Xiao Ke Qing (XKQ) granule has been clinically used to treat type 2 diabetes mellitus (T2DM) for 10 years in Chinese traditional medication. However, its mechanisms against hyperglycemia remain poorly understood. This study aims to investigate XKQ mechanisms on diabetes and diabetic liver disease by using the KKAy mice model. Our results indicate that XKQ can significantly reduce food and water intake. XKQ treatment also remarkably decreases both the fasting blood glucose and blood glucose in the oral glucose tolerance test. Additionally, XKQ can significantly decrease the serum alanine aminotransferase level and liver index and can alleviate the fat degeneration in liver tissues. Moreover, XKQ can ameliorate insulin resistance and upregulate the expression of IRS-1, PI3K (p85), p-Akt, and GLUT4 in the skeletal muscle of KKAy mice. XKQ is an effective drug for T2DM by ameliorating insulin resistance and regulating the PI3K/Akt signaling pathway in the skeletal muscle.
Animals ; Blood Glucose ; drug effects ; metabolism ; Diabetes Mellitus, Type 2 ; drug therapy ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Female ; Glucose Tolerance Test ; Glucose Transporter Type 4 ; metabolism ; Hypoglycemic Agents ; pharmacology ; Insulin ; blood ; Insulin Resistance ; Liver ; drug effects ; pathology ; Mice ; Mice, Inbred C57BL ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; drug effects

OBJECTIVE: To construct an effective lentiviral vector for RNA interference (RNAi) with human glucose transporter 3 (GLUT3)gene.
 METHODS: Four pairs of shRNA sequences against different parts of GLUT3-mRNA were separately cloned into the RNAi plasmid vector pLV-shRNA by recombinant DNA technology to construct shRNA expression vectors pLV-shRNA-GLUT3-1, pLV-shRNA-GLUT3-2, pLV-shRNA-GLUT3-3, and pLV-shRNA-GLUT3-4. The vectors were transfected into HeLa cells to detect the effectiveness of GLUT3 gene silencing. One of effective vectors was selected and co-transfected into 293T cells with lentivirus packaging plasmids to obtain packaged lentivirus particles LV-GLUT3. After viral titer determination, U251 glioblastoma cells were infected with LV-GLUT3 at a multiplicity of infection (MOI) of 10. Finally, the expression of GLUT3 protein was detected by Western blot. 
 RESULTS: DNA sequencing demonstrated that the shRNA sequences were successfully inserted into the pLV-shRNA vectors. In HeLa cells, the expression of GLUT3-mRNA was significantly down-regulated by the recombinant vectors compared with negative control. The recombinant lentivirus LV-GLUT3 harvested from 293T cells had a titer of 1.5×10(9) TU/mL. After infection with LV-GLUT3, the expression of GLUT3 protein in U251 glioblastoma cells was down-regulated. 
 CONCLUSION An effective lentiviral shRNA expression vector targeting the GLUT3 gene is successfully constructed and can be used for further study on the functions of GLUT3 gene.
Genetic Vectors ; Glucose Transporter Type 3 ; genetics ; HEK293 Cells ; HeLa Cells ; Humans ; Lentivirus ; Plasmids ; RNA Interference ; RNA, Messenger ; genetics ; RNA, Small Interfering ; genetics ; Transfection

Glucose transporter type 1 deficiency syndrome (GLUT1-DS) is caused by impaired glucose transport across the blood-brain barrier (BBB) and characterized by infantile seizures, developmental delay, acquired microcephaly, spasticity, ataxia, and a low cerebrospinal glucose concentration (hypoglycorrhachia). A diagnosis of GLUT1-DS is biochemically established in neurologically impaired patients with hypoglycorrhachia in the normoglycemia. GLUT1-DS can be confirmed by mutation analysis of the solute carrier family 2 (facilitated glucose transporter), member 1 (SLC2A1) gene or reduced 3-O-methyl-D-glucose uptake into erythrocytes. The patient was a 12-year-old boy born at term. He had experienced seizures from 4 months of age. Electroencephalography (EEG) did not show epileptiform activity. Brain magnetic resonance imaging (MRI) revealed mild diffuse cortical atrophy and ventricular dilatation. Furthermore, he showed developmental delay, mental retardation, and ataxia, which all became more apparent with age progression. For 7 years, he had experienced paroxysmal episodes of atonic behavioral changes that were aggravated before meals or when he became tired. When he was 12 years old, cerebrospinal fluid (CSF) analysis revealed a low glucose concentration in the normal serum glucose and lactate levels. Under the impression of GLUT1-DS, mutation analysis of the SLC2A1 gene by direct sequencing was performed using white blood cells, and c.680-2delA of intron 5 was found. We describe a GLUT1-DS patient with a typical natural history of GLUT1-DS through a long term follow-up visits, with a novel splice site mutation (SLC2A1: c.6802delA).
3-O-Methylglucose ; Ataxia ; Atrophy ; Blood Glucose ; Blood-Brain Barrier ; Brain ; Cerebrospinal Fluid ; Child ; Diagnosis ; Dilatation ; Electroencephalography ; Erythrocytes ; Follow-Up Studies ; Glucose ; Glucose Transport Proteins, Facilitative* ; Glucose Transporter Type 1 ; Humans ; Intellectual Disability ; Introns ; Lactic Acid ; Leukocytes ; Magnetic Resonance Imaging ; Male ; Meals ; Microcephaly ; Muscle Spasticity ; Natural History ; Seizures

BACKGROUNDSuppression of myostatin (MSTN) has been associated with skeletal muscle atrophy and insulin resistance (IR). However, few studies link MSTN suppression by ladder-climbing training (LCT) and IR. Therefore, we intended to identify the correlation with IR between LCT and to analyze the signaling pathways through which MSTN suppression by LCT regulates IR.

METHODSThe rats were randomly assigned to two types of diet: normal pellet diet (NPD, n = 8) and high-fat diet (HFD, n = 16). After 8 weeks, the HFD rats were randomly re-assigned to two groups (n = 8 for each group): HFD sedentary (HFD-S) and high-fat diet ladder-climbing training (HFD-LCT). HFD-LCT rats were assigned to LCT for 8 weeks. Western blotting, immunohistochemistry and enzyme assays were used to measure expression levels and activities of MSTN, GLUT4, PI3K, Akt and Akt-activated targets (mTOR, FoxO1 and GSK-3β).

RESULTSThe LCT significantly improved IR and whole-body insulin sensitivity in HDF-fed rats. MSTN protein levels decreased in matching serum (42%, P = 0.007) and muscle samples (25%, P = 0.035) and its receptor mRNA expression also decreased (16%, P = 0.041) from obese rats after LCT. But the mRNA expression of insulin receptor had no obvious changes in LCT group compared with NPD and HFD-S groups (P = 0.074). The ladder-climbing training significantly enhanced PI3K activity (1.7-fold, P = 0.024) and Akt phosphorylation (83.3%, P = 0.022) in HFD-fed rats, significantly increased GLUT4 protein expression (84.5%, P = 0.036), enhanced phosphorylation of mTOR (4.8-fold, P < 0.001) and inhibited phosphorylation of FoxO1 (57.7%, P = 0.020), but did not affect the phosphorylation of GSK-3β.

CONCLUSIONSThe LCT significantly reduced IR in diet-induced obese rats. MSTN may play an important role in regulating IR and fat accumulation by LCT via PI3K/Akt/mTOR and PI3K/Akt/FoxO1 signaling pathway in HFD-fed rats.

Animals ; Blotting, Western ; Diet, High-Fat ; adverse effects ; Glucose Tolerance Test ; Glucose Transporter Type 4 ; metabolism ; Immunohistochemistry ; Insulin Resistance ; physiology ; Male ; Myostatin ; metabolism ; Obesity ; etiology ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Quadriceps Muscle ; metabolism ; Rats ; Rats, Sprague-Dawley

Modified Si-Miao-San (mSMS) is composed of Rhizoma Coptidis, Cortex Phellodendri, Rhizoma Coptidis Semen Coicis and Atractylodes Rhizome. The prescription is used for the management of diabetes and insulin resistance in the clinic. This study aims to investigate its regulation of glucose disposal in adipocytes. Differentiated 3T3-L1 adipocytes were stimulated with conditioned medium derived from activated macrophages to induce insulin resistance and observed the effects of Mac-CM on insulin-mediated glucose uptake along the insulin receptor substrate-1/PI3K/Akt signaling pathway. Moreover, its regulation of AMPK phosphorylation was also investigated. mSMS enhanced AMPK phosphorylation and promoted basal glucose uptake in adipocytes; mSMS inhibited NF-κB activation by reducing P65 phosphorylation and improved insulin-stimulated IRS-1 tyrosine and Akt phosphorylation, leading to the restoration of insulin-mediated glucose uptake when cells were exposed to inflammatory stimulation. These beneficial effects were diminished in the presence of the AMPK inhibitor compound C. mSMS positively regulated AMPK activity, and this action contributed to improving insulin PI3K signaling by the beneficial regulation of IRS-1 function through inhibition of inflammation in adipocytes.
3T3-L1 Cells ; Adenosine Monophosphate ; metabolism ; Adenylate Kinase ; metabolism ; Adipocytes ; drug effects ; metabolism ; Animals ; Atractylodes ; Coix ; Coptis ; Diabetes Mellitus ; drug therapy ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Glucose ; metabolism ; Glucose Transporter Type 4 ; metabolism ; Inflammation ; metabolism ; prevention & control ; Insulin ; metabolism ; Insulin Receptor Substrate Proteins ; metabolism ; Insulin Resistance ; Mice ; NF-kappa B ; metabolism ; Phellodendron ; Phosphatidylinositol 3-Kinases ; metabolism ; Phosphorylation ; Phytotherapy ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction

OBJECTIVETo investigate the effect of Jiaotai Pill (, JTP) at different constitutional proportions on insulin signaling through phosphatidylinositol 3-kinase (PI3K) pathway in the skeletal muscle of diabetic rats.

METHODSThe rat model of type 2 diabetes mellitus (T2DM) was established by intravenous injection of a small dose of streptozotoein plus high fat diet feeding. JTP at the same dosage of cinnamon and the increasing dosage of Coptis chinensis was administered to diabetic rats for nine weeks respectively. Plasma glucose and insulin levels were assayed. The expressions of proteins were determined by Western blot method.

RESULTSAll the three formulations of JTP decreased plasma glucose and fasting insulin levels as well as increased the protein expressions of insulin receptor β (InsRβ) subunit, insulin receptor substrate-1 (IRS-1), PI3K p85 subunit and glucose transporter 4 (GLUT4) in skeletal muscle. Meanwhile, JTP increased the tyrosine phosphorylation of InsRβ subunit and IRS-1, and reduced the serine phosphorylation of IRS-1 in skeletal muscle. Interestingly, the effect of JTP on improving insulin sensitivity was not dose-dependent. In contrast, JTP containing the least amount of Coptis chinensis exhibited the best effect.

CONCLUSIONJTP at different constitutional proportions attenuates the development of diabetes in a rat model of T2DM. The mechanism might be associated with enhancing insulin signaling through PI3K pathway in the skeletal muscle.

Animals ; Body Weight ; drug effects ; Diabetes Mellitus, Experimental ; drug therapy ; enzymology ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Glucose Tolerance Test ; Glucose Transporter Type 4 ; metabolism ; Homeostasis ; drug effects ; Insulin ; metabolism ; Insulin Receptor Substrate Proteins ; metabolism ; Insulin Resistance ; Male ; Muscle, Skeletal ; drug effects ; enzymology ; metabolism ; pathology ; Phosphatidylinositol 3-Kinases ; metabolism ; Phosphorylation ; Phosphotyrosine ; metabolism ; Protein Subunits ; metabolism ; Rats ; Rats, Wistar ; Receptor, Insulin ; metabolism ; Signal Transduction ; drug effects

OBJECTIVETo study the effect of total saponins from Entada phaseoloides (TSEP) on islet morphology and skeletal muscle PI3K pathway-related protein expression of type 2 diabetic rats.

METHODType 2 diabetic rats were induced by high-fat diet and low-dose streptozotocin and then randomly divided into 5 groups, i.e. the normal control, the model group, the positive control drug (200 mg x kg(-1) metformin), the low-dose TSEP (25 mg x kg(-1)) group and the high-dose TSEP (50 mg x kg(-1)). Three weeks later, the islet morphology of rat pancreas were observed by HE staining, and protein expressions of insulin receptor substrate-1 (IRS-1), phosphatidylinositol 3-kinase (PI3K), protein tyrosine phosphatase-1B (PTP-1 B) and glucose transporter 4 (GLUT4) in rat skeletal muscle were detected by Western blot.

RESULTCompared with the modal group, TSEP administration groups showed relatively normal structures, clear pancreatic cells and intact capsula structures in pancreatic tissue pathological sections, with the number of pancreatic islets close to the normal control group. Meanwhile, above TSEP administration groups showed increased IRS-1, PI3K and GLUT4 protein expressions in their skeletal muscle tissues and decreased PTP-1B protein expression compared with the model group.

CONCLUSIONTSEP has an effect on protecting pancreatic tissues of type 2 diabetic rats and intervening in abnormal expression of proteins in skeletal muscle tissues.

Animals ; Diabetes Mellitus, Type 2 ; drug therapy ; Fabaceae ; chemistry ; Glucose Transporter Type 4 ; analysis ; Islets of Langerhans ; drug effects ; pathology ; Male ; Muscle, Skeletal ; drug effects ; pathology ; Phosphatidylinositol 3-Kinases ; analysis ; Rats ; Rats, Sprague-Dawley ; Saponins ; therapeutic use

This study is to investigate the effect of low doses of insulin (1 u x kg(-1)) and selenium (180 microg x kg(-1)) in combination on general physiological parameters and insulin signal molecules in cardiac muscle of STZ-induced diabetic rats. The levels of blood glucose were estimated using One Touch SureStep Blood Glucose meter. HbA1c levels were estimated using microcolumn assay. TG and TC were estimated using enzymatic assay. The levels of PI3K and GLUT4 in cardiac muscle were examined by immunoblotting and immunohistochemistry. The result showed that insulin in combination with selenium could significantly lower blood glucose and blood lipid levels and markedly restored the PI3K and GLUT4 levels in cardiac muscle. It could be concluded that there was cooperation between insulin and selenium, and that treatment of diabetic rats with combined doses of insulin and selenium increased cardiac glucose uptake by upregulating the level of PI3K-mediated GLUT4 in cardiac muscle, eventually ameliorating myocardial dysfunction.
Animals ; Antioxidants ; pharmacology ; Blood Glucose ; metabolism ; Cholesterol ; blood ; Diabetes Mellitus, Experimental ; blood ; metabolism ; Drug Therapy, Combination ; Glucose Transporter Type 4 ; metabolism ; Glycated Hemoglobin A ; metabolism ; Hypoglycemic Agents ; pharmacology ; Insulin ; pharmacology ; Male ; Myocardium ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Selenium ; pharmacology ; Signal Transduction ; drug effects ; Triglycerides ; blood

OBJECTIVETo investigate the effect of exercise on the expressions of phosphatidylinositol 3 kinase (PI3K) and protein kinase B (PKB) phosphorylation, protein and glucose transport proteins (GLUT4) at both the protein and mRNA levels in the skeletal muscles of type 2 diabetic rats.

METHODSThirty male SD rats were randomly divided into control group with normal diet feeding, diabetic group and diabetic exercise group with high-fat diet feeding. After 8 weeks of the high-fat diet, each rat received an intraperitoneal injection of streptozotocin (STZ, 30 mg/kg). Three weeks after the injection, the rats were rated for the presence of diabetes, and the rats in the exercise groups took swimming training for 4 weeks; all the groups maintained their assigned diets. The gastrocnemius of all the rats were dissected 48 h after the last training session. Western blotting was applied to detect the phosphorylation and protein expression of PI3K and PKB and the protein expression of GLUT4. The expression of GLUT4 mRNA was determined by RT-PCR.

RESULTSCompared with the diabetic group, the diabetic rats in the exercise group showed significantly increased protein expression and phosphorylation of PKB (P<0.05) and elevated GLUT4 protein and mRNA expressions in the skeletal muscles (P<0.01, P<0.05).

CONCLUSIONExercise training can modulates insulin signal transduction through the protein expression and phosphorylation of the protein kinase to promote glucose uptake in the skeletal muscle of type 2 diabetic rats.

Animals ; Diabetes Mellitus, Experimental ; metabolism ; Diabetes Mellitus, Type 2 ; metabolism ; Glucose Transporter Type 4 ; metabolism ; Insulin ; metabolism ; Male ; Muscle, Skeletal ; metabolism ; Phosphatidylinositol 3-Kinases ; genetics ; metabolism ; Phosphorylation ; Physical Conditioning, Animal ; Proto-Oncogene Proteins c-akt ; genetics ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

OBJECTIVETo observe the changes in the expression of glucose transporter-3 (GLUT3) in the cerebral cortex of rats during aging and investigate the role of GLUT3 in the aging process of the nervous system.

METHODSThe cerebral tissues were collected from rats of 3, 18, 24, and 30 months old (10 in each age group), and the expression of GLUT3 in the cerebral cortex was detected by immunohistochemistry.

RESULTSUnder optical microscope, GLUT3-positive cells were found in every group. Within the age range of 3 to 8 months, GLUT3-positive cells increased significantly with age (P<0.01), but at 24-30 months of age, the number of GLUT3-positive cells reduced significant with age (P<0.01).

CONCLUSIONThe expression changes of GLUT3 ir the cerebral cortex of rats during aging indicate that GLUT3 plays an important role in the maturation and aging of the nervous system.

Aging ; Animals ; Brain ; metabolism ; Cerebral Cortex ; metabolism ; Glucose Transporter Type 3 ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley