Lack of Augmenter of Liver Regeneration Disrupts Cholesterol Homeostasis of Liver in Mice by Inhibiting the AMPK Pathway.

It is well known that excessive cholesterol accumulation within hepatocytes deteriorates nonalcoholic fatty liver disease (NAFLD). Augmenter of liver regeneration (ALR) has been reported to alleviate NAFLD through anti-apoptosis; however, whether ALR could protect liver from cholesterol-induced NAFLD remains unclear. Mice with heterozygous deletion of Gfer (the gene for ALR, Gfer +/-) were generated, and liver steatosis was induced by either choline-deficient ethionine-supplemented, methionine choline-deficient diet for 4 weeks, or high-fat diet for 16 weeks. The results showed that Gfer +/- mice developed a more severe fatty liver phenotype than Gfer +/+ mice. The livers of Gfer +/- mice exhibited a higher concentration of cholesterol and low-density lipoprotein compared with the normal mice. Transcriptome-based analysis predicts low-density lipoprotein receptor (LDLR) primarily involved in the metabolic pathway. The experiments further indicate that cholesterol accumulation within hepatocytes is closely associated with enhancing the expression of LDLR and activation of sterol regulatory element binding protein 2 (SREBP2). Because adenosine monophosphate-activated protein kinase (AMPK) is a critical regulator of SREBP2 activation, we measured whether the activity of AMPK was regulated by ALR. We found that knockdown of ALR expression inhibited the phosphorylation of LKB1, an upstream activator of AMPK, followed by AMPK inactivation and SREBP2 maturation/nuclear translocation, leading to extensive cholesterol accumulation. Meanwhile, cellular oxidative stress increased as a result of ALR knockdown, indicating that ALR might also have a role in suppressing reactive oxygen species production. Conclusion: Our results confirm that ALR regulates cholesterol metabolism and alleviates hepatic steatosis probably through the LKB1-AMPK-SREBP2-LDLR pathway in vivo and in vitro, providing a putative mechanism for combating fatty liver disease.

Overexpression of the immediate early response 5 gene increases the radiosensitivity of HeLa cells.

The effects of the immediate early response 5 (IER5) gene on the sensitivity of HeLa cells to radiation remain unclear. In the present study, stably transfected HeLa cells resulting in the knockdown or overexpression of IER5 were investigated. In addition, xenografts of normal, IER5-silenced and -overexpressed HeLa cells were injected into nude mice and examined. The results demonstrated that the radiosensitivity of the IER5-overexpressed HeLa cells was significantly increased compared with that of the normal and IER5-silenced cells. The upregulation of IER5 effectively decreased cell proliferation and IER5 silencing promoted cell proliferation compared with that in the normal HeLa cells. Following irradiation of the cells with IER5 knockdown, cell cycle was arrested at the G2/M phase and an increase in the proportion of S phase cells was observed. By contrast, the overexpression of IER5 led to an increase in the proportion of G1 phase cells. Furthermore, the upregulation of IER5 inhibited tumor growth in vivo. The present findings demonstrate that the IER5 gene affects the radiosensitivity of HeLa cells and serves an important role in cell proliferation, suggesting that this gene may be a potential radiotherapeutic target in cervical cancer.

Differential composition of gut microbiota among healthy volunteers, morbidly obese patients and post-bariatric surgery patients.

The modulation of the gut microbiota was recently deemed one of the mechanisms responsible for the excellent outcomes of bariatric surgery. However, to date, only few studies have assessed this, and they have high heterogeneity. In the present study, next-generation 16S ribosomal DNA amplicon sequencing was used to characterize the gut microbiota of healthy volunteers, as well as patients prior to and after sleeve gastrectomy (SG) or Roux-en-Y gastric bypass (RYGB). Significant differences in α diversity, ß diversity and species were identified between the different groups/time-points. The results demonstrated excellent outcomes of SG and RYGB. The ß diversity was lower in healthy volunteers compared with that in morbidly obese patients with or without type 2 diabetes mellitus. At 3 months after SG, the α diversity was increased and the ß diversity was decreased. The abundance of certain species changed significantly after SG and RYGB. It was also revealed that the abundance of certain microbes was significantly correlated with the body mass index, fasting blood glucose and glycosylated haemoglobin. It may be concluded that bariatric surgery may cause obvious alterations in the gut microbiota and compared with healthy volunteers and obese patients without bariatric surgery, the microbiota composition of post-bariatric surgery has unique characteristics. However, studies with a larger cohort and longer follow-up may be required to confirm these results.

Deficiency in augmenter of liver regeneration accelerates liver fibrosis by promoting migration of hepatic stellate cell.

BACKGROUND: Augmenter of liver regeneration (ALR) protects liver from various injuries, however, the association of ALR with liver fibrosis, particularly its effect on hepatic stellate cells (HSC), remains unclear. In this study, we investigated the impact of ALR on the activation of HSC, a pivotal event in occurrence of liver fibrosis. METHODS: Liver fibrosis was induced in vivo in mice with heterozygous ALR knockdown (ALR-KD) by administration of CCl4 or bile duct ligation. The effect of ALR-KD and ALR-overexpression on liver fibrosis was studied in mice and in HSC cells as well. RESULTS: Hepatic collagen deposition and expression of α-smooth muscle actin (α-SMA) were significantly increased in the ALR-KD mice compared to wild-type mice. In vitro, ALR-shRNA resulted in the activation of HSC cell line (LX-2). Furthermore, ALR-shRNA promoted LX-2 cell migration, accompanied by increased filamentous actin (F-actin) assembly. The ALR-KD-mediated increase in HSC migration was associated with mitochondrial fusion, resulting in mitochondria elongation and enhancing ATP production. In contrast, ALR transfection (ALR-Tx) decelerated HSC migration and inhibited F-actin assembly, concomitantly enhancing mitochondrial fission and reducing ATP synthesis. Mechanically, stimulation of HSC migration by ALR-shRNA was attributed to the increased mitochondrial Ca2+ influx in HSCs. Treatment of ALR-shRNA-cells with Ruthenium Red (RuR), a specific inhibitor of mitochondrial calcium uniporter (MCU), significantly suppressed mitochondrial Ca2+ influx, HSC migration, mitochondrial fusion and ATP production. ALR-KD-induced HSC migration was verified in vitro in primary mouse HSCs. CONCLUSION: Inhibition of ALR expression aggravates liver fibrosis, probably via promoting HSC migration and mitochondrial fusion.

Lack of hepatic stimulator substance expression promotes hepatocellular carcinoma metastasis partly through ERK-activated epithelial-mesenchymal transition.

Hepatocellular carcinoma (HCC) is one of the most lethal malignancies due to its high frequency of metastasis via the epithelial-mesenchymal transition (EMT) pathway. Hepatic stimulator substance (HSS) can protect hepatocytes from injury and promote liver growth. Recent studies indicated that HSS expression is increased in HCC tissues; however, whether HSS expression is potentially associated with HCC metastasis, particularly through the EMT pathway, remains largely unknown. In this study, the relationship between HSS expression and HCC metastasis was investigated in clinical samples of HCC. Meanwhile, the regulation of HCC metastasis and EMT progression by HSS were also analyzed in both in vitro and in vivo models. The results showed that the expression of 23 kDa HSS was significantly decreased among HCC tissues with angioinvasion. A decrease in HSS predicted poor prognosis with a lower survival rate. Furthermore, the growth of xenograft tumors after inoculating MHCC97H-HSS-shRNA (HCC) cells into nude mice was notably accelerated compared to those inoculated with HSS-expressing cells. Further analysis revealed that knockdown of HSS expression in both MHCC97H and HepG2 cells could enhance the migration of these HCC cells. Concurrently, interference of HSS expression by shRNA promoted conversion of morphologically epithelial-like HCC cells into mesenchymal-like cells, together with downregulations of epithelial markers (such as E-cadherin and zonula occludens-1) and upregulation of mesenchymal-like makers (such as α-SMA, ß-catenin, and fibronectin). Furthermore, it was demonstrated that, as well as promoting EMT, HSS-shRNA induced the phosphorylation of extracellular signal-regulated kinase (ERK) and elevated the expression of the EMT-related transcription factor Snail. Specific inhibition of HSS-shRNA-induced ERK phosphorylation by PD98059 attenuated HCC cell migration in a dose-dependent manner. In conclusion, we demonstrated that downregulation of HSS expression contributes to HCC metastasis partially through the ERK-activated EMT pathway.

Deceleration of liver regeneration by knockdown of augmenter of liver regeneration gene is associated with impairment of mitochondrial DNA synthesis in mice.

Hepatic stimulator substance, also known as augmenter of liver regeneration (ALR), is a novel hepatic mitogen that stimulates liver regeneration after partial hepatectomy (PH). Recent work has indicated that a lack of ALR expression inhibited liver regeneration in rats, and the mechanism seems to be related to increased cell apoptosis. The mitochondria play an important role during liver regeneration. Adequate ATP supply, which is largely dependent on effective mitochondrial biogenesis, is essential for progress of liver regeneration. However, ALR gene expression during liver regeneration, particularly its function with mitochondrial DNA synthesis, remains poorly understood. In this study, ALR expression in hepatocytes of mice was suppressed with ALR short-hairpin RNA interference or ALR deletion (knockout, KO). The ALR-defective mice underwent PH, and the liver was allowed to regenerate for 1 wk. Analysis of liver growth and its correlation with mitochondrial biogenesis showed that both ALR mRNA and protein levels increased robustly in control mice with a maximum at days 3 and 4 post-PH. However, ALR knockdown inhibited hepatic DNA synthesis and decelerated liver regeneration after PH. Furthermore, both in the ALR-knockdown and ALR-KO mice, expression of mitochondrial transcription factor A and peroxisome proliferator-activated receptor-γ coactivator-1α were reduced, resulting in impaired mitochondrial biogenesis. In conclusion, ALR is apparently required to ensure appropriate liver regeneration following PH in mice, and deletion of the ALR gene may delay liver regeneration in part due to impaired mitochondrial biogenesis.

Involvement of hepatic stimulator substance in the regulation of hepatoblast maturation into hepatocytes in vitro.

Hepatic stimulator substance (HSS), also known as augmenter of liver regeneration (ALR), acts as a hepatotrophic growth factor to promote liver regeneration after liver damage or partial hepatectomy. However, the expression and function of HSS during liver development in mammals remain largely unknown. In this work, the hepatoblasts were isolated from mice at embryonic day 13.5 (E13.5), and HSS expression and its role during hepatoblast maturation were investigated. The results showed that HSS expression was enhanced in the hepatoblasts compared with mouse primary hepatocytes. HSS expression (23 kDa) was significantly decreased if the hepatoblast maturation was induced by a combination of oncostatin M (OSM), dexamethasone (DEX), and hepatocyte growth factor (HGF). We also found that knockdown of HSS expression (mainly 23-kDa isoform) by siRNA promoted hepatoblast maturation and also activated the signal transducer and activator of transcription 3 (STAT3) phosphorylation levels. However, if STAT3 activity was blocked by a small-molecule inhibitor Stattic, then hepatocyte maturation could be abolished, suggesting that STAT3 was most likely a potential molecule responsible for HSS signaling. In summary, our results demonstrated for the first time that HSS might be an active factor participating in the regulation of liver development and hepatocyte maturation.

Downregulation of hepatic stimulator substance during the early phase of liver regeneration inhibits E-cadherin expression in mice.

Hepatic stimulatory substance (HSS), which encodes a sulfhydryl oxidase enzyme, promotes liver regeneration (LR) and maintains the viability of hepatocytes. Surprisingly, we found that the levels of the HSS mRNA and expressed protein were both strongly repressed at 12h after a 70% partial hepatectomy (PH) in mice. Understanding the mechanism and effect of this extraordinary suppression can provide a novel path for exploring the molecular function of HSS during LR. We observed that the EGF levels in the serum were negatively correlated with HSS expression in regenerating livers. Treating primary mouse hepatocytes or Hepa1-6 cells with EGF suppressed HSS mRNA expression. This suppression was transcriptional and was mediated by the effect of EGF on the phosphorylation of CCAAT/enhancer-binding protein ß (C/EBPß), which regulates HSS expression. We further showed that the enhanced phosphorylation of C/EBPß after PH promoted its interaction with the HSS promoter and repressed HSS expression at early time-points after PH. Interestingly, the knockdown of HSS caused a dramatic decrease in E-cadherin expression in hepatocytes. E-cadherin expression was also significantly suppressed at 12h after PH. Moreover, the pre-injection of HSS-expressing adenovirus vectors prevented E-cadherin suppression after PH. Treatment with C/EBPß siRNA reversed the EGF-mediated inhibition of HSS expression and led to enhanced E-cadherin expression and reduced cell migration. Our findings suggest that C/EBPß directly inhibits the HSS promoter after PH and that this inhibition can downregulate E-cadherin expression. These data provide novel insight into the potential role of HSS in hepatic structural reconstruction during LR.

Epidermal growth factor down-regulates the expression of human hepatic stimulator substance via CCAAT/enhancer-binding protein ß in HepG2 cells.

hHSS (human hepatic stimulator substance), acting as a hepatotrophic growth factor, promotes liver regeneration. However, the regulatory mechanisms for hHSS transcription are still poorly understood. In the present study, we investigated transcription of hHSS triggered by EGF (epidermal growth factor) and the role of C/EBPß (CCAAT/enhancer-binding protein ß) as a potential core factor responsible for hHSS transcription in HepG2 cells. The results show that EGF suppresses hHSS mRNA expression at early time points. Using a promoter deletion assay, we identified a proximal region (-358/-212) that is required for EGF suppression. Overexpression of C/EBPß enhances EGF suppression of hHSS, and mutation of the C/EBPß-binding site at -292/-279 or siRNA (short interfering RNA) interference abolishes EGF suppression. Furthermore, using an electrophoretic mobility-shift assay and chromatin immunoprecipitation analysis, we found that C/EBPß specifically binds to the -292/-279 site that is responsible for EGF inhibition. Moreover, using a knockin (overexpression) and knockdown strategy (siRNA), we confirmed that C/EBPß is a key factor responsible for inhibition of hHSS mRNA expression. Pre-treatment with an inhibitor of JNK (c-Jun N-terminal kinase) or down-regulation of JNK1 with specific siRNA reverses EGF-inhibited hHSS expression. Our results provide a crucial regulatory mechanism for EGF in hHSS transcription within the promoter proximal region.

Increased hepatic UCP2 expression in rats with nonalcoholic steatohepatitis is associated with upregulation of Sp1 binding to its motif within the proximal promoter region.

Uncoupling protein-2 (UCP2) is a mitochondrial inner-membrane carrier protein that is involved in the control of fatty acid metabolism. To understand the mechanism of the transcriptional regulation of ucp2 in the pathogenesis of nonalcoholic steatohepatitis (NASH), we cloned 500 bp upstream of the ucp2 exon 1 from a rat liver cDNA library and identified cis-acting regulatory elements. The transcriptional start site was identified as "C," -359 bp from the ATG codon. A reporter gene assay showed that deletion of the nucleotide sequence between -264 and -60 bp resulted in a significant decrease in promoter activity in HepG2 and H4IIE cells. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) revealed that the increase in promoter activity is related to an enhanced ability of Sp1 to bind to its motifs at -84 to -61 bp within the ucp2 proximal promoter. Overexpression of exogenous Sp1 in H4IIE cells also increased the promoter activity. We demonstrated that the expression of UCP2 mRNA and protein is markedly increased in rats with nonalcoholic steatohepatitis (NASH). Coincidently, levels of Sp1 binding to -84/-61 bp were also increased. Overall, our data indicate that the Sp1-binding site located at the proximal promoter is involved in the regulation of rat UCP2 expression.

Down-regulation of hepatic nuclear factor 4alpha on expression of human hepatic stimulator substance via its action on the proximal promoter in HepG2 cells.

hHSS (human hepatic stimulator substance) stimulates hepatocyte growth. To understand the mechanism controlling hHSS expression, we analysed the proximal promoter activity and identified two regulatory regions (-212/-192 and -152/-132) that were important for transcription in HepG2 cells. Using the luciferase reporter assay, gel-shift experiments and ChIP (chromatin immunoprecipitation), we found that the transcription factors HNF4alpha (hepatocyte nuclear factor 4alpha) and Sp1 (stimulating protein-1) were essential for hHSS promoter activity and could directly bind to regions -209/-204 and -152/-145 respectively. We also confirmed that activation and repression of hHSS transcription induced by Sp1 and HNF4alpha resulted from binding of these factors to these two cis-elements respectively. Overexpression of HNF4alpha led to a dramatic repression of the promoter activity and, in contrast, the activity was markedly elevated by overexpression of Sp1. Furthermore, overexpression of HNF4alpha1, one of the HNF4alpha isoforms, resulted in a dramatic suppression of the promoter activity. Moreover, repression of HNF4alpha expression by siRNA (small interfering RNA) remarkably enhanced the hHSS mRNA level. It has been reported previously that expression of HNF4alpha is functionally regulated by dexamethasone. To further confirm the transcriptional control of HNF4alpha on hHSS, we tested the effect of dexamethasone on hHSS transcription in HepG2 cells. In the present study we have demonstrated that the expression of the hHSS gene was down-regulated at the transcriptional level by dexamethasone in HepG2 cells. A deletion and decoy assay revealed that binding of HNF4alpha to nucleotides -209/-204 was responsible for the suppression of hHSS promoter activity by dexamethasone. Increases in the HNF4alpha-binding activity and expression were simultaneously observed in an electrophoretic mobility-shift assay and Western blot analysis. These results suggested that Sp1 activates hHSS basal expression, but HNF4alpha inhibits hHSS gene expression.

Identification of human hepatic stimulator substance gene promoter and demonstration of dual regulation of AP1/AP4 cis-acting element in different cell lines.

Human hepatic stimulator substance (hHSS) is a newly identified growth-promoting factor in the liver. HSS is capable of stimulating hepatic regeneration in partial hepatectomized rats, thus, promoting growth of hepatic tumor cells. To understand and elucidate the transcriptional regulation of hHSS gene, the 4890bp of 5'-flanking region of the gene have been isolated and sequenced. The transcriptional start site, located at 248nt upstream from the ATG starting codon, was identified by 5'-rapid amplification cDNA end (5'-RACE). The classical promoter sequences, such as TATA box or GAATT were not identified in the promoter region, instead a GC-rich segment was formed (>70%) by expanding to a longer than 400bp, and immediately upstream from the ATG start codon. The transient transfection assays, using promoter deletion constructs, showed that hHSS promoter was efficiently capable in driving the reporter expression not only in HepG2 cells, but also in Cos7 cells. A region spanning nucleotides in the range of -447 to -358bp revealed a negative regulation on promoter activity in HepG2 cells, but with positive regulation in Cos7 and Hela cells. The promoter activity was obviously influenced by AP1/AP4 (-375/-369nt) mutation in these three cell lines. EMSAs showed that the site was recognized by AP1 in HepG2 cell, and only by an AP4 protein in Cos7 cells. The c-Jun bound to the promoter was further verified by supershift in HepG2 cells and human liver tissue. Chromatin immuno-precipitation (ChIP) demonstrated that there was a direct association of c-Jun with hHSS promoter in HepG2 cells. The c-Jun strongly suppressed hHSS promoter activity in transient expression analyses in HepG2 cells. Mutations in the AP1 binding sites rescued suppression caused by c-Jun, suggesting this was a direct regulation of the hHSS promoter. In contrast, there was no significant effect in c-Jun over-expressed Cos7 and Hela cells. The tissue-specific function of c-Jun in hHSS promoter activity may in part help explain the differences in biology function of hHSS between liver and non-liver cells.

[Effect of the regulation of IGFs system components in retinoic acid-induced congenital clubfoot].

The fetal rat models with congenital clubfoot were constructed by treating 24 Wistar rats with all trans retinoic acid (ATRA). The MC-3T3-E1 cells were cultured with ATRA, 17 beta-estrogen (E2) or combinations of the two chemicals. The flow cytometer was used to determine the cell proliferation. The insulin-like growth factor-II (IGF-II ) and IGF conjugated protein-6 (IGFBP-6) mRNA level in rat calvaria bone tissue and MC-3T3-E1 cells were detected by northern blotting analysis and reverse transcription polymerase chain reaction. The congenital clubfoot of fetal rat was induced by ATRA in concentration of 100 approximately 140 mg/kg with dosage-dependence effect. The expression of IGF-II mRNA and cell proliferation were enhanced by E2(1 x 10(-6) mol/L) in rat calvaria bone tissue and MC-3T3-E1 cells, whereas the IGFBP-6 mRNA was increased. ATRA(1 x 10(-6) mol/L), however, inhibited the effect of E2 on regulation of IGF- II gene and IGFBP-6 gene as well as MC-3T3-E1 cell proliferation. These findings provide the evidence that ATRA can induce congenital skeleton malformation and congenital clubfoot in pregnant Wistar rats. IGF-II and IGFBP-6 are important regulative factors for skeleton development and osteoblast proliferation in rat.

[Antiestrogenic effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the gene expression of insulin-like growth factors family in osteoblast cells].

OBJECTIVE: To investigate the antiestrogenic effect of environment teratogen on the gene expression of insulin-like growth factors (IGFs) family in osteoblast cells during rat skeleton development. METHODS: The fetal rat models with congenital skeleton malformation were constructed by treating 20 female Wistar rats with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on pregnant day 10. The MC-3T3-E1 cells were cultured with estrogen, TCDD, or a combination of the two chemicals for 24 hours. The IGF-II and IGFBP-6 mRNA levels in rat calvaria bone tissue and MC-3T3-E1 cells were detected by reverse transcription-polymerase chain reaction. Flow cytometer was used to determine the cell proliferation. RESULTS: TCDD at the concentration of 5-15 microg/kg induced developmental skeleton defect of fetal rat, and the effect was dose-dependent. The expression of IGF-II mRNA gene was enhanced by estrogen in rat calvaria bone tissue and MC-3T3-E1 cells, whereas IGFBP-6 mRNA was decreased. Estrogen increased the cell proliferation in MC-3T3-E1 cells. TCDD, however, inhibited the effect of estrogen on regulation of IGF-II gene and IGFBP-6 gene as well as MC-3T3-E1 cell proliferation. CONCLUSION: These findings provide the evidence that TCDD can induce congenital fetal skeleton malformation under the condition of high estrogen level in pregnant Wistar rats. TCDD has antiestrogenic effect and hence exerts negative influence on the osteoblast cells through target IGF-II and IGFBP-6 of IGFs family.

[Correlation analysis between WNK4 gene and essential hypertension].

OBJECTIVE: To investigate association of mutation in WNK4 gene with essential hypertension and to analyze the expression of WNK4 gene. METHODS: cSNP in WNK4 gene in a small samples was detected by sequencing, then PCR-RFLP was performed in 98 patients with essential hypertension and 95 control subjects. The expression profile of WNK4 gene was tested by RT-PCR. RESULTS: A cSNP was detected in WNK4 gene exon7 G1662A, and there were significant differences in the distribution of allele frequency of G1662A between essential hypertension group and control group. WNK4 gene were expressed in the tissues of kidney, brain, lung, heart, spleen and intestine of fetus. CONCLUSION: WNK4 gene is well correlated with essential hypertension.

[Distribution and significance of cSNP in KCNA7 gene as a novel NIDDM candidate gene in the population of northeast China].

To investigate the distribution and significance of a coding single nucleotide polymorphism (cSNP) of the novel NIDDM candidate gene,KCNA7 in the population of Northeast China, 97 patients with NIDDM and 141 controls were tested. Genotypes of KCNA7 gene T418M(C/T) were performed by PCR-RFLP, and SSCP was used to detect other unknown variations near the C/T site of KCNA7 gene. As a result, no significant difference was observed in the distribution of genotypes of T418M(C/T) between NIDDM and control group. Clinical biochemical examinations showed no significant difference between genotypes in both INDDM and control group, and no other SNPs were found near the C/T site of KCNA7 gene. This study demonstrates the frequency of this cSNP complies well with the Hardy-Weinberg equilibrium in normal group, T418M(C/T) is only a polymorphic maker of KCNA7 gene,and the possibility of association between NIDDM and KCNA7 can not be excluded.

[Polymorphism analysis of 825C/T of the G-protein beta 3 subunit in high risk population of hypertension in the northeast China].

PCR-RFLP was used for genotyping of GNB3 C/T in 133 patients with EH and 257 healthy control subjects in the high risk population and in 98 patients with EH and 110 healthy control subjects in general population. Some biochemical tests were performed also. The association between the polymorphism and blood pressure was analyzed by the case-control study. Not association between GNB3 825C/T polymorphism and blood pressure was found in both populations. CT and TT genotypes in women of the high risk population are associated with diastolic blood pressure, serum sodium and calcium. Although GNB3 is not a susceptible gene of hypertension in the northeast Chinese, it still has some effects on regulation of the blood pressure in susceptible women.

[Cloning and analysis of human alpha-1B glycoprotein precursor gene: a novel member of human immunoglobulin superfamily].

A cDNA of 1694 bp was cloned from human liver Marathon cDNA by means of Rapid Amplification of cDNA Ends (RACE). The cDNA has an open reading frame at 43-1530 bp encoding 495 amino acid residues and a 17-residue signal peptide. There are four Igc2 domains in the translated polypeptide, which is highly homologous to the human alpha-1B glycoprotein isolated from human plasma. Upon this, we conclude that this cDNA is the alpha-1B glycoprotein precursor gene which has never been cloned before, and it may be a novel member of immunoglobulin superfamily and may involved in the cell recognition and the regulation of cell behavior.