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Objective:To investigate the effect of methyltransferase like 14 (METTL14) on the proliferation and invasion of breast cancer (BC) cells by regulating cyclin L2 (Cyclin L2, CCNL2) through m6A modification.Methods:Cancer tissues and paracancerous tissues of BC patients in Yantaishan hospital were collected from Aug. 2018 to Feb. 2020. The expression levels of m6A, METTL14 and CCNL2 in tissues were detected by high performance liquid chromatography/mass spectrometry (HPLC/MS) and qRT-PCR. Dual-luciferase reporter assay, qRT-PCR, and western blot were used to verify the regulatory relationship between METTL14 and CCNL2. RIP experiments verified the regulatory relationship between YTH domain-containing family protein (YTHDF2) and CCNL2. Cell viability was detected by MTT method, and cell invasion ability was detected by Transwell.Results:Compared with normal cells (0.24±0.02) and tissues (0.18±0.02) , BC cells MCF-10A (0.47±0.03, t=11.05, P<0.001) and HS-578T (0.41±0.03, t=8.17, P=0.001) and BC tissues (0.39±0.02, t=12.86, P<0.001) m6A level increased. Compared with normal tissues (1.00±0.26) (0.84±0.07) , METTL14 mRNA (1.57±0.28, t=13.50, P<0.001) and protein levels (1.66±0.11, t=10.89, P<0.001) in BC tissues were significantly increased high. Compared with the control group (100.00±10.11) (1.00±0.12) , the BC cell invasion ability (54.15±6.21, t=6.69, P=0.003) and activity (0.64±0.06, t=4.65, P=0.010) were weakened. Compared with the control group (100±11.05) (1±0.13) , the BC cell invasion ability (175.31±13.45, t=7.49, P=0.002) and activity (2.16±0.16, t=9.75, P=0.002) in the METTL14 overexpression group were enhanced, and the effects of METTL14 on cell invasion (137.41±12.64, t=3.56, P=0.024) and activity (1.64±0.15, t=5.59, P=0.005) were partially reversed after m6A inhibitor treatment change. Compared with normal tissues, CCNL2 expression was down-regulated in BC tissues, and the interaction between CCNL2 and METTL14 was confirmed. Compared with the control group (1.00±0.1) (0.64±0.05) , knockdown of METTL14 could make CCNL2 mRNA (1.67±0.05) . 0.13, t=7.08, P=0.002) and protein (1.09±0.09, t=7.57, P=0.002) were up-regulated. METTL14 knockout enhanced the stability of CCNL2 mRNA through a YTHDF2-dependent pathway, compared with sh-METTL14 group (50.47±5.16) (0.52±0.05) , BC cell invasion ability of sh-METTL14+sh-CCNL2 group (71.69±6.41, t=4.47, P=0.011) and activity (0.64±0.05, t=2.94, P=0.042) were improved. Conclusion:METTL14 inhibits the expression of CCNL2 through m6A modification to enhance the invasion and activity of BC cells.
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Objective:To investigate the effects of circ_000543 derived from hypoxic exosomes on proliferation and invasion of breast cancer (BC) cells.Methods:Bioinformatic website was used to predict the abnormal expression of circ_000543 in BC tissue and the transcription factor that might regulate circ_000543. Double luciferase report experiment and ChIP assay were used to confirm the regulation relationship between YY1 and circ_000543. Exosomes were separated from normal BC cells and BC cells under hypoxic condition, and qRT-PCR was adopted to detect the expression of circ_000543 in exosomes. The expression of circ_000543 and YY1 in exosomes was intervened and the exosomes were cocultured with BC cells under normoxia. CCK8 and Transwell assay was used to detect the proliferation and invasive ability individually.Results:qRT-PCR experiment found that, compared with MCF-10A cells (1±0.11) and exosomes isolated from normoxic cells (1±0.10), circ_000543 expression was up-regulated in BC cells (1.59±0.13) and exosomes derived from cells under hypoxic condition (1.63±0.12) ( t=6.001, P=0.004; t=6.986, P=0.002). Exosomes derived from cells under hypoxic condition promoted proliferation and invasion of BC cells under normoxia. Inhibition of circ_000543 partially offset the effects of exosomes. YY1 induced the expression of circ_000543 in BC cells as a transcription factor. The expression of circ_000543 was inhibited when YY1 expression was down-regulated; at the same time, down-regulation of YY1 inhibited the effects of exosomes on proliferation and invasion of BC cells. Conclusion:The transcription factor YY1 promoted proliferation and invasion of BC cells by inducing hypoxic breast cancer-derived exosomal circ_000543.
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OBJECTIVE:To investigate the effects of alanyl-glutamine dipeptide-intensified early enteral nutrition (EN) sup-port on nutritional indexes,immune indexes,renal indexes and complications of sepsis patients. METHODS:A total of 112 cases of sepsis admitted into our hospital during May 2013-Jan. 2015 were selected and divided into observation group and control group according to random number table,with 56 cases in each group. Both groups received routine antibiotic therapy and early EN support(48 h)with nitrogen supplement 0.2 g/kg,calories 25 kcal/kg and nonprotein calories 19-21 kcal/kg each day. Observation group was additionally given Alanyl-glutamine for injection 0.5 g/kg with 0.9% sodium chloride injection 100 mL,continuous pump within 24 h,for 4 d. The levels of nutritional indexes (ALB,PAB,Hb),immune indexes (CRP,IgG,IgA and IgM), APACHEⅡ scores,SOFA scores,liver and renal function indexes (the levels of ALT,AST,Cr and BUN) were compared be-tween 2 groups before and after treatment. The prognosis and the occurrence of complication were also observed in 2 groups. RE-SULTS:Before treatment,there was no statistical significance in the levels of ALB,PAB,Hb,CRP,IgG,IgA,IgM,ALT, AST,Cr,BUN and APACHEⅡ scores,SOFA scores between 2 groups(P>0.05). After treatment,the levels of ALB and PAB in observation group were increased significantly compared to before treatment,and the observation groups was significantly higher than control group,with statistical significance(P<0.05). APACHEⅡ score and SOFA score of 2 groups were decreased significantly compared to before treatment,and the observation group was significantly lower than the control group,with statis-tical significance(P<0.05). CRP of 2 groups were decreased significantly while IgG were increased significantly;the observa-tion group was significantly better than the control group,with statistical significance (P<0.05). Cr and BUN levels of 2 groups were decreased significantly compared to before treatment,and the level of Cr in observation group was significantly con-trol group,with statistical significance (P<0.05). The time of ICU stay,ventilator supporting time and antibiotics application time in observation group were significantly shorter than control group,and the incidence of diarrhea and gastric retention were significantly lower than control group,with statistical significance (P<0.05). CONCLUSIONS:Alanyl-glutamine dipeptide-in-tensified early EN support can significantly improve immune function and liver and renal function of sepsis patients and reduce the occurrence of complications.
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OBJECTIVE:To study the general pattern,characteristics and related risk of adverse drug reaction(ADR)induced by Monosialotetrahexosylganglioside sodium injection,and to provide reference for safe use of drugs in the clinic. METHODS:Us-ing“monosialotetrahexosylganglioside”and“ADR”as keywords,literautres were retrieved from CJFD,Wanfang and VIP database according to inclusion and exclusion criteria in Oct. 2014,and cases reports were extracted and analyzed statistically. RESULTS:A total of 19 literatures were included,involving 56 patients,with the ratio of male to female was 6.17∶1,and the most of patients aged 0-9 years,≥60 years,accounting for 44.64%(25 cases),30.36%(17 cases). Multiple organs or systems were involved in ADR;the most common reactions are fever (29 cases) and shiver (28 cases),etc.,and severe ADR can cause green-barry syn-drome,anaphylactic shock,etc. CONCLUSIONS:Thus,it is important to pay attention to ADR caused by Monosialotetrahexosyl-ganglioside sodium injection,to prevent the occurrence of serious ADR,to perfect drug instructions,and to ensure clinical safe ad-ministration.
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Objective To analyze the prognostic factors for patients with hepatic encephalopathy (HE) in acute-on-chronic liver failure(ACLF).Methods A retrospective analysis was performed on 159 ACLF patients with HE.The hepatic encephalopathy was determined to baseline,the patients were divided into survivors(n =13) and nonsurvivors(n =146),The 32 factors affecting the prognosis were analyzed by Cox proportional hazard model with SPSS.Results One-month,three-month,and six-month survival rates were 20.13％,10.06％ and 8.18％,respectively.Cox regression analysis of prognostic factors showed that it could the stage of hepatic encephalopathy and HRS significantly improve the survival rate of the patients with HE in acute-on-chronic liver failure.The stage of hepatic encephalopathy and HRS could significantly decrease the survival rate of the patients(x2 =18.344,11.368,all P ＜ 0.05),elevated the levels of hepatic encephalopathy (relative risk (RR) =1.591) and HRS (RR =1.809) indicate worse prognosis with hepatic encephalopathy in acute-on-chronic failure.Conclusion The stage of hepatic encephalopathy and HRS were independent risk factor sof prognosis in acute-on-chronic liver failure.
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Objective To investigate the expressions of Th17, CD4 +CD25 +Treg, HLA-DR mRNA in peripheral blood of children with EV71-induced hand, foot and mouth diseases ( HFMD ) and their clinical significance.Methods Stratified random sampling was used to select 60 children with HFMDs from Liaocheng People’s Hospital from February to October, 2014, including 20 mild, 20 severe and 20 critically ill cases.Twenty healthy children were also enrolled as the control group.All the children with HFMDs were given ribavirin (10 mg/kg) for the treatment.The percentages of Th17 and CD4 +CD25 +Treg cells in CD4 +T cells of peripheral blood were determined by flow cytometry, and the expression of HLA-DR mRNA in peripheral blood mononuclear cells was detected by reverse transcription polymerase chain reaction ( RT-PCR).Analysis of variance and SNK-q test were used to compare the expressions of Th17, CD4 +CD25 +Treg and HLA-DR mRNA among groups, and Pearson correlation analysis was performed to reveal the correlations between HLA-DR mRNA and Th17, CD4 +CD25 +Treg.Results Compared with the control group, the expression of Th17 was increased, while CD4 +CD25+Treg and HLA-DR mRNA expressions were decreased in children with HFMDs on d1 of treatment (F=310.4, 81.5 and 545.4, P0.05), but Th17 level in fatal was still higher and CD4 +CD25 +Treg level was still lower than those in control group (t=16.4 and 12.0, P<0.05).After 10 d of treatment, HLA-DR mRNA expressions in mild group and severe group were increased to the normal level.HLA-DR mRNA expression in surviving patients of critical ill group was still lower than that in mild group and severe group (P<0.05), but was higher than that in fatal patients (t=7.8, P<0.05).Pearson correlation analysis showed that, HLA-DR mRNA was negatively correlated with Th17 level (r=-0.770, P<0.01), and positively correlated with CD4 +CD25 +Treg level (r=0.883, P<0.01).Conclusion The expressions of Th17, CD4 +CD25 +Treg cells, and HLA-DR mRNA are correlated with the severity of HFMD, and may be used for evaluation of disease severity and prediction of disease outcomes.
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Objective To study the pharmacokinetics and bioequiavailability of Glipizide in healthy volunteers. Methods The plasma levels of Glipizide were determined at 30 min,1,2,3,4,5,6,8,10,12,14 h by HPLC after single-dose oral cross administration of 10 mg Glipizide tablets of two brands in 18 healthy male volunteers aged 23 to 25 years old,with the interval of one week. Results Glipizide tablets of the two brands were as follows,Tmax: (2.39?0.50),(2.33?0.49) h; Cmax: (675.3?151.7),(647.1?166.7)g/L; AUC_ 0-t : (3 565.4? 733.4) ,(3 304.8?588.4) ?g?h -1 ?L -1 . Conclusion The two preparations were bioequivalent.
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Objective To investigate the roles of sphingosine kinase (SPK) on the regulation of proliferation, migration and apoptosis of gastric carcinoma cells. Methods BGC-823 cells were infected with adenoviral carriers bringing the wild type SPK gene and SPK mutant (SPK DN ), and their effect on the migration and 5-FU-induced apoptosis of BGC-823 cells was evaluated. The adenovirus-mediated expression of SPK was detected by Western blot, and cellular SPK enzyme activity was assayed. The cell migration was investigated by Transwell Migration Technology. Results The adenovirus-mediated wild type SPK gene increased the cellular SPK activity, thus leading to enhanced migration and resistance to 5-FU-induced apoptosis of BGC-823 cells. Whereas overexpression of SPK mutant (SPK DN ) decreased the cellular SPK activity, thus inhibited the migration and sensitized BGC-823 to 5-FU-induced apoptosis. Conclusion SPK plays important roles in the regulation of migration and apoptosis of gastric carcinoma cells, therefore it may be of a new target for tumor treatment.