, the effects of 4-HIL on liver glycogen synthesis and hepatic insulin resistance have not been described. The aim of this study was to investigate the effects of 4-HIL on glycogen synthesis in a tumor necrosis factor-α (TNF-α)-induced insulin resistance model using HepG2 cells.
  
  
  METHODS
  HepG2 cells were divided into eight groups: control; TNF-α; and 5, 10, or 20 μM 4-HIL without or with TNF-α. Glycogen and protein expression were evaluated using a glycogen assay kit and western blotting, respectively.
  
  
  RESULTS
  Glycogen levels did not differ between the 4-HIL groups and control (P>0.05), but were decreased significantly in the TNF-α group (P<0.05), indicating the establishment of insulin-resistant HepG2 cells. Adding 20 μM 4-HIL to TNF-α-treated cells increased glycogen levels (P<0.05). Relative to the control group, the P-IRS-1/IRS-1 and P-JNK/JNK ratios were increased (P<0.001) in the TNF-α group, whereas the P-AKT/AKT and P-GSK/GSK ratios were decreased (P<0.001). When 20 μM 4-HIL was added to TNF-α-treated cells, the P-IRS-1/IRS-1 and P-JNK/JNK ratios decreased (P<0.001 and P<0.05, respectively), whereas the P-AKT/AKT and P-GSK/GSK ratios increased (P<0.05 and P<0.001).
  
  
  CONCLUSIONS
  4-HIL directly or indirectly reversed TNF-α reduced glycogen levels by inhibiting JNK and IRS-1 (Ser(307)) phosphorylation and increasing AKT (Ser(473)) and GSK-3 phosphorylation. These findings demonstrate that 4-HIL modulates hepatic insulin resistance at the molecular level, and suggest that it is a novel potential therapeutic agent for the treatment of insulin resistance in patients with diabetes.
authors: Department of Integrated Chinese Traditional and Western Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Furong
authors: F
authors: Lu
authors: Department of Endocrinology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Qin
authors: Q
authors: Cai
authors: Department of Endocrinology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Mohammad Ishraq
authors: MI
authors: Zafar
authors: Department of Neurosurgery, Wuhan Central Hospital No. 26, Shengli Street, Wuhan 430014, China.
authors: Lun
authors: L
authors: Cai
authors: Department of Endocrinology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Wen
authors: W
authors: Du
authors: Department of Endocrinology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Liumeng
authors: L
authors: Jian
authors: Department of Endocrinology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Liping
authors: L
authors: Li
authors: Department of Endocrinology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology 1277, Jiefang Avenue, Wuhan 430022, China.
authors: Feng
authors: F
authors: Gao
keywords: 3T3-L1 adipocytes; Fenugreek; JNK signaling pathway; L6 myotubes; insulin resistance