Effect of Silencing DNMT1 gene on histone methylation modulation, cell proliferation and apoptosis in Molt-4 cell line / 中国药理学通报

Aim To investigate the effect of small in-terfering RNA(siRNA) targeting DNMT1 gene on cell proliferation, apoptosis and histone modulation in acute lymphoid leukemia cell line, Molt-4. Methods The small interfering RNA targeting DNMT1 gene was transfected into Molt-4 cells by LipofectamineTM 2000. The DNMT1 mRNA and protein level were detected by RT-PCR and Western blot. Cell proliferation was de-termined by MTT. Cell apoptosis was measured by Flow Cytometry. The expression of Bcl-2, procaspase-3, P15, histone methylation and histone acetylation was detected by Western blot. Results DNMT1 was suppressed by siRNA targeting DNMT1 in a concentra-tion-dependent manner. DNMT1 siRNA suppressed cells proliferation and induced apoptosis in Molt-4 cells. Apoptotic rate was (4. 27 ± 1. 42)% , (15. 25 ± 1. 54)% , (35. 63 ± 2. 54)% , (66. 27 ± 3. 02)%after transfecting with DNMT1 siRNA at 0, 30, 60, 120 nmol·L - 1 for 24 hours, P < 0. 05. The expres-sion of Bcl-2, procaspase-3 was suppressed and P15 was promoted after transfecting of DNMT1 siRNA. DN-MT1 siRNA downregulated histone methylated H3K9 and upregulated histone methylated H3K4. The altera-tion of histone acetylation of H3 was not seen. Conclu-sion DNMT1 siRNA suppresses DNMT1 efficiently in Molt-4 cells. The depletion of DNMT1 downregulates histone methylation of H3K9, and upregulates histone methylation of H3K4. It inhibits cell growth and in-duces cell apoptosis in Molt-4 cell line.

Antiproliferative effect of silencing LSD1 gene on Jurkat cell line and its mechanism / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of silencing LSD1 gene by RNA interference on the proliferation, apoptosis on human lymphocytic leukemia Jurkat cell line and its mechanism.</p><p><b>METHODS</b>The hairpin- like oligonucleotide sequences targeting LSD1 gene was transfected into Jurkat cells by lipofectamine(TM) 2000. The LSD1 mRNA and protein were detected by RQ- PCR and Western blot. Cell growth was determined by MTT. Cell apoptosis was analyzed by flow cytometry. The expression of Bcl-2, Bax, procaspase- 3, and histone H3K4me, H3K4me2, H3K4me3, Act- H3, H3K9me were detected by Western blot.</p><p><b>RESULTS</b>LSD1 mRNA was markedly suppressed by the shRNA targeting LSD1. LSD1 shRNA suppressed the proliferation and induced cells apoptosis of Jurkat cells. The cell apoptotic rate was (41.34±3.58)%, (3.45±1.54)%, (1.76±0.52)% in LSD1 shRNA, Neg-shRNA and Blank respectively, the difference among them was statistically significant (P<0.05). LSD1 shRNA down- regulated the expressions of Bcl- 2 and procaspase- 3, and up- regulated the expression of Bax. The methylation of H3K4me1, me2 and acetylation of Act- H3 improved without change of the methylation of H3K4me3.</p><p><b>CONCLUSIONS</b>Deplete of LSD1 gene maybe through modifying the methylation of histone H3K4 to promote the cell apoptosis and inhibit cell growth in Jurkat cell line.</p>

Effect of phenelzine on the proliferation, apoptosis and histone methylation and acetylation of Molt-4 cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of monoamine oxidase inhibitor phenelzine on proliferation, apoptosis and histone modulation in acute lymphoblastic leukemia cell line Molt-4 cells.</p><p><b>METHODS</b>The effect of Phenelzine on cell proliferation was detected by MTT. Apoptotic rate was measured by flow cytometry. The variation of apoptosis associated proteins Caspase-3, Bcl-2 and Bax, cyclin-dependent kinase inhibitor p21, tumor suppressor protein p15, and the expression level of histone methylation of H3K4, H3K9 and histone acetylation of H3, DNMT1 were detected by Western Blot.</p><p><b>RESULTS</b>① Molt-4 cell proliferation rates were (87.68±3.54)%, (67.84±3.24)%, (51.48±3.37)%, (28.72±2.56)% respectively after exposured to phenelzine at 5, 10, 15, 20 μmol/L for 24 h, P<0.05. ② After 10 μmol/L of phenelzine exposure for 24, 48, 72 h, cell proliferation rates were (67.84±3.24)%, (50.24±2.01)%, (40.31±2.25)%, P<0.05. ③ The apoptotic rates were (13.64±2.58)%, (31.24±3.42)%, (56.37±4.26)% after phenelzine treatment at 5, 10, 20 μmol/L for 24 h, which was concentration dependent. ④ Phenelzine could upregulate the expression of Bax, caspase-3, p21, and downregulate Bcl-2 expression. Phenelzine upregulated the methylation level of histone H3K4me1, H3K4me2 and histone acetylated H3, while it didn't change the level of histone H3K4me3, H3K9me1, H3K9me2. ⑤ Phenelzine inhibited DNMT1 expression and promoted p15 expression.</p><p><b>CONCLUSIONS</b>Phenelzine increased the methylation of histone H3K4me1, H3K4me2, acetylation of histone H3 and p21, and decreased the expression of DNMT1 and p15, and ultimately inhibited the proliferation and apoptosis of Molt-4 cells.</p>

Effects of silencing LSD1 gene on Molt-4 cells apoptosis / 中国药理学通报

Aim To observe the effect of the LSD1 gene on the proliferation and apoptosis of Molt-4 cells, a kind of human acute T-lymphoblastic leukemia cells. Methods siRNA fragment based on LSD1 gene was designed, filtered out and then transfected into Molt-4 cells. The effects of LSD 1 siRNA on Molt-4 cell prolif-eration were observed by the method of MTS. The cell apoptosis was analyzed by flow cytometry. The states of histone H3K4, H3K9 methylation, histone H3 acetyla-tion, p15, DNA methyltransferase 1 (DNMT1), and apoptosis-related proteins like Bcl-2 , procaspase-3 were evaluated by Western blot. Results Silencing LSD1 gene inhibited cell proliferation. Molt-4 cell pro-liferation rate was ( 99. 65 ± 1. 21 )%, ( 83. 02 ± 1. 69)%, (65. 72 ± 2. 16)%,and (41. 15 ± 2. 23)%respectively after the treatment of Molt-4 cells with 0 , 30, 60, 120 nmol·L-1 of LSD1 siRNA after 48 hours ( P < 0. 05 ) . Cell proliferation rate was ( 99. 86 ± 1. 35)%,(65. 72 ± 2. 16)%,(48. 26 ± 1. 92)%,and ( 37. 86 ± 1. 66 )% respectively after the transfection of Molt-4 cells with 60 nmol · L-1 of LSD1 siRNA after 0 , 24 , 48 , 72 hours ( P<0. 05 ) . Cell apoptosis rate was ( 3. 35 ± 1. 26 )%, ( 12. 16 ± 1. 74 )%, ( 32. 74 ± 2. 47 )%, ( 54. 64 ± 2. 58 )% respectively after transfection of LSD1 siRNA in indicated concentrations for 48 hours ( P <0. 05 ) . At the same time, the ex-pression levels of apoptosis-related proteins like Bcl-2 , procaspase-3 decreased. LSD1 siRNA inhibited LSD1 and LSD1 mRNA, and accumulated histone mono-, and di-methylation H3K4 and histone H3 acetylation. However, alteration of H3K4 trimethylation, H3K9 methylation was not detected. LSD1 siRNA downregu-lated DNA demethylase DNMT1 and upregulated p15 . Conclusions LSD1 siRNA can inhibit Molt-4 cell proliferation and induce apoptosis. Its mechanism may be associated with epigenetic regulation. In addition, it is expected to become a new target for leukemia treat-ment.

Clinical effect of different scheme of moxifloxacin and levofloxacin for elderly multi drug resistant pulmonary tuberculosis / 中国基层医药

Objective To explore the clinical effect of different scheme of moxifloxacin and levofloxacin for elderly multi drug resistant pulmonary tuberculosis.Methods 136 cases of elderly patients included in the study were randomly divided into the observation group 1 and observation group 2 with 68 cases in each group according to the sequence in group.The observation group 1 used moxifloxacin regimen for treatment of pulmonary tuberculosis while the observation group 2 used levofloxacin regimen.The two groups were treated for 18 months and observed the clinical curative effect,and the sputum negative,pulmonary lesions absorption,empty changes and adverse reactions.Results The total efficiency of the observation group 1 was 92.6％,which in the observation group 2 was 72.1％,the total efficiency of the two groups had a significant difference (x2 =9.917,P =0.002) ;the sputum negative conversion rate of the observation group 1 at the end of 3 months was significantly higher than that of the observation group 2 (x2 =4.115,P =0.043),no significant difference was found at the other time points (all P ＞ 0.05) ; after treatment,the obvious absorption + absorption of lung lesion in the observation group 1 was 59 cases,accounting for 86.8％,that in observation group 2 was 48 cases,accounting for 70.6％,two groups of lung lesions absorption had significant difference (x2 =5.303,P =0.021) ; the pulmonary cavity closure + reduced in the observation group 1 was 52 cases,accounting for 76.4％,that in the observation group 2 was 41 cases,accounting for 60.3％,with significant difference between the two groups (x2 =4.115,P =0.043) ;there was no significant difference in adverse reaction of the two groups (P ＞ 0.05).Conclusion Moxifloxacin anti tuberculosis regimen has a better curative effect in treatment of elderly patients with multi drug resistant pulmonary tuberculosis,which should be expanded the application.

Antiproliferative effect of silencing mTOR gene on MCL Jeko-1 cell line and its mechanism / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of silencing mTOR gene by RNA interference on proliferation and apoptosis, and its mechanism on mantle cell lymphoma Jeko-1 cell Line.</p><p><b>METHODS</b>The hairpin-like oligonucleotide sequences targeting mTOR gene was designed and transfected into Jeko-1 cells by lipofectamine TM 2000. The mTOR mRNA and protein were detected by RQ-PCR and Western blot. Cell growth was determined by MTT. Cell apoptosis was analyzed by flow cytometry. The expressions of Bcl-2, Bax, procaspase-3, procaspase-9, P70S6K,and p-P70S6K were detected by Western blot.</p><p><b>RESULTS</b>mTOR mRNA was markedly suppressed by shRNA targeting mTOR. mTOR shRNA suppressed proliferation and induced cells apoptosis of Jeko-1 cells. The cell apoptotic rates were (36.62 ± 3.24)%, (2.58 ± 1.04)%, (1.24 ± 0.30)% respectively, in mTOR shRNA, Neg-shRNA and Blank with statistically significant difference among them (P<0.05). mTOR shRNA down-regulated the expressions of Bcl-2, proCaspase3, proCaspase9 and p-70S6K, up-regulated the expression of Bax.</p><p><b>CONCLUSION</b>Deplete of mTOR gene may be realized through inhibiting the Akt/mTOR signaling pathway to promote the cell apoptosis and inhibit cell growth in Jeko-1 cell line.</p>

Effect of BIX-01294 on cell proliferation, apoptosis and histone methylation in Molt-4 line / 中国药理学通报

Aim To investigate the effect of the specif-ic inhibitor BIX-01294 of G9a on the proliferation, ap-optosis,DNA methylation and histone modulation of a-cute leukemia cell line, Molt-4. Methods Cells were cultured with different concentrations of BIX-01294 . Cell growth was determined by MTT. Cell apoptosis was analyzed by flow cytometry. The expression of Caspase-3, Bcl-2, Bax, P15, acetylated H3, H3 K9 me1 , H3 K9 me2 , H3 K9 me3 and H3 K27 me1 , H3K27me2 methylation were detected by Western blot. Results BIX-01294 downregulated bcl-2 , and upreg-ulated Bax, Caspase-3 and induced apoptosis in (5. 54 ± 1. 35)%, (10. 24 ± 2. 26)%, (32. 28 ± 3. 26)%, (47. 52 ± 4. 37 )% after 24 hours exposure to BIX-01294 in 0 , 1 , 2 , 4 μmol · L-1 . The difference be-tween them was statistically significant ( P <0. 05 ) . BIX-01294 inhibited DMNT1 and promoted P15 , which resulted in cell proliferation inhibition. Further studies showed that BIX-01294 decreased H3 K9 me1 , H3 K9 me2 , and H3 K27 me1 , H3 K27 me2 , and didn′t change protein expression of acetylated H3 and H3 K9 me3. Conclusion BIX-01294 inhibits G9a, resulting in downregulation of methylation of H3 K9 me1 , H3 K9 me2 , H3 K27 me1 and H3 K27 me2 and DMNT1 and P15 denovo. It inhibits cell proliferation and in-duces cell apoptosis.

Effect of downregulation the expression of HDAC1 on cells differentiation of HL-60 cells / 药学学报

This study is to investigate the effect of downregulation histone deacetylases 1 (HDAC1) gene by the technology of RNA interference on the differentiation of HL-60 cells line. The optimal segment targeting HDAC1 gene was designed and transfected into HL-60 cells by Lipofectamine 2000. The HDAC1 mRNA and protein level were detected by RT-PCR and Western blotting. The morphologic change of HL-60 cells was detected by an optical microscope with Wright-Giemsa. Cell differentiation was tested by NBT reduction assay. Expression of CD13, CD33 and CD14 was measured by flow cytometry. The results indicated that HDAC1 mRNA and protein were markedly suppressed by the siRNA targeting HDAC1 in a concentration-dependent manner. HDAC1 siRNA promoted cell differentiation. HL-60 cells became more mature in morphology after transfected to HDAC1 siRNA at a concentration of 30-60 nmol x L(-1) for 24 h. NBT reduction ability of HDAC1 siRNA with 30 nmol x L(-1) was 0.31 +/- 0.09, compared with negative control (0.20 +/- 0.02) (t = -3.1, P or = 120 nmol x L(-1). After transfection with 60 nmol x L(-1) HDAC1 siRNA to HL-60 cells, the expression of CD13 was (96.50 +/- 0.70)% in compared to siRNA-NC (3.39 +/- 0.68) % (t = 164.9, P 0.1). HDAC1 siRNA promoted cell differentiation in indicated concentration. HDAC1 might be one of the targets of gene therapy for leukemia.

Study on histone acetylation modulation and Akt signaling pathway inhibition by phenyhexyle isothiocyanate in prostate cancer PC3 cell line / 中华泌尿外科杂志

Objective To investigate phenyhexyle isothiocyanate (PHI) modulating histone acetylation and inhibiting Akt signaling pathway in prostate cancer cell line PC3 in vitro. Methods Apoptotic cells were measured by TUNEL assay. Histone acetylated H3, H4 and the Akt protein signaling pathway (Akt, p-Akt, mTOR, p-mTOR, p70S6K and p-p70S6K) were detected by Western blot. Results Apoptotic cells increased after exposure to PHI with concentration dependent. PHI significantly induced an accumulation of histone acetylated H3, H4. The change of Akt, mTOR, p70S6K proteins was not observed. Phosphorylation of Akt (p- Akt), mTOR (p-mTOR) and p70S6K (p-p70S6K) decreased after exposure to PHI for 7 h. Conclusions PHI can induce histone acetylation H3, H4 accumulation. PHI inhibits Akt signaling pathway resulting cell apoptosis. It might be a new anticancer agent.

Inhibition of gene p15 hypermethylation by phenylhexyl isothiocyanate in Molt-4 cells / 白血病·淋巴瘤

Objective To investigate the effect of phenylhexyle isothiocyanate (PHI) on demethylation and activation of transcription gene p15 in acute leukemia cell line Molt-4. Methods DNA sequencing and modified methylation specific PCR (MSP) were used to screen p15-M and p15-U mRNA after Moh-4 cells were treated with PHI. P15 mRNA was measured by RT-PCR. Pl5 protein was detected by Western blotting. Results Hypermethylation of gene pl5 was apparently attenuated and activation of transcription p15 gene was de novo after 5 days exposure to PHI. PHI enhanced both the expression of p15 mRNA and p15 protein in a concentration-dependent manner. The ratio of the gray scale of p15 mRNA strap was 0.17±0.12 in control, 0.29±0.14 in PHI 10 μmol/L, 0.55±0.07 in PHI 20 μmol/L, 0.93±0.13 in PHI 40 μmol/L. Conclusion PHI could active demethylation and transcription of gene p15.

Study on irregulatory modification of histone acetylation, methylation in diffuse large B-cell lymphoma / 白血病·淋巴瘤

Objective To investigate the expression of histone acetylated H3 and H4, methylated H3K4 and H3K9 in diffuse large B-cell lymphoma (DLBCL). Methods The expression of histone acetylated H3 and H4, methylated H3K4 and H3K9 were examined by SP immunohistochemistry technique in lymphoid tissue of 40 cases with DLBCL and 16 cases with proliferative lymphadenitis. Results The expression of histone acetylation of H3 and H4 were lower than that in proliferative lymphadenitis. Histone methylated H3K4 was lower in expression and H3K9 was in higher expression. There was a positive correlative expression between the global histone acetylation of H3 and H4, the global histone acetylation of H3, H4 and histone methylation of H3K4. Conclusion Improper modification of histone acetylations and methylations may play an important role in pathogenesis in DLBCL.

Clinical study of treatment of chronic hepatitis B with kurorinone combined with thymosin / 解放军医学杂志

Objective To evaluate the effects of kurorinone combined with thymosin on the treatment of chronic hepatitis B. Methods 178 patients with chronic hepatitis B were randomly divided into group A, B and C. 64 patients in group A received kurorinone combined with thymosin for 3 months. 58 patients in group B received kurorinone only, and 56 patients in group C received thymosin only. Results At the end of treatment, HBeAg and HBV DNA negative-transformed rates were respectively 51.8% and 53.6% in group A, but 32.7% and 34.6% in group B, 21.6% and 21.6% in group C, the differences were statistically significant (P

Roles of TNF-?,Endotoxin in Upper Gastrointestinal Bleeding in Cirrhosis Patients and the Preventive and Treatment Mechanism of Rhubarb / 中国医师杂志

Objective To explore the role of tumor necrosis factor (TNF-?) and endotoxin(LPS) in upper gastrointestinal bleeding in cirrhosis patients and the preventive and treatment mechanism of rhubarb. Methods 45 liver cirrhosis patients with upper gastrointestinal bleeding were treated with rhubarb, and its efficacy was compared with cemetidine's efficacy. Liver function, and serum levels of TNF-? and LPS were measured before and after the therapy. Results Patients’symptoms and liver function were improved, and serum levels of TNF-? and LPS significantly decreased after treated with rhubarb. Conclusions TNF-? and endotoxin might play an important role in upper gastrointestinal bleeding in cirrhosis patients. Rhubarb has protective and treatment effect on upper gastrointestinal bleeding in cirrhosis patients by reducing TNF-? and LPS release.

The Effect of Kurorinone on the Serum Level of TNF-? and IL-6 in patients with Chronic Hepatitis B / 中国医师杂志

Objective To explore the effects of kurorinone on the serum level of TNF-? and IL-6 in the patients with chronic hepatitis B (CHB). Methods 87 patients with CHB were randomly divided into groups A and B. Patients in group A (n=45) and group B (n=42) received kurorinone and diammonium glycyrrhizin treatment, respectively, for 3 months. The liver histopathological changes were observed, and liver function and the serum level of TNF-? and IL-6 in the patients with CHB were detected after treatment. Results Compared with group B, liver injury and liver function of the patients in group A were obviously ameliorated, and the serum level of TNF-? and IL-6 significantly decreased (P

Experimental study of the therapeutic effect of interferon-? on liver fibrosis in rats / 中华传染病杂志

0.05 ), However, all pasameters showed significantly milder or lower in interferon ? treated group than incontrol group at the 6th, 12th week( P

Clinical study of the interferon ?-2b combined with kurorinone in the treatment of chronic hepatitis B / 中华传染病杂志

Objective To study the efficacy of interferon ? 2b (IFN? 2b) combined with kurorinone in the treatment of chronic hepatitis B. Methods 146 patients with chronic hepatitis B were randomly divided into four groups(A,B,C,D). Based on the similar general treatment(group D),37 patients in group A was treated with IFN ? 2b and kurorinone. However, 37 patients in group B and 36 patients in group C received IFN ? 2b and kurorinone,respectively.Dynamic changes and relationships among liver histopathology,expression of TGF ? in liver tissue and serum levels of TGF ?,HBeAg,HBV DNA were studied using immunohistochemistry,radioimmunoassay and ELISA before and after treatment of IFN ? 2b and kurorinone. Results The score of histological hepatitic fibrosis,expression of TGF ? in liver tissue and serum levels of TGF ?,HBeAg, HBV DNA in group A showed significant lower than in group B and group C ( P