Identification and transcriptional activity analysis of core regulatory region of human guanylate binding protein 5 gene promoter / 中国生物制品学杂志

@#Objective To construct luciferase reporter plasmids of truncated fragments of different lengths of human guanylate binding protein 5(GBP5)gene promoter and analyze the transcriptional activity of each fragment to determine the core regulatory region.Methods GBP5promoter sequence was amplified by PCR,truncated into five fragments of different lengths and connected to pGL3-basic plasmid.The constructed recombinant plasmids pGL3-GBP5-11/21/31/41/51were transfected into 293FT cells and detected for luciferase activity.The binding sites of transcription factors in GBP5promoter region were predicted by JASPAR software,and Yin-Yang transcription factor 1(YY1)targeting the core regulatory region was selected and verified for the transcriptional regulatory activity.The CDS sequence of YY1 was amplified by PCR to construct the overexpression plasmid pIRES2-EGFP-YY1,which was then co-transfected to 293FT cells with plasmids pGL3-GBP5-21(-1 623 ～ +47 bp)and internal reference plasmid pRL-CMV,and detected for luciferase activity to analyze the regulation of transcription factor YY1 on GBP5 promoter activity.Results Colony PCR and double enzyme digestion identification proved that the plasmid of human GBP5 promoter reporter gene was correctly constructed;JASPAR software predicted that there were multiple transcription factor binding sites such as STAT1,YY1 and Foxp3 in GBP5promoter region.Double luciferase activity assay showed that pGL3-GBP5-21(-1 623 ～ +47 bp)showed the highest promoter activity,while the promoter activity of pGL3-GBP5-41(-520 ～ +47 bp)decreased significantly,suggesting that the core region of GBP5 promoter was located at upstream-1 623 ～-520 bp of 5 'UTR;Overexpression of YY1 significantly activated the GBP5 promoter activity and regulated the expression of GBP5.Conclusion The core regulatory region of human GBP5 promoter was located in upstream-1 623 ～-520 bp of the 5 'UTR,with a binding site of transcription factor YY1 existing in this region.Meanwhile,overexpression of YY1 significantly effected the activity of GBP5 promoter.

Effect of lncRNA HULC knockdown on rat secreting pituitary adenoma GH3 cells

Pituitary adenoma is one of the most common tumors in the neuroendocrine system. This study investigated the effects of long non-coding RNAs (lncRNAs) highly up-regulated in liver cancer (HULC) on rat secreting pituitary adenoma GH3 cell viability, migration, invasion, apoptosis, and hormone secretion, as well as the underlying potential mechanisms. Cell transfection and qRT-PCR were used to change and measure the expression levels of HULC, miR-130b, and FOXM1. Cell viability, migration, invasion, and apoptosis were assessed using trypan blue staining assay, MTT assay, two-chamber transwell assay, Guava Nexin assay, and western blotting. The concentrations of prolactin (PRL) and growth hormone (GH) in culture supernatant of GH3 cells were assessed using ELISA. The targeting relationship between miR-130b and FOXM1 was verified using dual luciferase activity. Finally, the expression levels of key factors involved in PI3K/AKT/mTOR and JAK1/STAT3 pathways were evaluated using western blotting. We found that HULC was highly expressed in GH3 cells. Overexpression of HULC promoted GH3 cell viability, migration, invasion, PRL and GH secretion, as well as activated PI3K/AKT/mTOR and JAK1/STAT3 pathways. Knockdown of HULC had opposite effects and induced cell apoptosis. HULC negatively regulated the expression of miR-130b, and miR-130b participated in the effects of HULC on GH3 cells. FOXM1 was a target gene of miR-130b, which was involved in the regulation of GH3 cell viability, migration, invasion, and apoptosis, as well as PI3K/AKT/mTOR and JAK1/STAT3 pathways. In conclusion, HULC tumor-promoting roles in secreting pituitary adenoma might be via down-regulating miR-130b, up-regulating FOXM1, and activating PI3K/AKT/mTOR and JAK1/STAT3 pathways.

Effect of active immunization against angiotensin II type 1 (AT1) receptor on hypertension & arterial remodelling in spontaneously hypertensive rats (SHR).

Background & objectives: angiotensin II receptor type 1 (AaT1) is known to be involved in the pathogenesis of hypertension. tThis study was undertaken to explore the effect of active immunization against AaT1 receptor on blood pressure and small artery remodelling in spontaneously hypertensive rat (SHR). Methods: Male SHR and Wistar rats aged two months were actively immunized with different peptides (AaTR12185、AaTR10014 and AaTR12181) corresponding to particular sequences of rat receptor, while another SHR group was given losartan (10 mg/kg/day) orally once a day. Aanti-AaT1 receptor antibodies were detected by ELISAa and blood pressure was measured. The effect of the antibodies on the artery and vascular smooth muscle cells (VSMCs) proliferation was studied. Results: all immunized animals produced antibodies against the particular peptides. The systolic blood pressure was decreased in the SHR immunized with peptide-AaTR12181 compared with the control. However, no changes were observed in the SHR immunized with other two peptides. The Wistar rats immunized with the three peptides did not show any changes in blood pressure. The media/lumen area ratio of the mesenteric artery was reduced in SHR immunized with and similar to that of the SHR treated with losartan. The antibody from SHR immunized with AaTR12181 had no effect on the proliferation of VSMC. But it could inhibit the proliferation caused by angiotensin II and its effect at the titre of 1:40 was similar to that of 1μmol/l losartan. Interpretation & conclusions: Our findings demonstrated that the antibody from SHR immunized with AaTR12181 had the effect of reducing blood pressure and target organ protection similar to losartan. Aactive immunization against AaT1 receptor may be a promising strategy in future for the treatment of hypertension.

The Occurrence of Primary Hepatic Adenoma in Deceased Donor Renal Transplant Recipient

Main findings: We reported a case of new-onset, multi-focal hepatic adenoma in an 18 year-old man with no classic risk factors occurring forty months after a renal transplant from a cadaver donor. Histopathology of the adenoma was examined and genotype and phenotype were also analyzed. Histopathologic examination of the adenoma showed no malignancy. Genotype and phenotype analysis revealed no HNF1α or β-catenin gene mutations and no inflammatory infiltration. The patient was well and disease-free postoperatively. Case hypothesis: Hepatic adenoma occurs mostly in those taking oral contraceptives or androgenic-anabolic steroids or in those with hereditary diseases. Hepatic adenoma in a renal transplant recipient is rare and has only been reported in one case with glycogen storage disease type Ia. Immunosuppressive treatment might have contributed to the development of the neoplasm. Promising future implications: Although malignant change occurs most often in β-catenin gene mutation hepatic adenoma, surgical resection of the adenoma in a patient under immunosuppressive therapy should be considered in order to avoid the possibility of malignant transformation or hemorrhagic rupture. .

Observation on therapeutic effect of "Shandong Xiaoying" needling method for treatment of cerebral infarction / 中国针灸

<p><b>OBJECTIVE</b>To search for an effective method for treatment of cerebral infarction.</p><p><b>METHODS</b>Sixty cases of cerebral infarction were randomly divided into an observation group and a control group, 30 cases in each group. All of them were treated with routine needling method, and "Shandong Xiaoying" needling was added to the observation group. The therapeutic effects, recovery of myodynamia and daily living ability were compared in the two groups.</p><p><b>RESULTS</b>There was no significant difference between the two groups in the total effective rate for recovery of upper limb myodynamia (P > 0.05), but there was a very significant difference in the total effective rate for recovery of lower limb myodynamia and increase of daily living ability between the two groups (P < 0.01).</p><p><b>CONCLUSION</b>"Shandong Xiaoying" needling has markedly therapeutic effect on cerebral infarction.</p>