MicroRNA-124a Protects the Myocardium Against Ischemia Reperfusion Injury Through Regulation of the Notch Signaling Pathway

Abstract Introduction: This study's objective is to investigate the effect of downregulation of micro ribonucleic acid (miR)-124a on myocardial injury after ischemia reperfusion (I/R) in rats. Methods: Sprague Dawley (SD) rats (n=20) were divided into four groups - sham, I/R, I/R+miR-124a antagomir (I/R+ant-miR-124a), and I/R+ant-normal control (NC). The pathomorphological and infarct size variance of injured myocardial tissues with IR were conducted with hematoxylin (HE) and triphenyltetrazolium chloride (TTC) staining. The expression levels of miR-124a, BAX, nuclear factor kappa B (NF-KB), Notch1, and Hes1 were examined by quantitative real-time polymerase chain reaction or Western blot in myocardium. The inflammatory cytokines interleukin (IL)-6, IL-1β, and tumor necrosis factor alpha (TNF-α) were detected by the enzyme-linked immunosorbent assay, as well as the activity of lactate dehydrogenase (LDH) and creatine kinase (CK) in serum by colorimetry. Results: The expression of miR-124a was increased in the I/R group. Compared with I/R and I/R+ant-NC groups, after downregulating miR-124a, the expression of IL-6, IL-1β, TNF-α, BAX, NF-KB, LDH, and CK were decreased, but the expression of Notch1 and Hes1 were increased. In HE staining, myocardial tissue edema, red blood cell exudation, and myocardial fiber arrangement disorder were accompanied by inflammatory cell infiltration and local necrosis in the I/R group. However, the pathological injury of myocardial tissue was alleviated after downregulating miR-124a. Additionally, TTC results showed that the myocardial infarction area was decreased in the I/R+ant-miR-124a group. Conclusion: Downregulation of miR-124a expression through Notch pathway can significantly reduce myocardial damage after 24 hours of I/R in SD rats. Therefore, miR-124a may become a potential therapeutic target for I/R injury.

Effect of wheat-grain moxibustion at Yinbai (SP 1) on the menstrual period prolongation after IVF-ET down-regulation treatment / 中国针灸

OBJECTIVE@#To compare the clinical effect between wheat-grain moxibustion at Yinbai (SP 1) and oral administration of dydrogesterone tablet for menstrual period prolongation after down-regulation treatment of in vitro fertilization embryo transfer (IVF-ET).@*METHODS@#A total of 54 patients with prolonged menstrual period after down-regulation treatment of IVF-ET were randomly divided into an observation group and a control group, 27 cases in each one. In the observation group, when the menstrual period delayed more than 7 days, the wheat-grain moxibustion at Yinbai (SP 1) was performed, once a day, with an interval of 1 day between two 3-day treatments; when the menstrual blood was cleaned, the ovulation was continued and the eggs were taken. In the control group, when the menstrual period delayed more than 7 days, the oral administration of dydrogesterone tablet was provided, 10 mg each time, twice a day; when the menstrual blood was cleaned, the ovulation was continued and the eggs were taken. The number of days for menstrual blood to be cleaned, the area change of uterine cavity hemorrhage, the morphology of endometrium, the blood supply of endometrium, the number of oocytes obtained, the grade of frozen embryo and the clinical effect were observed between the two groups after treatment.@*RESULTS@#Compared with the control group, the number of days for menstrual blood to be cleaned was shorter in the observation group after treatment (0.05). The cured rate in the observation group was 100.0% (27/27), higher than 33.3% (9/27) in the control group (<0.05).@*CONCLUSION@#The wheat-grain moxibustion at Yinbai (SP 1) could more effectively treat prolonged menstrual period after IVF-ET down-regulation treatment, which is beneficial to the preparation of the endometrium, and has no effect on the oocyte collection and embryo culture.

G-quadruplex forming region within WT1 promoter is selectively targeted by daunorubicin and mitoxantrone: A possible mechanism for anti-leukemic effect of drugs

Wilms tumor 1 (WT1) has long been overexpressed in acute myeloid leukemia and has a prognostic value in its diagnosis.Lately, the formation of G-quadruplexes in oncogenic promoters like WT1 has been widely investigated since stabilizationof these structures leads to transcriptional inhibition of the oncogene. Daunorubicin and mitoxantrone considered as crucialcomponents of almost all standard acute myeloid leukemia induction regimens. Herein we have proposed a probablemolecular mechanism of action through which the drugs may stabilize WT1 promoter G-quadruplexes. Differential pulsevoltammetry, circular dichroism, and polyacrylamide gel electrophoresis, electrophoretic mobility shifts assay, polymerasechain reaction (PCR) stop assays, and quantitative RT-PCR were performed in order to better understanding the nature ofinteractions between the drugs and G-quadruplexes. Data revealed that both drugs had potential to stabilize G-quadruplexesand down-regulate WT1 transcription but daunorubicin exposed more silencing impact. The results illustrated the therapeutic association of these two commercial FDA-approved drugs in WT1 transcriptional down-regulation. Since WT1 hasknown as a transcriptional regulator of at least 137 target genes, so the new data are significant for the development of newapproaches to regulating WT1 and other target genes by employing special drugs in cancer treatment.

Analysis of relative factors affecting the pregnancy outcome of frozen-thawed embryo transfer in patients with endometriosis / 上海交通大学学报（医学版）

Objective:To explore the factors affecting the pregnancy outcome of frozen-thawed embryo transfer (FET) in endometriosis (EMT) patients in order to provide reference for the clinical selection of FET strategies. Methods:A total of 329 EMT patients who received blastocyst FET at the Reproductive Medicine Center, Department of Obstetrics & Gynecology, The 900th Hospital of the Joint Logistics Support Force, PLA, from Jan. 2015 to Dec. 2017 were analyzed retrospectively. The patients were divided into three groups according to endometrial preparation protocols, ages, and endometrial thickness on the day of progesterone conversion, respectively. By endometrial preparation protocols, the three groups included gonadotropin-releasing hormone agonist (GnRH-a) down-regulation+ hormone replacement therapy (HRT) group (GnRH-a+HRT group, A1 group, n=138), HRT group (B1 group, n=52), and natural cycle (NC) group (C1 group, n=139). By ages, the three groups included 35 years old group (C2 group, n=59). By endometrial thickness on the day of progesterone conversion, the three groups included 12 mm group (C3 group, n=37). The differences in pregnancy outcomes among EMT patients with blastocyst FET were compared under different grouping factors. Results:The endometrium of A1 group was significantly thicker than that of B1 group (P=0.041), the implantation rate and clinical pregnancy rate of B1 group were significantly higher than those of C1 group (P=0.000, P=0.003). Compared with A1 group, the implantation rate of B1 group was significantly higher (P=0.023), while it was significantly lower in group C1 (P=0.027). The abortion rate of A2 group was significantly higher than that of B2 group (P=0.007). Compared with A3 group, the implantation rate of B3 group was significantly higher (P=0.041), while it was significantly lower in C3 group (P=0.026). Conclusion:HRT endometrial preparation protocol for EMT patients with blastocyst FET can improve the implantation rate and clinical pregnancy rate, and reduce the abortion rate and ectopic pregnancy rate, which may be an economical and efficient endometrial preparation protocol in clinical.

Effects of three kinds of endometrial preparation on pregnancy outcomes of patients with frozen-thawed embryo transfer / 吉林大学学报(医学版)

Objective: To investigate the effects of three kinds of endometrial preparation (normal hormone replacement, half- and full-dose of long-acting gonadotropin-releasing hormone agonist (GnRH-a) down-regulation combined with hormone replacement) on the pregnancy outcomes in the frozen-thawed embryo transfer (FET) cycle in the patients with endometriosis (E M T), adenomyosis or repeated implantation failure (RIF) for unknown reasons, and to provide a basis for the selection of clinical endometrial preparation method. Methods: A total of 191 patients with EMT, adenomyosis or RIF for unknown reasons underwent FET treatment were selected. The patients were divided into normal hormone replacement group (n=63), half-dose GnRH-a group (n=61) and full-dose GnRH-a group (n=67) according to the endometrial preparation method. The clinical data of patients in each group such as age, body mass index (BMI), duration of infertility, the number of embryo transfer cycles, the number of embryos transferred, the endometrial thickness on the day of conversion and transplantation, the rate of high-quality embryos transferred, the intrauterine clinical pregnancy rate and the embryo implantation rate were analyzed retrospectively and compared. Results: There were no significant differences of the general clinical data of the patients in FET cycles in three groups such as age, BMI, duration of infertility, the number of embryo transfer cycles, the number of embryos transferred, the endometrial thickness on the day of conversion and transplantation, and the rate of high-quality embryos transferred (P

Colonic Transit Disorder Mediated by Downregulation of Interstitial Cells of Cajal/Anoctamin-1 in Dextran Sodium Sulfate-induced Colitis Mice

BACKGROUND/AIMS: Interstitial cells of Cajal (ICC) and their special calcium-activated chloride channel, anoctamin-1 (ANO1) play pivotal roles in regulating colonic transit. This study is designed to investigate the role of ICC and the ANO1 channel in colonic transit disorder in dextran sodium sulfate (DSS)-treated colitis mice. METHODS: Colonic transit experiment, colonic migrating motor complexes (CMMCs), smooth muscle spontaneous contractile experiments, intracellular electrical recordings, western blotting analysis, and quantitative polymerase chain reaction were applied in this study. RESULTS: The mRNA and protein expressions of c-KIT and ANO1 channels were significantly decreased in the colons of DSS-colitis mice. The colonic artificial fecal-pellet transit experiment in vitro was significantly delayed in DSS-colitis mice. The CMMCs and smooth muscle spontaneous contractions were significantly decreased by 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB), an ANO1 channel blocker, and NG-Nitro-L-arginine methyl ester hydrochloride (L-NAME), an inhibitor of nitric oxide synthase activity, in DSS-colitis mice compared with that of control mice. Intracellular electrical recordings showed that the amplitude of NPPB-induced hyperpolarization was more positive in DSS-colitis mice. The electric field stimulation-elicited nitric-dependent slow inhibitory junctional potentials were also more positive in DSS-colitis mice than those of control mice. CONCLUSION: The results suggest that colonic transit disorder is mediated via downregulation of the nitric oxide/ICC/ANO1 signalling pathway in DSS-colitis mice.

Downregulation of OsAGO17 by artificial microRNA causes pollen abortion resulting in the reduction of grain yield in rice

Background: Pollen development is an important reproductive process that directly affects pollen fertility and grain yield in rice. Argonaute (AGO) proteins, the core effectors of RNA-mediated silencing, play important roles in regulating plant growth and development. However, few AGO proteins in rice were reported to be involved in pollen development. In this study, artificial microRNA technology was used to assess the function of OsAGO17 in pollen development. Results: In this study, OsAGO17, a rice-specific gene, was specifically expressed in rice pollen grains, with the highest expression in uninucleate microspores. Downregulation of OsAGO17 by artificial microRNA technology based on the endogenous osa-miRNA319a precursor was successfully achieved. It is found that downregulation of OsAGO17 could significantly affect pollen fertility and cause pollen abortion, thus suggesting that OsAGO17 functions in rice pollen development. In addition, the downregulation of OsAGO17 mainly caused a low seed-setting rate, thereby resulting in the reduction of grain yield, whereas the downregulation of OsAGO17 did not significantly affect rice vegetative growth and other agricultural traits including number of florets per panicle, number of primary branch per panicle, and 100-grain weight. Furthermore, the result of subcellular localization analysis indicated that the OsAGO17 protein was localized to both the nucleus and the cytoplasm. Conclusion: These results represent the first report of the biological function for OsAGO17 in rice and indicate that OsAGO17 may possibly play crucial regulatory roles in rice pollen development. It helps us to better understand the mechanism of pollen development in rice.

Three GnRH Agonist Prototols of Down Regulation on Pituitary Suppression and Pregnancy Outcome of In Vitro Fertilization in Women with Normal Ovarian Reserve : A Randomized-Control Clinical Trial / 中山大学学报(医学科学版)

[Objective] To investigate the effects of different doses of gonadotropin releasing hormone agonist (GnRH-α) on the down-regulation of normal ovarian reserve,and compared the down-regulation level as well as the clinical outcome of in vitro fertilization and embryo transfer (IVF-ET) cycles.[Methods] This RCT study included 63 infertility couples of age＜35 yrs.women with normal ovarian reserve function who were intended to received GnRH-α long protocol treatment.Of the 63 women were randomly divided into three groups according to the dose of triptorelin,21 received daily 0.05 mg short-acting GnRH-α,21 received daily 0.1 mg short-acting GnRH-α,while 21 received reduced-dose depot of 1.25 mg GnRH-αt.[Results] In the three groups,the average duration of down-regulation reached after injection of GnRH-α,the level of LH and E2,the total number of antral follicles,the number of antral follicles of ＜4 mm and 8～9 mm were similar.The serum follicle-stimulating hormone level on the day of gonadotropin initiation were significantly higher in the two short-acting groups compared with the long-acting group [(3.92 ± 1.12) U vs.(3.03 ± 1.14) U vs.(2.05 ± 1.12) U,P＜ 0.001].Four hours after the GnRHa injection,the serum FSH,LH levels were higher in short-acting 0.05 mg group than the short-acting short-acting 0.1 mg group.Both number of days of gonadotropin stimulation and gonadotropin doses were similar in three groups.On the day of hCG administration,the numbers of 14-18mm diameter follicles [(3.91 ±2.12) vs.(5.81 ±3.55) vs.(6.43±3.39),P＜0.001] as well as the proportion of follicles with diameter ≥18 mm/≥10 mm [(33.1％± 13.2％) vs.(24.0％±12.4％) vs.(30.1％±12.2％),P＜0.05],were both statistically significant different in three groups.Although serum LH level on hCG day was significantly increased in 0.05 mg group [(2.47±1.33) U vs.(1.80±0.69) U vs.(1.43±0.53) U,P＜0.05].No premature LH surge and premature ovulation was observed.The number of retrieved oocyteswas significant different [(10.14±4.80) vs.(11.51±2.42) vs.(12.79±2.73),P＜0.05].However,no significant differences was found regard to the number of MII oocytes,and the serum estrogen level per egg was significant higher in 0.05 mg group [(282.33±42.13) U vs.(221.62±32.02) U vs.(200.03±37.89) U,P＜0.001].The live birth rate (LBR) of these three groups in fresh cycles were 61.9％,55.0％,and 50.0％,respectively.The cumulative LBR were 85.7％,76.2％,and 75.0％,respectively.A increased trend was observed in the clinical pregnancy rate,cumulative clinical pregnancy rate and cumulative LBR in 0.05 mg group than the other two groups.[Conclusion] For women with normal ovarian reserve,as the GnRH-α dosage decreased,the down-regulation of pituitary reduced,while serum LH levels on the day of hCG trigger increased.The number of oocytes retrieved was decreased,the proportion of cycles which retrieved ＞ 15 oocytes was also lower.However,the average estrogen level per egg was significant increased,and a better clinical outcome of IVF-ET was received.

Clinical Effects of Different Doses of Long-acting Gonadotropin Releasing Hormone Agonist (GnRH-a) Versus Daily Injections of Short-acting GnRH-a in IVF Cycles / 医学研究杂志

Objective Clinical efficacy was compared among single injections of different doses of long acting gonadotropin releasing hormone agonist (GnRH-a),and daily injections of short-acting GnRH-a in order to evaluate different methods of ovarian stimulation for in vitro fertilization (IVF) cycles.Methods A retrospective study of 214 patients who underwent IVF assisted fertility treatments was conducted.Patients were allocated into four study groups:the short protocol (group A),in which daily injections of 0.1 mg GnRH-a was administered in the mid-luteal phase until the day of human chorionic gonadotropin (hCG) administration (see below);or the long protocol (group B,C & D),in which single injections of 3.75mg,2.0mg,or 0.9mg of long-acting GnRH-a was given in the mid-luteal phase,respectively.Stimulation with gonadotropins (Gn) started when pituitary down-regulation was established.When vaginal ultrasonographic scans showed that at least two follicles had reached 16-20mm in diameter,Gn stimulation was withdrawn,and serum estradiol (E2),progesterone (P),and luteinizing hormone (LH) were determined.Additionally,human chorionic gonadotropin (hCG) was administered that evening.Egg collection was performed 38 hours after hCG injection and the standard IVF procedure was performed.Results There were no statistically significant differences amongst the four groups when measuring serum LH levels,number of oocytes,number of fertilized eggs,number of good quality embryos,and clinical pregnancy rate.The total amount of Gn administered was almost identical when comparing group A and group D,as well as when comparing group B and group C.However,Group A and D required less Gn stimulation to exhibit follicles of 16-20mm in diameter,compared to group B and C (P ＜0.005).Moreover,there was a significant difference in the time required for ovulation induction between group A and group C,where group A had a shorter time to ovulation.The fertilization rate was statistically different between group B and other groups (P ＜ 0.005).Conclusion Through our data analysis,we conclude based on outcome,cost,side-effects,and simplification of treatments,that the 0.9mg long-acting GnRH-a treatment is eminent for ovarian stimulation for IVF.

Inhibitory effects of dexamethasone on proliferation of fibroblasts derived by idiopathic orbital inflammatory pseudotumor / 中华实验眼科杂志

Background Idiopathic orbital inflammatory pseudotumor (IOIP) is a common orbital disease, but its etiology is still unclear,so the effect of glucocorticoid treatment is unsatisfied.Objective This study was to investigate the effects of dexamethasone on orbital fibroblasts from IOIP patients and explore the action machanism.Methods Six pieces of IOIP tissues from 6 IOIP patients and 3 pieces of normal orbital connective tissues from lacrimal gland prolapse patients were obtained during the surgery in Beijing Tongren Hospital from November 2011 to January 2012.The orbital fibroblasts were cultured using explant culture method.The morphology of the cells were observed under the optical microscope,and biomarks of the cells were detected by immunochemistry.The growth and proliferation of the cells were assayed using WST-8.The expression of ICAM-1 in the cells in both the control group and the IOIP group was detected by immunochemistry.The fibroblasts were incubated in 96-well plates, and different concentrations of dexamethasone (0,1 × 10-3 , 1 × 10-4 , 1 × 10-5 and 1 × 10-6 mol/L) were respectively added into the medium for 24,48 and 72 hours,and then the proliferation of the cells was detected by WST-8 assay.The contents of ICAM-1 in different concentrations of dexamethasone groups were assayed by ELISA.Results The characteristics of the cells were similar between the control group and the IOIP group with the spindle shape and long protructions.The cells showed the positive response for vimentin and absent response for desmin, S-100, cytokeratin (CK).Compared with the control group,the growth speed of fibroblasts was fast in the IOIP group.The proliferative values of the cells (absorbancy) were gradually reduced with the increase of dexamethasone concentrations (F ion =36.27,P=0.00) and the lapse of acting time (Ftime =3.69 ,P=0.00).In cultured cells without dexamethasone for 24,48 and 72 hours,the mean expression levels of ICAM-1 were 0.298±0.008,0.312±0.003 and 0.319±0.011, showing a gradually increasing trend.However,the expression of ICAM-1 was gradually reduced with the increases of concentrations and the lapse of acting time of dexamethasone (Fconcentration =75.17,P=0.00;Ftime =3.11,P=0.00).Conclusions Occurrence and development of IOIP is probably associated with the over-expression of ICAM-1 in orbital fibroblasts.Dexamethasone plays anti-inflammation and treating effects on IOIP by down-regulating the expression of ICAM-1 and inhibiting the proliferation of orbital fibroblasts.

Comparative Study of Long-and Short-acting Triptorelin on Pituitary Down-regulation in Long Protocol / 昆明医科大学学报

Objective To explore the efficacy of long- and short-acting triptorelin on pituitary down-regulation in long protocol and the pregnancy outcome in vitro fertilization and embryo transfer (IVF-ET) . Methods Three hundred and seventeen patients for IVF-ET were enrolled as study and randomized them into two groups. In group A (n=145), patients received single dose subcutaneous injection of 1.25 mg long-acting diphereline in mid-luteal phase. In group B (n=172), patients received 0.1 mg/d subcutaneous injection of short-acting diphereline in mid-luteal phase for 14-18 days until pituitary suppression were got,and then reduced to 0.05 mg/d until the injection of HCG. Results The dose and the days of gonadotropin administration in group A were bigger and longer than those in group B ( 0.05) . But clinical pregnancy rate of group B had increasing trend. Conclusion Administration of short-acting diphereline has the similar effect with that of long-acting diphereline on pituitary down-regulation in long protocol. Short-acting diphereline requires lower amount of gonadotropin and is more flexible,so it should be recommended.

Down-regulation of human cytomegalovirus UL138, a novel latency-associated determinant, by hcmv-miR-UL36.

MicroRNAs (miRNAs) are small RNAs, 19–23 nucleotides in length, which regulate a variety of cellular processes. Human cytomegalovirus (HCMV) encodes only one intronic miRNA: human cytomegalovirus microRNA UL36 (hcmv-miR-UL36). In this study, we found that over-expression of hcmv-miR-UL36 resulted in a more than threefold increase in HCMV DNA synthesis at 24 h post infection. Fifteen putative targets of hcmv-miR-UL36 were identified using hybrid PCR, one being the HCMV UL138 gene that has previously been identified as a novel latency-associated determinant of HCMV infection. Down-regulation of UL138 expression by hcmv-miR-UL36 was validated using luciferase reporter assays and Western blot analysis in HEK293 cells. In the presence of hcmv-miR-UL36, we observed a 74.6% decrease in luciferase activity and a 46.2% decrease in HCMV UL138 protein expression. Our results indicate that hcmv-miR-UL36 may be a viral miRNA contributing to HCMV replication.

Prognostic Implication of 15-Hydroxyprostaglandin Dehydrogenase Down-Regulation in Patients with Colorectal Cancer

PURPOSE: Prostaglandin (PG) E2 is known to be closely related to cancer progression and is inactivated by 15-hydroxyprostaglandin dehydrogenase (PGDH). 15-PGDH is shown to have tumor suppressor activity and to be down-regulated in various cancers, including colorectal cancer (CRC). Therefore, we evaluated the expression of 15-PGDH and its prognostic effect in patients with CRC. METHODS: 15-PGDH expression was examined by using immunohistochemistry in 77 patients with CRC. Its prognostic significance was statistically evaluated. RESULTS: Negative 15-PGDH expression was noted in 55.8% of the 77 cases of CRC. 15-PGDH expression showed no correlation with any of the various clinicopathologic parameters. The status of lymph node metastasis, tumor-node-metastasis stages, and pre-operative carcinoembryonic antigen levels showed significant prognostic effect. However, univariate analysis revealed down-regulation of 15-PGDH not to be a predictor of poor survival. The 5-year overall survival rate was 71.7% in the group with positive expression of 15-PGDH and 67.1% in the group with negative expression of 15-PGDH, but this difference was not statistically significant (P = 0.751). CONCLUSION: 15-PGDH was down-regulated in 55.8% of the colorectal cancer patients. However, down-regulation of 15-PGDH showed no prognostic value in patients with CRC. Further larger scale or prospective studies are needed to clarify the prognostic effect of 15-PGDH down-regulation in patients with colorectal cancer.

Computational Analysis of miRNA and Target mRNA Interactions: Combined Effects of The Quantity and Quality of Their Binding Sites / 生物化学与生物物理进展

MicroRNAs (miRNAs) act by binding to complementary sites on target messenger RNA (mRNA) to induce mRNA degradation and/or translational repression. To investigate the influence of miRNAs at transcript levels, two human miRNAs (miR-1 and miR-124) were transfected into HeLa cells and microarrays used to examine changes in the mRNA profile showed that many genes were downregulated and that the fold decreases in levels of these target mRNAs differed remarkably. Features depicting interactions between miRNAs and their respective target mRNAs, such as the number of putative binding sites, the strength of complementary matches and the degree of stabilization of the binding duplex, were extracted and analyzed. It was found that, for a given target mRNA, both the quality and quantity of miRNA binding sites significantly affected its degree of destabilization. To delineate these types of interactions, a simple statistical model was proposed, which considers the combined effects of both the quality and quantity of miRNA binding sites on the degradation levels of target mRNAs. The analysis provides insights into how any animal miRNA might interact with its target mRNA. It will help us in designing more accurate methods for predicting miRNA targets and should improve understanding of the origins of miRNAs.

Computational Analysis of miRNA and Target mRNA Interactions:Combined Effects of The Quantity and Quality of Their Binding Sites / 生物化学与生物物理进展

MicroRNAs(miRNAs) act by binding to complementary sites on target messenger RNA(mRNA) to induce mRNA degradation and/or translational repression.To investigate the influence of miRNAs at transcript levels,two human miRNAs(miR-1 and miR-124) were transfected into HeLa cells and microarrays used to examine changes in the mRNA profile showed that many genes were downregulated and that the fold decreases in levels of these target mRNAs differed remarkably.Features depicting interactions between miRNAs and their respective target mRNAs,such as the number of putative binding sites,the strength of complementary matches and the degree of stabilization of the binding duplex,were extracted and analyzed.It was found that,for a given target mRNA,both the quality and quantity of miRNA binding sites significantly affected its degree of destabilization.To delineate these types of interactions,a simple statistical model was proposed,which considers the combined effects of both the quality and quantity of miRNA binding sites on the degradation levels of target mRNAs.The analysis provides insights into how any animal miRNA might interact with its target mRNA.It will help us in designing more accurate methods for predicting miRNA targets and should improve understanding of the origins of miRNAs.

Progress in the mechanism of post-traumatic insulin resistance / 肠外与肠内营养

Insulin resistance(IR) develops following major injury to the body, including serious trauma, surgical infection and burn. Metabolic disorders due to IR have a serious impact on energy production and attenuate body capacity of anti-infection and anti-shock. Therefore, the research about the mechanism of post-traumatic insulin resistance will be beneficial to improving patients' metabolism situation.

Research on the induction of T cell subset response via oral vaccination with liposome-mediated recombinant pCDNA3.1~+/Ag85A plasmid / 中国免疫学杂志

Objective:To prepare recombinant pCDNA3.1+/Ag85A DNA vaccine encoding tuberculosis Ag85A gene mediated with LipofectamineTM 2000 and to study the effect on T cell subpopulation via oral vaccination.Methods:Recombinant plasmid containing Ag85A using liposome as a vector was constructed and administered to C57BL/6 mice via oral route.Determination of the contents of IFN-r,IL-4 in serum of C57BL/6 mice was performed by double antibody sandwich ELISA.Furthermore,the ability of splenocytes to secret IFN-? and IL-4 was tested by ELISPOT method.When the mice were vaccinated with recombinant eukaryotic expressing vector 5 weeks later,titers of serum antibody against Ag85A were detected by ELISA.The percentage of the CD4+ and CD8+ T cell subsets in the splenocytes was determined by flow cytometry.Results:Lymphocytes obtained from the spleen of liposomal pcDNA3.1+/Ag85A vaccine-immunized mice exhibited lower IFN-? production and higher IL-4 production than those of pcDNA3.1+ vector immunized mice.The number of spleen MNC secreting IFN-? stimulated by Ag85A protein in vitro was significantly lower than that of plasmid vector group.Liposomal pcDNA3.1+/Ag85A vaccine immunized mice elicited higher Ag85A-specific antibodcy titres.The percentage of the CD4+ and CD8+ T cell subsets in the splenocytes was decreased.The subset both were shown the profile of Th2 responses.Conclusion:The oral recombinant plasmid pCDNA3.1+/Ag85A mediated with LipofectamineTM 2000 It shows down-regulation effect on the subsets of CD4+ T cells and CD8+ T cells.The regimen has good immunogenecity and could induce Th2 type humoral response in C57BL/6 mice.The immunization suppresses secretion of IFN-?.It can greatly enhance the titres of Ag85A-specific antibodies.LipofectamineTM 2000 can act as an adjuvant through comparation with negative control group.

Downregulation of Protein Kinase C Isozymes on Cholesteatoma / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: The exact pathogenesis of cholesteatoma remains unknown in spite of several theories that have been formulated. The most characteristic histologic finding of cholesteatoma is the proliferation of the squamous cell lining of the lesion. Protein Kinase C (PKC) is a family of phospholipid-dependent serine/threonin protein kinase that sends extracellular signals across the cell surface in order to regulate epithelial cell groweth and differentiation. This study attempted to provide the evidence for the role of PKC in cholesteatoma. MATERIALS AND METHODS: Twenty-five cholesteatoma specimens were obtained from patients for western blotting, immunohistochemical study, RT-PCR, and densitometry. RESULTS: The results of western blotting revealed that considerably lower levels of PKCalpha, PKCbeta, and PKCepsilon protein were detected in cholesteatoma than in the posterior auricular skin. In the immunohistochemical study, PKCalpha, PKCbeta, and PKCepsilon were detected both in the basal and suprabasal layer of posterior auricular skin, but they were not detectable in cholesteatoma. The results of PCR for PKCalpha, PKCbeta, and PKCepsilon showed that there were no differences between cholesteatoma and posterior auricular skin regarding the mRNA expression. CONCLUSION: Downregulation of PKCalpha, PKCbeta, and PKCepsilon in cholesteatoma suggests that abnormal epithelial growth is a possible mechanism of cholesteatoma. The results suggest the following there is an abnormal signal transduction through the PKC pathway in cholesteatoma: downregulation of PKC takes place in the post-transcription phase, and downregulation of PKC is associated with prolonged chronic inflammation of cholesteatoma.

Assessment of luteal rescue and desensitization of macaque corpus luteum brought about by human chorionic gonadotrophin and deglycosylated human chorionic gonadotrophin treatment.

The objective of the current study was to investigate the mechanism by which the corpus luteum (CL) of the monkey undergoes desensitization to luteinizing hormone following exposure to increasing concentration of human chorionic gonadotrophin (hCG) as it occurs in pregnancy. Female bonnet monkeys were injected (im) increasing doses of hCG or dghCG beginning from day 6 or 12 of the luteal phase for either 10 or 4 or 2 days. The day of oestrogen surge was considered as day '0' of luteal phase. Luteal cells obtained from CL of these animals were incubated with hCG (2 and 200 pg/ml) or dbcAMP (2.5, 25 and 100 μM) for 3h at 37°C and progesterone secreted was estimated. Corpora lutea of normal cycling monkeys on day 10/16/22 of the luteal phase were used as controls. In addition the in vivo response to CG and deglycosylated hCG (dghCG) was assessed by determining serum steroid profiles following their administration. hCG (from 15-90 IU) but not dghCG (15-90 IU) treatment in vivo significantly (P < 0·05) elevated serum progesterone and oestradiol levels. Serum progesterone, however, could not be maintained at a elevated level by continuous treatment with hCG (from day 6-15), the progesterone level declining beyond day 13 of luteal phase. Administering low doses of hCG (15-90 IU/day) from day 6-9 or high doses (600 IU/day) on days 8 and 9 of the luteal phase resulted in significant increase (about 10-fold over corresponding control P < 0·005) in the ability of luteal cells to synthesize progesterone (incubated controls) in vitro. The luteal cells of the treated animals responded to dbcAMP (P < 0·05) but not to hCC added in vitro. The in vitro response of luteal cells to added hCG was inhibited by 0, 50 and 100% if the animals were injected with low (15-90 IU) or medium (100 IU) between day 6-9 of luteal phase and high (600 IU on day 8 and 9 of luteal phase) doses of dghCG respectively; such treatment had no effect on responsivity of the cells to dbcAMP. The luteal cell responsiveness to dbcAMP in vitro was also blocked if hCG was administered for 10 days beginning day 6 of the luteal phase. Though short term hCG treatment during late luteal phase (from days 12-15) had no effect on luteal function, 10 day treatment beginning day 12 of luteal phase resulted in regain of in vitro responsiveness to both hCG (P < 0·05) and dbcAMP (P < 0·05) suggesting that luteal rescue can occur even at this late stage. In conclusion, desensitization of the CL to hCG appears to be governed by the dose/period for which it is exposed to hCG/dghCG. That desensitization is due to receptor occupancy is brought out by the fact that (i) this can be achieved by giving a larger dose of hCG over a 2 day period instead of a lower dose of the hormone for a longer (4 to 10 days) period and (ii) the effect can largely be reproduced by using dghCG instead of hCG to block the receptor sites. It appears that to achieve desensitization to dbcAMP also it is necessary to expose the luteal cell to relatively high dose of hCG for more than 4 days.

Modification of Epidermal Growth Factor Receptor (EGFR) Expression after Aminoglycoside-induced Acute Tubular Necrosis in Rat / 대한비뇨기과학회지

Acute tubular necrosis induced by aminoglycoside-antibiotics is followed by a regenerative Process which leads to the restoration of damaged tubules. It is well known that tubular regeneration is mediated by polypeptide growth factors such as epidermal growth factor (EGF) In the absence of nephrotoxic alterations, EGF is immunolocalized in distal tubules, whereas epidermal growth factor receptor(EGFR) immunostaining is localized in proximal tubules. After acute tubular necrosis, the sign of regeneration is accompanied by redistribution of EGF immunoreactivity from distal to proximal tubules and a reduction of total immunoreactive EGF due to a decrease of tissue-bound proEGF. However, it is controversial whether EGFR is down- or up-regulated during this regenerative process. We evaluated the time course of the regenerative response subsequent to tubular damage induced by aminoglycoside, with a particular attention paid to EGFR in order to examine whether it is down- or up-regulated. Female Sprague-Dawley rats were treated during 4 and 8 consecutive days with a daily dose of 80 mg/kg gentamicin i.p. Groups of experimental animals (n=10) were terminated at increasing time intervals (5, 9, 12, 16 days) after cessation of treatment. One hour before sacrifice, each individual received i.p. 200mg/kg Blood for the immunohistochemical demonstration of cell proliferation (S-phase cell). Blood was collected at the time of sacrifice to measure serum creatinine and BUN levels. EGFR immunoreactivity was revealed on paraffin-embedded tissue section by immunohistochemical staining using polyclonal anti-EGFR antibody. Upon immunostaining sections in control kidneys, immunoreactive EGFRs exhibited a quite specific and restricted distribution since they were confined to proximal tubules. But proximal tubules undergoing regenerative repair were characterized by a disappearance of EGFR, which expressed BrdU in immunohistochemical sections for BrdU. Beyond the episode of tubular regeneration, proximal tubules recovered immunoreactive EGFR. The results suggest that the apparent loss of EGFR could be due to a process of receptor down-regulation in proximal tubules displaying evidence of regenerative response.