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ObjectiveTo observe the effect of modified Gegen Qinliantang on the expression levels of proteins related to the farnesoid X receptor/small heterodimer partner/peroxisome proliferator-activated receptor α （FXR/SHP/PPARα） signaling pathway in the liver tissue of db/db model mice with type 2 diabetes mellitus （T2DM） and explore the underlying mechanism of action of modified Gegen Qinliantang. MethodThirty db/db mice were randomly divided into model group， metformin group （0.2 g·kg-1）， and high-， medium-， and low-dose modified Gegen Qinliantang groups （31.9， 19.1， 6.4 g·kg-1）， with 6 mice in each group. An additional six m/m mice were assigned to the blank group. Respective drugs were administered via oral gavage for 12 weeks. Mouse body weight， fasting blood glucose （FBG）， total cholesterol （TC）， triglyceride （TG）， high-density lipoprotein cholesterol （HDL-C）， and low-density lipoprotein cholesterol （LDL-C） levels were measured. Oil red O staining was used to observe hepatic lipid accumulation and periodic acid-schiff （PAS） staining was used to assess hepatic glycogen deposition. Ammonium ferric sulfate staining was used to observe cholesterol deposition in intestinal tissues. Western blot was employed to detect the expression of FXR， cholesterol 7α-hydroxylase （CYP7A1）， SHP， and PPARα proteins in liver tissues， and enzyme-linked immunosorbent assay （ELISA） was used to measure serum free fatty acid （FFA） levels. ResultAt the end of the treatment， compared with the blank group， the model group exhibited significant increases in mouse body weight， FBG， FFA， TC， TG， and LDL-C levels （P<0.01）， along with significant hepatic lipid droplets， reduced hepatic glycogen， noticeable cholesterol accumulation in intestinal tissues， significantly decreased expression of FXR， SHP， PPARα proteins， and significantly increased expression of CYP7A1 protein in liver tissues （P<0.01）. Compared with the model group， the metformin group and the high- and medium-dose modified Gegen Qinliantang groups demonstrated significant reductions in mouse body weight， FBG， FFA， TC， TG， LDL-C levels （P<0.05， P<0.01）， significant increases in HDL-C levels （P<0.05， P<0.01）， decreased hepatic lipid accumulation， increased hepatic glycogen， reduced intestinal cholesterol accumulation， significantly increased expression of FXR， SHP， PPARα proteins， and significantly decreased expression of CYP7A1 protein in liver tissues （P<0.01）. ConclusionModified Gegen Qinliantang may regulate the FXR/SHP/PPARα signaling pathway to suppress FFA levels and improve lipid metabolism in T2DM mice.
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Objective To investigate the therapeutic effects of emodin on acute cholestatic hepatitis and mec-hanism thereof. Methods Fifty Sprague - Dawley (SD)rats were randomly divided into 5 groups and were treated with emodin,ursodeoxycholic acid,dexamethasone,or normal saline respectively for 4 days. On the fifth day gastric perfusion of alpha - naphthylisothiocyanate(ANIT)was performed to establish models of choiestatic hepatitis. Four to six hours after the establishment of model the above mentioned agents were given continuously. Forty - eight hours after the model establishment blood samples were collected from abdominal aorta to examine the total bilirubin(TB),direct bilirubin (DB),alanine aminotransferase(ALT),total bile acid(TBA),aspartate aminotransferase(AST),alkaline phosphatase (ALP),gamma glutamine transferase(GGT). Specimen of liver was collected to undergo pathological examination. Real - time PCR was used to detect the mRNA expression of farnesoid X receptor(FXR),small heterodimer partner (SHP ),bile salt export pump (BSEP ),uridine diphosphate glucuronosyltransferase 2 family polypeptide B4 (UGT2B4). Results The serum levels of total bilirubin (TB),direct bilirubin (DB),alanine aminotransferase (ALT),total bile acids (TBA),aspartate aminotransferase (AST),and alkaline phosphatase (ALP)of the model group were respectively (68. 1 ± 26. 1)μmol/ L,(46. 3 ± 20. 1)μmol/ L,(483 ± 228)U/ L,(159. 1 ± 57. 9)μmol/L,(2. 0 ± 0. 5)U/ L,(996 ± 382)U/ L,(324 ± 120)U/ L. The levels of TB,DB,ALT,TBA,AST,ALP of the emodin group were respectively (15. 0 ± 8. 7)μmol/ L,(10. 8 ± 3. 9)μmol/ L,(147 ± 71)U/ L,(60. 1 ± 22. 7)μmol/ L, (295 ± 104)U/ L,(222 ± 59)U/ L,and were all significantly lower than those of the model group (all P < 0. 05). The levels of TB,DB,ALT,TBA,AST,GGT,ALP of the emodin group were all significantly lower than those of the ursode-oxycholic acid group (all P < 0. 05). The levels of TB,DB,ALT,TBA,GGT,AST were all significantly lower than those of the dexamethasone group (all P < 0. 01). The expression levels of FXR,SHP,BSEP,UGT2B4 mRNA in the emodin group (1. 087 ± 0. 285,0. 892 ± 0. 390,0. 902 ± 0. 149,1. 785 ± 0. 403)were all significantly higher than those of the model group (0. 152 ±0. 088,0. 559 ±0. 194,0. 561 ±0. 123,0. 177 ±0. 039,all P <0. 05). Conclusions By decreasing the levels of TB,DB,ALT,TBA,AST,ALP and reducing pathological changes,emodin has a protective effect on cholestatic hepatitis. It has better effects than ursodeoxycholic acid and dexamethasone. These effects may be due to promoting FXR,SHP,BSEP and UGT2B4 expression.
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Objective To investigate the effects of inducible hypothermia on expressions of peroxisome proliferator activated receptor gamma coactivator-1 (PGC-1) and small heterodimer partner (SHP) in rat model of hemorrhagic shock.Methods SD rats were randomly (random number) divided into three groups:control (C),normothermia (N) and hypothermia (H).Exsanguination was carried out in rats by continuously drawing out venous blood (25 mL/kg) over 15 minutes to establish hemorrhagic shock model.Then,rectal temperatures of rats were maintained by body surface cooling to 32℃ in H group and by body surface warming to 38℃ in N group,respectively.After a shock period of 60 minutes,rats received the infusion of whole blood of their own and lactated Ringer's solution (1 ∶ 2) treatment for 60 min.Rats were warmed to 38℃ by body surface warming and monitored for 3 h after resuscitation.Hematocrit (Hct),base excess (BE),lactate (Lac),and glucose (Glu) were recorded before modeling and after different lengths of hemorrhagic shock period (HSP).The expressions of PGC-1 mRNA and SHP mRNA and the levels of their protein in liver were tested by real-time reverse transcription polymerase chain reaction (qRT-PCR) and western blotting,respectively.Results The H group had lower lactate levels and higher BElevels than the N group [(6.3±2.1) vs.(10.4±1.5) and (-4.7±2.5) vs.(-9.0±3.2)] (P＜ 0.05).At 72 hours after modeling,there were four survivors in the N group and seven survivors in the H group (P ＜ 0.05,Log Rank).The expressions of PGC-1 mRNA and SHP mRNA increased in N group.Hypothermia resuscitation down-regulated PGC-1 mRNA expression,meanwhile,increased expression of SHP mRNA.Both Hypothermia and Normothermia resuscitation increased SHP protein levels,but decreased PGC-1 protein levels.Conclusions Inducible hypothermia ameliorated acidosis and energy metabolism imbalance through adaptive regulation in PGC-1 and SHP.
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BACKGROUND: Expression of hepatic cholesterol 7alpha-hydroxylase (CYP7A1) is negatively regulated by orphan nuclear receptor small heterodimer partner (SHP). In this study, we aimed to find whether thyroid hormone regulates SHP expression by modulating the transcriptional activities of liver receptor homolog-1 (LRH-1). METHODS: We injected thyroid hormone (triiodothyronine, T3) to C57BL/6J wild type. RNA was isolated from mouse liver and used for microarray analysis and quantitative real-time polymerase chain reaction (PCR). Human hepatoma cell and primary hepatocytes from mouse liver were used to confirm the effect of T3 in vitro. Promoter assay and electrophoretic mobility-shift assay (EMSA) were also performed using human hepatoma cell line RESULTS: Initial microarray results indicated that SHP expression is markedly decreased in livers of T3 treated mice. We confirmed that T3 repressed SHP expression in the liver of mice as well as in mouse primary hepatocytes and human hepatoma cells by real-time PCR analysis. LRH-1 increased the promoter activity of SHP; however, this increased activity was markedly decreased after thyroid hormone receptor beta/retinoid X receptor alpha/T3 administration. EMSA revealed that T3 inhibits specific LRH-1 DNA binding. CONCLUSION: We found that thyroid hormone regulates the expression of SHP mRNA through interference with the transcription factor, LRH-1.
Subject(s)
Animals , Child , Humans , Mice , Bile Acids and Salts , Carcinoma, Hepatocellular , Cell Line , Child, Orphaned , Cholesterol , Cholesterol 7-alpha-Hydroxylase , DNA , Hepatocytes , Liver , Microarray Analysis , Real-Time Polymerase Chain Reaction , Receptors, Thyroid Hormone , RNA , RNA, Messenger , Thyroid Gland , Thyroid Hormones , Transcription FactorsABSTRACT
OBJECTIVES: Orphan nuclear receptor small heterodimer partner (SHP) is involved in osteoblastic differentiation. This study was undertaken to demonstrate a role of SHP in in vivo bone development using microcomputed tomographic (microCT) analysis of SHP knockout (KO) mice. MATERIAL & METHODS: Tibia bones were harvested from 1-, 4-, 8- and 20-week-old wild type (WT) and SHP KO mice. The microarchitecture of tibial bone was analyzed using a microCT (Skyscan 1172; Skyscan, Kontich, Belgium). Samples were scanned at a resolution of 17 microm (isotropic). The X-ray was operated with 50 kV, 200 microA of energy, 1.2 sec of exposure time, and a 0.5 mm thick aluminum filter. Projections were acquired over an angular range of 180degrees. For quantification of the bone mineral density (BMD), the microCT was calibrated using 2 standard phantoms with densities of 0.25 and 0.75 g/cm3. The image slices were reconstructed and analyzed using CT analyzer software (CTan, Skyscan). RESULTS: The CT values of tibial trabecular bone were significantly decreased in SHP KO compared to WT at 20-week-old mice determined by microCT; (bone volume / tissue volume [BV/TV, 40%], BMD [80%], and trabecular number [Tb.N, 50%]). However, the CT values were not significantly different between WT and SHP KO in cortical bone. Furthermore, the qualitative indices of trabecular bone such as the structure model index (SMI) and the polar moment inertia (PMI) did not differ between WT and SHP KO mice. CONCLUSION: These microCT results supports that SHP may act as a positive regulator of trabecular bone formation.
Subject(s)
Animals , Child , Humans , Mice , Aluminum , Bone Density , Bone Development , Child, Orphaned , Mice, Knockout , Osteoblasts , Osteogenesis , Tibia , X-Ray MicrotomographyABSTRACT
Objective: To investigate the changes of cholesterol 7alpha- hydroxylase(CYP7A1) ,farnesoid X receptor(FXR), and small heterodimer partner (SHP) expression in liver tissues of bile duct ligated (BDL) rats,and to analyze the relationship between their expression. Methods: Obstructive cholestasis rat model was induced by bile duct ligation,and rats were sacrificed on day 1,3,and 14 after operation. The hepatic tissues were collected and the total mRNA was extracted. The CYP7A1 mRNA expression was determined by real-time RT-PCR. The protein expression of FXR and SHP in the nuclei was determined by Western blotting analysis. Results: Compared with sham-operated group, experimental obstructive cholestasis group had significantly enhanced CYP7A1 mRNA expression and decreased FXR,SHP expression(P<0. 001,P<0. 05). Conclusion: The up-regulation of CYP7A1 may be associated with the down-regulation of FXR and SHP.
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Objective To evaluate the relationship of small heterodimer partner(SHP) gene and birth weight in China. Methods A cohort study of 191 normal pregnant women was conducted. Both maternal and cord blood samples were collected. PCR-RFLP was used to detect the polymorphism of SNP-rs7504 of SHP. Results (1) The frequency of both neonatal and maternal C allele and (TC+CC) genotype increased significantly with birth weight (P=0.004, OR=3.168; P=0.005, OR=3.315; P=0.013, OR=2.495; P=0.013, OR=2.495). (2) The babies were heavier if they were C allele carrier. The average increase of birth weight was 246.3 g comparing the neonates with TC+CC genotype with those with TT genotype [(3658.7?400.94)g vs (3412.4?444.4)g, P=0.005]. The average birth weight of those maternal C allele carriers was 210.3 g heavier than those non-C allele carriers[(3628.9?405.5) g vs (3418.6?449.0 g]. (3) The fetal C allele was associated with maternal weight in pregnancy, prepregnant BMI, paternal height and weight. Women with C allele were heavier and had higher BMI without statistical significance comparing with those non-C allele carriers. Neither neonatal nor maternal SHP gene was associated with blood glucose and insulin level. (4) Multiple factors analysis showed that birth weight was related to maternal height, weight gain during pregnancy, prepregnant BMI, maternal and cord blood insulin level. After adjustment, the neonatal birth weight remained significantly correlated with cord blood SHP (P=0.0354), but not with maternal SHP gene (P=0.0711). Conclusions SHP gene is associated with newborns birth weight and may affect fetal growth.
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OBJECTIVE: We inversigated Small Heterodimer Partner (SHP) gene mutation in Korean Polycystic Ovarian Syndrome (PCOS) patients. SHP protein regulates the activity of nuclear receptors which regulate the cellular development and differentiation. Recently, the mutation of SHP gene was found in the obesity and diabetes patients in Japanese group, and suggested that its mutation may involved in pathogenic mechanism of PCOS. METHODS: This study was performed in 20 PCOS patients and 20 normal women. The DNAs were extracted from the peripheral bloods, and amplified at each exon (1 and 2) of SHP gene by PCR method. Subsequently, each PCR product was digested with the restriction enzyme indicated below for studying restriction fragment length polymorphism (RFLP). After enzyme digestion, the results of RFLP were compared PCOS patients with control women to find any sequence variation. RESULTS: We examined 9 regions of exon 1 with Msp I, Pvu II, Dde I and 3 regions of exon 2 with Pst I, Dde I. There is no heterozygous or homozygous mutation in patients and control women at these restriction sites. CONCLUSION: The genetic analysis at our restriction sites in the SHP gene did not show any genetic variation in Korean PCOS patients. Our PCR-RFLP analysis was not covered the entire SHP gene (68 bp/ 1,006 bp), we need to further analysis of the entire SHP gene.