BACKGROUND: Radiation (IR)-induced DNA damage triggers cell cycle arrest and has a suppressive effect on the tumor microenvironment (TME). Wee1, a cell cycle regulator, can eliminate G2/M arrest by phosphorylating cyclin-dependent kinase 1 (CDK1). Meanwhile, programed death-1/programed death ligand-1 (PD-1/PDL-1) blockade is closely related to TME. This study aims to investigate the effects and mechanisms of Wee1 inhibitor AZD1775 and anti-PD-1 antibody (anti-PD-1 Ab) on radiosensitization of hepatoma. METHODS: The anti-tumor activity of AZD1775 and IR was determined by 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT) assay on human and mouse hepatoma cells HepG2, Hepa1-6, and H22. The anti-hepatoma mechanism of AZD1775 and IR revealed by flow cytometry and Western blot in vitro . A hepatoma subcutaneous xenograft mice model was constructed on Balb/c mice, which were divided into control group, IR group, AZD1775 group, IR + AZD1775 group, IR + anti-PD-1 Ab group, and the IR + AZD1775 + anti-PD-1 Ab group. Cytotoxic CD8 + T cells in TME were analyzed by flow cytometry. RESULTS: Combining IR with AZD1775 synergistically reduced the viability of hepatoma cells in vitro . AZD1775 exhibited antitumor effects by decreasing CDK1 phosphorylation to reverse the IR-induced G2/M arrest and increasing IR-induced DNA damage. AZD1775 treatment also reduced the proportion of PD-1 + /CD8 + T cells in the spleen of hepatoma subcutaneous xenograft mice. Further studies revealed that AZD1775 and anti-PD-1 Ab could enhance the radiosensitivity of hepatoma by enhancing the levels of interferon γ (IFNγ) + or Ki67 + CD8 T cells and decreasing the levels of CD8 + Tregs cells in the tumor and spleen of the hepatoma mice model, indicating that the improvement of TME was manifested by increasing the cytotoxic factor IFNγ expression, enhancing CD8 + T cells proliferation, and weakening CD8 + T cells depletion. CONCLUSIONS This work suggests that AZD1775 and anti-PD-1 Ab synergistically sensitize hepatoma to radiotherapy by enhancing IR-induced DNA damage and improving cytotoxic CD8 + T cells in TME.
Humans ; Animals ; Mice ; Carcinoma, Hepatocellular/radiotherapy* ; Cell Cycle Proteins/metabolism* ; Protein-Tyrosine Kinases/genetics* ; Apoptosis ; Programmed Cell Death 1 Receptor ; Cell Line, Tumor ; G2 Phase Cell Cycle Checkpoints ; Liver Neoplasms/radiotherapy* ; Tumor Microenvironment ; Pyrazoles ; Pyrimidinones

[摘 要] 肠道微生物是存在于人体内的庞大生态系统，它与机体形成一个相互关联的共存体。近年来得益于分子生物技术的蓬勃发展，肠道微生物组学研究表明，肠道微生物在很大程度上左右了肿瘤的发生与发展，除了微生物本身的直接作用之外，由它们引起的宿主炎症免疫系统、代谢功能方面的改变也间接发挥了重要作用，而这些变化对于后续抗肿瘤治疗也产生一定影响。基于肠道微生物的重要作用，在此基础上开发出来的微生态制剂在抗肿瘤治疗中的临床应用价值也逐渐显现，可为今后的抗肿瘤治疗提供辅助作用。本文从肠道微生物的地位、肠道微生物影响肿瘤发展的机制和对抗肿瘤治疗的影响，以及肠道微生物的临床应用等四个方面展开论述，为基于肠道微生物的抗肿瘤制剂技术的研发和临床应用提供新的思路和启示。

The aim of the present study was to investigate whether the physiological features of Ano1 were affected by enhanced green fluorescent protein (EGFP) fusing at Ano1 C-terminal. The eukaryotic expression vectors of Ano1 and EGFP-Ano1 were constructed, and these plasmids were transfected into Fischer rat thyroid follicular epithelial (FRT) cells using liposome. The expression and location of Ano1 were examined by using inverted fluorescence microscope. The ability of Ano1 to transport iodide was detected by kinetics experiment of fluorescence quenching. The results showed that both Ano1 and EGFP-Ano1 were expressed on FRT cell membrane and could be activated by Ca(2+). There was no significant difference of the ability to transport iodide between Ano1 and EGFP-Ano1. These results suggest Ano1 and EGFP-Ano1 have similar physiological feature.
Animals ; Anoctamin-1 ; Cell Membrane ; physiology ; Chloride Channels ; metabolism ; Epithelial Cells ; physiology ; Genetic Vectors ; Green Fluorescent Proteins ; metabolism ; Microscopy, Fluorescence ; Plasmids ; Rats ; Recombinant Fusion Proteins ; metabolism ; Thyroid Gland ; cytology ; Transfection

OBJECTIVE: To establish the hairy roots culture system of Atractylodes japonica Koidz. ez Kitam. and study the hairy roots growth and analyze the polysaccharide content in hairy roots culturing system.

Objective: To study the chemical constituents from the dried rhizome of Lepidium meyenii (Maca) cultivated in Jilin. Methods: The chemical constituents were isolated and purified with macroreticular resin, silica gel, ODS column chromatography, preparation HPLC, etc. The structures of the compounds were identified by physicochemical properties and various spectroscopic methods. Results: Thirteen compounds were isolated as N-methyl-3-hydroxy-benzeneacetamide (1), N-benzyloctadecanamide (2), benzeneacetamide (3), benzylamine (4), 3-methoxyphenylacetic acid (5), phenylacetic acid (6), 4-hydroxy-3-methoxybenzoic acid (7), nicotinic acid (8), 3, 4-dihydroxy-benzoic acid methyl ester (9), adenosine (10), L-valine (11), daueosterol (12), and β-sitosterol (13). Conclusion: Compound 1 is identifled as a new natural compound, and compounds 3-6 and 9-12 are obtained from L. meyenii for the first time.

Objective: To explore the effect of ginsenoside Rgl on the ubiquitin-modified protein aggregation in the cortex after cerebral ischemia reperfusion (I / R) injury in the rats, and to further clarify the therapeutic mechanism of ginsenoside Rgl in the cerebral I/R injury. Methods: The middle cerebral artery occlusion (MCAO) model was set up with suture method for 1. 5 h of embolization. A total of 72 rats were divided into sham operation group, I/R model group, positive drug control (nimodipine) group, low, middle, and high doses 10, 20, and 40 mg ' k g - 1) of ginsenoside Rgl groups. All 12 rats in each group were given intraperitoneal injection. TTC staining and Longa' s score method were used to detect the infarction areas and the neurological deficit scores of the rats in various groups 24 h after modeling. The death of neurons in the cortex and hippocampus after cerebral ischemia of the rats in various groups were observed with HE staining. Immunohistochemistry and Western blotting method were used to detect the expression of ubiquitin-modified protein aggregation in the cortex of the rats in various groups. Results: Compared with I/R group, the percentages of infarction areas of the rats in nimodipine group and ginsenoside Rgl groups were significantly decreased (P < 0 . 05). and the neurological deficit scores were decreased (P < 0 . 05). The HE staining results showed that compared with sham operation group, the neurons in I/R model group were sparse, showing fragmentation and dissolution; compared with I/R model group, the phenomena of cell nucleus fragmentation, dissolution and powder staining in nimodipine group and different doses of ginsenoside Rgl groups were all improved to different degrees. The immunohistochemical results showed that compared with sham operation group, the positive expression level of ubiquitin-modified protein in I/R model group was increased significantly (P < 0 . 05); compared with I/R model group, the positive expression levels of ubiquitin-modified protein in nimodipine group and different doses of ginsenoside Rgl groups were decreased (P < 0 . 05), especially in high dose of ginsenoside Rgl group (P < 0 . 05). The Western blotting results showed that compared with sham operation group, the level of ubiquitin-modified protein aggregates in I/R model group was significantly increased (P < 0 . 0 5); compared with I/R model group, the levels of ubiquitin-modified protein aggregates in nimodipine group and different doses of ginsenoside Rgl were decreased (P < 0 . 05), especially in high dose of ginsenoside Rgl group. Conclusion: Ginsenoside Rgl can inhibit the formation of ubiquitin-modified protein aggregates induced by I/R injury in the cortex, thereby alleviating the I/R injury in the rats.

Many pathological phenomena of male infertility are related to epigenetic changes in male germ cells. Epigenetic regulation during spermatogenesis plays an important role in mitotic/meiotic divisions and spermiogenesis. The histones have various post-translational modifications on different amino acid residues during spermatogenesis. These modifications are crucial to the precise regulation of spermatogenesis. Moreover, the histone-to-protamine transition will occur during spermiogenesis. Many studies have also found that abnormal changes of histone modifications during spermatogenesis may damage the sperm development, leading to male sterility. This article reviews the changes of histone modifications during spermatogenesis, the regulation of the development of male germ cells, and the relationship between histone abnormalities and male sterility.
Epigenesis, Genetic ; Histones ; metabolism ; Humans ; Infertility, Male ; physiopathology ; Male ; Spermatogenesis

The aim of the present study was to establish a cell model of volume-regulated anion channel subunit LRRC8A and investigate the physiological characteristics of LRRC8A. The eukaryotic expression vectors of LRRC8A and YFP-H148Q/I152L were constructed and transfected into Fischer rat thyroid (FRT) cells by Lipofectamine 2000. The FRT cell lines co-expressing LRRC8A and YFP-H148Q/I152L were obtained by antibiotic screening. The expression of LRRC8A and YFP-H148Q/I152L in FRT cells was detected by the inverted fluorescence microscope. The fluorescence quenching kinetic experiment was done to verify the function and effectiveness of the cell model. Then the cell model was utilized to study the physiological characteristics of LRRC8A, such as the characteristics of anion transport, the opening of LRRC8A by osmotic pressure, the effect of anion transport velocity, and the effect of chloride channel inhibitors on LRRC8A anion channel. The results of the inverted fluorescence microscope showed that LRRC8A was expressed on the cell membrane and YFP-H148Q/I152L was expressed in the cytoplasm. The results of fluorescence quenching kinetic test showed that under the condition of low osmotic state, LRRC8A could transport some kinds of anions, such as iodine and chloride ions. Osmotic pressure played a key role in the regulation of LRRC8A volume-regulated anion channel opening. Chloride channel inhibitors inhibited ion transport of LRRC8A channel in a dose-dependent manner. It is suggested that LRRC8A has the characteristics of classic volume-regulated anion channels by using the cell model of FRT cells co-expressing LRRC8A and YFP-H148Q/I152L.
Animals ; Anions ; Cells, Cultured ; Chloride Channels ; antagonists & inhibitors ; Ion Transport ; Membrane Proteins ; physiology ; Microscopy, Fluorescence ; Rats ; Rats, Inbred F344 ; Thyroid Gland ; cytology ; Transfection

OBJECTIVETo study therapeutic effects of Sky-bone expander for the treatment of osteoporotic vertebral compressed fractures.

METHODSFifteen patients (18 vertebrae) suffering from vertebral compression fractures were treated with Sky bone expander system which expanded and reconstructed the vertebral body with PMMA cement. The clinical effect was evaluated by observing the changing of visual analogue scale (VAS). The preoperative and postoperative mean VAS scores were compared by paired-sample t test. All the patients were followed up by telephone or clinic consulting after being discharged from our hospital.

RESULTSThe procedure was performed successfully in 15 patients. The operation time ranged from 45 to 110 minutes (65 minutes per vertebra on average). The patients were followed up and the duration ranged from 6 to 12 months (8 months on average). The mean VAS score of the patients were improved significantly at the third postoperative day compared with those before the operation (2.5 +/- 1.3, vs 7.7 +/- 1.1, all P < 0.05). The mean VAS score at the end of the follow-up was 2.2 +/- 1.2.

CONCLUSIONSky bone expander system provides significant pain relief effect in the cases of osteoporotic vertebral compression fractures, shortens the duration of lying in bed, and its procedure is convenient with few complications.

Aged ; Female ; Fractures, Compression ; surgery ; Humans ; Male ; Middle Aged ; Osteoporosis ; complications ; Pain Measurement ; Spinal Fractures ; surgery ; Vertebroplasty

BACKGROUNDLung cancer is one of the most common cancers worldwide and its mortality rate has increased year after year. Molecular biology has contributed to make people's understanding to the disease at the gene level. A new idea will be given to the early diagnosis and treatment for lung cancer with the method of molecular biology which can be used to find the association between some genes and lung cancer. The aim of this study is to evaluate the relationship between polymorphism of codon 72 (BstU I single nucleotide polymorphism, BstU I SNP) in p53 gene and susceptibility and radiosensitivity of non-small cell lung cancer (NSCLC) in Chinese population.

METHODSThe BstU I single nucleotide polymorphic sites at condon 72 in exon 4 of p53 gene in 50 patients with NSCLC as well as 50 healthy controls were inspected by polymerase chain reaction-restricted fragment length polymorphism assay and the relationship between BstU I SNP and susceptibility and radiosensitivity of NSCLC was analysed by case-control test.

RESULTSThe allelic distribution of the three genotypes (A1/A1, A1/A2, A2/A2) in healthy controls was 32.0%, 42.0% and 26.0% respectively, which differed slightly from that of lung cancer patients, which was 28.0%, 32.0% and 40.0%. These allelic and genotype differences between control and lung cancer groups (A1/A1 OR=0.83, 95% CI 0.36-1.27 and A2/A2 OR=1.90, 95% CI 0.82-4.42) were insignificant. The patients with A1/A1 or A2/A2 genotype were sensitive to radiotherapy and the patients with A1/A2 were not sensitive to radiother-apy (Chi-square=9.2, P < 0.05), the effective rate to radiotherapy were 71.4%, 70.0% and 25.0% respectively.

CONCLUSIONSThere is no significant relationship between the BstUI SNP in p53 and susceptibility in NSCLC .The p53 BstU I SNP is closely associated with radiosensitivity of NSCLC in the Northern Chinese population.