1.SHIP2 sensitizes gastric cancer cells BGC-823 to paclitaxel through upregulation of Bim
Yanmei GE ; Yan YE ; Linjie ZHANG
Chinese Pharmacological Bulletin 2014;(5):701-705,706
Aim To study the sensitivity of human gastric cancer BGC-823 cells to paclitaxel after trans-fection of SHIP2 ( The SH2 domain containing inositol 5-phosphatase 2 ) cDNA. Methods Apoptotic cells were determined by the propidium iodide method using flow cytometry. The levels of protein and mRNA ex-pression were measured by Western blot analysis and qRT-PCR, respectively. pCMV6-SHIP2 plasmid and empty vector were transiently transfected into BGC-823 cells, respectively. Stable cell lines were established after infecting BGC-823 cell with GV112-Puromycin and GV112-Puromycin-Bim( Bcl-2 interacting mediator of cell death) lentivirus particles. pCMV6-SHIP2 plas-mid was transiently transfected into the stable cell lines. Results BGC-823 cells were relatively insensi-tive to paclitaxel compared with SGC-7901 cells. The apoptotic rate was only (25. 6 ± 1. 6)% after the treat-ment with 0 . 3 μmol · L-1 paclitaxel for 48 h in BGC-823 cells. The expression levels of Bim protein and mRNA in BGC-823 cells treated with paclitaxel at dif-ferent time points were not significantly changed. The expression of Bim protein was increased after transfec-tion of pCMV6-SHIP2 plasmid, and the apoptotic rate was up to ( 50. 8 ± 0 . 9 )% in BGC-823 cells treated with paclitaxel for 48h. The expression of Bim protein was significantly inhibited after infecting with GV112-Puromycin-Bim lentivirus particles. The apoptotic rate of infected BGC-823 cells was only ( 27. 6 ± 1. 6 )%after treatment upon paclitaxel for 48h. Conclusion Overexpression of exogenous SHIP2 can increase the expression of Bim, induce apoptosis and enhance sen-sitivity of BGC-823 cells to paclitaxel.
2.Preparation of internal quality control substance of real-time PCR to analyze UU-DNA and the design quality control programs
Yanmei GE ; Di ZHANG ; Suyi FAN ; Hang YUAN ; Yuan MAO
International Journal of Laboratory Medicine 2016;37(8):1070-1071,1074
Objective Developing an internal quality control substance of Ureaplasma urealyticum(UU)‐DNA for real‐time PCR to establish an internal quality control system and preliminary evaluation its clinical value .Methods Internal quality control sub‐stance was prepared by mixing samples which Ct value were 24-25(positive sample) and 32 -33(weak positive sample) ,respec‐tively .At the same time ,selecting samples that test results were negative as negative control .The target value ,standard deviation (s) and coefficient of variation(CV) of internal quality control substance were defined by“instant method”for the first 20 runs and Levey‐Jennings quality control(QC) chart after the first 20 runs .Using the“Westdard” multi‐rule quality control methods to ana‐lyze the detection results .Exporting OPSPecs chart by quality control rules in Unity Real Time (URT ) system and setting up new quality control rules according with OPSPecs chart .Results 131 times of the detection of quality control substance were performed totally .The first 20 runs were defined by“instant method”and later 111 runs were defined by Levey‐Jennings QC chart ,the results were stable of quality control substance and reasonable quality control rules .Conclusion Preparing of internal quality control sub‐stance of UU‐DNA used in real‐time PCR might be easy and stable .So ,the internal quality control substance of UU‐DNA could be worthy for practical application in this PCR laboratory .Design internal quality control rules based OPSPecs chart in molecular de‐tection is very simple and practical .
3.Influence of bloody cervical cell samples on HPV genotyping detection
Yanmei GE ; Hang YUANG ; Suyi FAN ; Yuan MAO ; Peng CAO
International Journal of Laboratory Medicine 2017;38(11):1499-1501
Objective To investigate the influence of bloody cervical cell samples on human papillomavirus(HPV) genotyping detection using flowcytometry fluorescence hybridization kit.Methods According to the concentration of hemoglobin(Hb),cervical cell samples were divided into five groups,including group A with Hb of 0 g/L(10 cases),group B with 0 g/L
4. Clinical and genetic characteristics analysis of Chinese children with glutamate dehydrogenase type of congenital hyperinsulinemia
Ge LYU ; Zidi XU ; Peipei HUI ; Qiao ZENG ; Min LIU ; Jie YAN ; Yuyun WU ; Yanmei SANG
Chinese Journal of Pancreatology 2020;20(1):47-51
Objective:
To analyze the clinical features and genetic characteristics of Chinese children with glutamate dehydrogenase type of congenital hyperinsulinism (GDH-HI).
Methods:
Pedigrees with 10 GDH-HI children admitted to Beijing Children′s Hospital from February 2008 to December 2018 were selected as subjects. Clinical features, the detection of pathogenic genes and follow-up data were retrospectively analyzed. Polymerase chain reaction DNA (PCR-DNA) direct sequencing method and second generation sequencing technique were used to analyze the GLUD1 genetic sequences of 10 GDH-HI children and their relatives.
Results:
Of the 10 GDH-HI children, 9 had normal birth weight and 1 was a giant. Nine patients were accompanied by asymptomatic hyperammonemia, and one had normal blood ammonia. 9 had ever been treated with diazoxide, which was all effective. All 10 children carried GLUD1 gene mutations, 5 patients carried c. 965C>T (p.R322H) GLUD1 gene mutation, and the remaining 5 cases carried c. 1388A>T (p.N463I), c. 1495C>A(p.G499C), c. 1493C>T(p. S498L), c. 1519G>A(p.H507Y) and c. 1388A>G(p.N463S), respectively. 9 cases (90%) had de novo mutations, and 1 case had paternal autosomal dominant inheritance. 8 children were followed up in long term. One child had spontaneous remission in 8 years after being diagnosed, and seven patients required long-term oral diazoxide to maintain normal blood glucose levels, two of whom had epilepsy.
Conclusions
The birth weight of children with GDH-HI in China was usually normal. A small number of GDH-HI children had normal serum ammonia levels. Most of the GLUD1 gene mutations in GDH-HI children in China were de novo mutations, among which the GDH p. R322H mutation was a hot spot mutation in Chinese children with GDH-HI. Most of GDH-HI children were diazoxide-responsive. As the disease progresses, some children may have epilepsy, and a few children have a tendency to relieve by themselves.