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Objective: To construct a recombinant lentiviral expression vector pCDH-Daxx-EGFP to investigate the effect of Daxx on the proliferation of vascular smooth muscle cells (VSMCs)., Methods: The recombinant lentiviral expression vector pCDHDaxx-EGFP was constructed using PCR-based accurate synthesis method. After identification by sequencing and enzyme digestion, the recombinant lentiviral vector was contransfected into 293T cells with lentivirus packaging vector. The recombinant lentivirus particles were collected and purified to infect VSMCs, whose expression of Daxx was detected with Western boltting. The cells infected with the empty vector pCDH-EGFP or pCDH-Daxx-EGFP were incubated in serum-free medium or in the presence of angiotensin Ⅱ (AngⅡ). The cell viability was determined with MTT assay, and the cell cycle changes were analyzed with flow cytometry. The cell migration ability was assessed using a scratch wound healing assay. The expression of p-Akt protein in the cells was detected using Western blotting., Results: Double enzyme digestion and sequencing confirmed successful construction of the recombinant plasmid. Compared with the cells infected with the empty vector, the cells infected with pCDH-Daxx-EGFP exhibited significantly increased expressions of Daxx protein ( P < 0.05). AngⅡ treatment of the cells infected with the pCDH-Daxx-EGFP, as compared with the cells infected with the empty vector, significantly lowered the cell viability, S phase cell ratio and cell migration ability ( P < 0.05), and significantly decreased the expression level of p-Akt protein ( P < 0.05)., Conclusions: We successfully constructed the recombinant lentiviral vector pCDH-Daxx-EGFP and overexpressed Daxx in primary cultured VSMCs using this vector. Daxx overexpression can inhibit AngⅡ-induced proliferation and migration in VSMCs probably by regulating p-Akt protein.

Objective: To report on a case of 10p15.3 microdeletion syndrome and to explore its clinical and molecular characteristics., Methods: The patient was subjected to whole exome sequencing (WES), with his clinical features discussed in the light of literature review., Results: The patient presented with global developmental delay, hypotonia, autistic-like traits, mild facial dysmorphism and other features including short stature, small hands and feet, congenital heart disease and feeding difficulty. WES has detected deletions of ZMYND11, DIP2C, LARP4B, TUBB8, GTPBP4, IDI2, IDI1, WOR37 and ADARB2 genes on the short arm of chromosome 10. Among these, ZMYND11 gene been previously associated with intellectual disability., Conclusion: The patient's phenotype was closely correlated with that of 10p15.3 microdeletion syndrome. Haploinsufficiency of the ZMYND11 gene may underlie the manifestations of 10p15.3 microdeletion syndrome.

Recent research in epigenetics suggests that defects in epigenetic regulation of ribosomal DNA (rDNA) transcription may contribute to tumorigenesis. ATRX/DAXX complex is involved in the establishment and maintenance of the silence of the rDNA gene through H3K9me3 modification at histone variant H3.3. The ATRX/DAXX-related genes are frequently mutated in some types of tumors, which may increase rDNA transcription and promote cancer development and progression. In this review, we focus on the mechanism that abnormal transcription of rDNA potentially influences tumorigenesis. We also summarize the epigenetic regulatory mechanism of rDNA transcription, which may provide new theoretical support for drug development based on rDNA transcriptional regulation.

Objective: To investigate the interaction between nuclear transcriptional factor E26 transformation specific 1 (Ets1) and peroxiredoxin 1 (Prx1) in nicotine-induced oral precancerous lesion cells. Methods: Human oral precancerous lesion dysplastic oral keratinocyte (DOK) cells were cultured and divided into nicotine group, control group, knockdown group and knockdown control group. The nicotine group, knockdown group and knockdown control group were treated with 1 μmol/L nicotine for 7 days while the control group was untreated. Western blotting, co-immunoprecipitation (Co-IP) and chromatin immunoprecipitation (ChIP) were performed to detect Prx1 and Ets1 protein expression, Prx1 and Ets1 protein interaction, combined activity of Ets1 with PRDX1 gene promoter region in nicotine group and control group DOK cells. In nicotine group, DOK cells were transfected with siRNA or lentivirus to knockdown Ets1 and Prx1 expression. Prx1 and Ets1 protein expression was examined by Western blotting. Results: Nicotine increased the expression of Prx1 and Ets1 protein in DOK cells. The relative expression of Prx1 and Ets1 was 0.71±0.02, 0.12±0.01 in nicotine group and 0.53±0.06, 0.01±0.01 in control group ( P= 0.009, P= 0.000). Co-IP showed that Prx1 could form protein complex with Ets1. The expression of Prx1 and Ets1 complex protein was increased in nicotine group. ChIP revealed that nicotine upregulated the combination of transcriptional factor Ets1 with PRDX1 gene promoter region, and the enrichment fold was 80.9±19.7 in nicotine group and 13.8±1.2 in control group ( P= 0.004). Ets1 and Prx1 protein expression was knocked down. The relative expression of Ets1 and Prx1 was 0.60±0.06, 0.48±0.03 in knockdown group and 0.83±0.08, 0.80±0.06 in knockdown control group ( P= 0.016, P= 0.002). Ets1 knockdown suppressed the expression of Prx1 ( P= 0.002). Conversely, Prx1 knockdown also inhibited the expression of Ets1 significantly ( P= 0.000). Conclusions: In oral precancerous lesion cells, Ets1 directly regulates Prx1 expression and nicotine might promote the development of oral precancerous lesion by magnifying the positive feedback signal pathway between Ets1 and Prx1.

Objective: To investigate the role of JAZF1 gene rearrangement in the diagnosis and differential diagnosis of endometrial stromal sarcomas by fluorescence in situ hybridization (FISH). Methods: JAZF1 gene rearrangement was analyzed by FISH in 129 cases of ESS diagnosed from January 2008 to December 2016 including 105 cases of low-grade endometrial stromal sarcoma (LG-ESS), 21 cases of high-grade endometrial stromal sarcoma (HG-ESS) and 3 cases of undifferentiated uterine sarcoma (UUS). Sixteen cases of the related tumours in uterus were also collected as control group. The results were compared with our previous studies of JAZF1/JJAZ1 fusion gene in ESS by RT-PCR. Results: Detection of JAZF1 gene rearrangement by FISH was successfully analyzed in 144 cases. JAZF1 gene alteration was detected in 63 cases, all of which were LG-ESS, with an overall positivity of 60.6% (63/104), while no JAZF1 gene rearrangement was found in all other cases. JAZF1 gene rearrangement was present in LG-ESS with classic histology (69.3%, 52/75), smooth muscle differentiation (2/10), sex cord-like differentiation (4/5), fibromyxoid change (1/5), clear cell change (0/1), skeletal muscle differentiation (0/1), and schwannoma-like palisading pattern (0/1). The different components in all the cases of LG-ESS with variant histology had the clonal origin, with or without JAZF1 gene alteration. Compared to the results of JAZF1/JJAZ1 fusion gene by RT-PCR, the positive rate of JAZF1 gene rearrangement in LG-ESS by FISH (61.9%, 26/42) was significantly higher than that of RT-PCR (30.0%, 12/40; P <0.01). Conclusions: JAZF1 gene rearrangement is present only in LG-ESS, but not in HG-ESS, UUS or other related tumours in uterus. The frequency of JAZF1 gene rearrangement varies between classic LG-ESS and different morphologic variants. It is frequently, but not consistently, present in classic LG-ESS and less often positive in variant cases. The results of JAZF1 gene alterations in LG-ESS with different morphologic variants support the contention that the endometrial stromal and their variant morphologic components have the same clonal origin. Detection of JAZF1 gene rearrangement by FISH is very useful for the diagnosis and differential diagnosis of ESS.

Objective: To investigate the expression characteristic of the Daxx gene in the mouse testis and its role in spermatogenesis., Methods: Real-time PCR, Western blot and immunofluorescence were used in examining the expression characteristics of DAXX in the testis tissue from wild-type, Sertoli cell-specific androgen receptor knockout (SCARKO) and androgen receptor knockout (ARKO) mice at different postnatal weeks ., Results: The Daxx gene was highly expressed in the testis tissue and mainly in the nuclei of the wild-type mice at 4 postnatal weeks. Compared with the wild-type, the ARKO mice showed a markedly decreased expression of DAXX (0.299±0.026), which displayed a polar distribution in the spermatogenic cells (0.853±0.058) and exhibited no significant difference in the SCARKO mice (1.000±0.015)., Conclusions: The Daxx gene expression is the highest in the middle-stage development of the mouse testis, significantly decreased in ARKO mice as compared with the wild-type, and its location influenced by specific AR knockout in Sertoli cells. DAXX may be involved in the regulation of spermatogenesis in mice.

Objective: To investigate the effects of juxtaposed with another zinc finger gene 1 (JAZF1) over-expression on the levels of pro-inflammatory cytokines in high-fat diet (HFD)-induced mouse fatty liver and its associated mechanisms. Methods: Twenty male C57BL/6J (3 weeks old) and 10 male JAZF1 transgenic (JAZF1-Tg) mice were randomly divided into three groups: wide-type with normal diet (NF group, n = 10), wide-type with high-fat diet (HF group, n = 10), and JAZF1-Tg with high-fat diet (HJ group, n = 10). All mice were fed with the corresponding diet for 12 weeks, and their food consumption and body weight were measured periodically. After 12 weeks, fasting blood glucose (FBG), insulin (INS), total cholesterol (TC), triglyceride (TG), free fatty acids (FFA), and alanine aminotransferase (ALT) in the blood and liver tissue from each group were measured. TG concentration in liver tissue was determined using an enzymatic assay, and the mRNA expression of tumor necrosis factor-α(TNF-α), monocyte chemoattractant protein-1 (MCP-1), and interleukin-8 (IL-8) in the liver was measured by RT-PCR. In addition, the expression of p-JNK/JNK, p-p-38 MAPK/p-38 MAPK, p-ERK/ERK, IκBα, andβ-actin (reference) in the liver was determined using Western blot.. Results: (1) Body weight, FBG, INS, TC, and ALT were significantly reduced in the HJ group compared with those of the HF group (31.19±0.81 vs 36.07±1.43, 6.94±0.32 vs 8.14±0.36, 31.09±2.12 vs 45.21±3.34, 3.05±0.07 vs 3.81±0.08, 54.75±4.92 vs 68.09±5.15, respectively; P < 0.05). There were no significant differences in TG and FFA between the HJ and HF groups (0.72±0.05 vs 0.81±0.03, 0.81±0.4 vs 0.87±0.03; both P > 0.05). (2) There was no significant difference in liver TG concentration between the HJ and HF groups (35.49±3.17 vs 38.26±3.59, P > 0.05). (3) Compared with the HF group, the HJ group had significantly reduced mRNA expression of TNF-α, MCP-1, and IL-8 (2.54TNF-αvs 8.64±0.73, 1.19±0.73,vs 3.93±0.18, 5.09±0.48 vs 9.09±0.89; all P < 0.01), significantly reduced protein expression of p-JNK and p-p-38 MAPK (0.92±0.06 vs 1.51±0.01, 1.07±0.04 vs 1.45±0.04; both P < 0.01), and significantly increased protein expression of IκBα(0.99±0.06 vs 0.79±0.05, P < 0.01) in liver tissue. However, no significant difference was observed in the p-ERK level between the HJ and HF groups ( P > 0.05). Conclusion: Upregulation of JAZF1 expression can significantly inhibit the expression of TNF-α, MCP-1, and IL-8 in the liver of mice on HFD. This attenuation may be closely associated with the reduced activation of the JNK, p-38 MAPK, and NF-κB pathways.

Objective: To study the morphologic features, immunophenotype and significance of expression of JAZF1-SUZ12 and YWHAE-FAM22 fusion genes in endometrial stromal sarcoma (ESS)., Methods: Fifty-three cases of ESS were retrieved and the pathologic features were reviewed. Immunohistochemical study for estrogen receptor, progesterone receptor, CD10, cyclin D1, smooth muscle actin, desmin and H-caldesmon were carried out using tissue microarray technology. Reverse transcription-polymerase chain reaction (RT-PCR) was applied for detection of expression of JAZF1-SUZ12 and YWHAE-FAM22 fusion genes in 47 cases of ESS and 12 cases of other spindle cell neoplasia in uterus (including 2 cases of undifferentiated sarcoma, 3 cases of leiomyosarcoma, 3 cases of leiomyoma, 4 cases of adenosarcoma and 2 cases of uterine tumor resembling ovarian sex cord tumor)., Results: The 53 cases of ESS studied included 43 cases of low-grade ESS and 10 cases of high-grade ESS. As for low-grade ESS, in addition to the classic morphologic features, smooth muscle differentiation was present in 7 cases (16.3%), sex cord-like differentiation in 2 cases (4.7%), rhabdoid differentiation in 1 case (2.3%), clear cell changes in 1 case (2.3%) and schwannoma-like palisading pattern in 1 case (2.3%). As for high-grade ESS, sex cord-like differentiation (1 case), mucinous microcystic changes (1 case) and focal clear cell changes (1 case) were also observed. The expression rate of estrogen receptor, progesterone receptor, CD10, cyclin D1, smooth muscle actin, desmin and H-caldesmon was 86.0%, 81.4%, 74.4%, 2.3%, 23.3%, 23.3% and 4.7% in low-grade ESS, respectively, and was 1/10, 6/10, 6/10, 7/10, 1/10, 1/10 and 0 in high-grade ESS, respectively. RNA extraction was successful in 47 cases of ESS, including 39 cases of low-grade ESS and 8 cases of high-grade ESS. The positive rate of JAZF1-SUZ12 fusion gene was 30.8% (12/39) in low-grade ESS. The positive rate of YWHAE-FAM22 fusion gene was 12.5% (1/8) in high-grade ESS. The 14 control cases were all negative for JAZF1-SUZ12 and YWHAE-FAM22 fusion genes., Conclusions: As uncommon pathologic pattern may occur in both low-grade ESS and high-grade ESS, detection of JAZF1-SUZ1 and YWHAE-FAM22 fusion genes by RT-PCR would be helpful in diagnosis and differential diagnosis of ESS, especially for those tumors which lack typical morphologic features.

Objective: To investigate the effects of JAZF1 overexpression on the pro-inflammatory cytokines in hepatic steatosis., Methods: The model of hepatic steatosis was established by incubating hepatocytes with palmitic acid (PA) at 0, 0.125, 0.25, 0.5 and 1 mM dose and for 0, 6, 12, 24 and 48 hours, after which recombinant adenovirus expressing JAZF1 (Ad-JAZF1) was introduced to up-regulate expression. Triglyceride level was measured by GOD. Cell viability was detected by CCK-8. The mRNA and protein expression of TNF-alpha, MCP-1, IL-8 and JAZF1 was examined by RT-PCR, ELISA, and western blotting., Results: The PA-treated hepatocytes showed dose-dependent significant increases in TNF-alpha, MCP-1 and IL-8 mRNA expression for doses up to 0.25 mM; there were no significant increases for the highest doses of 0.5 and 1 mM. The 0.25 mM PA-treated hepatocytes showed time-dependent significant increases in TNF-alpha, MCP-1 and IL-8 mRNA expressions (FTNF-alpha = 26.51, FMCP-1 = 57.20, FIL-8 = 353.85, P less than 0.01), with the maximum level reached at 12 h and followed by a gradual decrease with longer treatment times. JAZF1 mRNA and protein expression was markedly increased in hepatocytes infected with Ad-JAZF1 (P less than 0.01). However, the AP-treated hepatocytes with JAZF1 overexpression showed down-regulation of TNF-alpha, MCP-1 and IL-8 mRNA expression (decreased by 89.69%, 77.68%, and 83.21%, respectively) and secretion (37%, 37% and 41%, respectively, P less than 0.01)., Conclusion: Stimulation of hepatocytes by the PA fatty acid in vitro promotes mRNA expression of TNF-alpha, MCP-1 and IL-8, but overexpression of JAZF1 inhibits the PA-induced expression and secretion of these factors.

Objective: To investigate the expression of death domain associated protein (Daxx) and human papillomavirus 16 E6 protein (HPV16 E6) in cervical lesions and analyze their significance., Methods: Immunohistochemical SABC method was used to detect the expression of Daxx and HPV16 E6 in 194 cases of cervical tissues with different lesions., Results: (1) The positive expression rate of Daxx was 28.57% (12/42), 40.00% (18/45), 65.91% (29/44), 66.67% (42/63), respectively, in chronic cervicitis, cervical intraepithelial neoplasia I-II (CIN I-II), CIN III and cervical squamous cell carcinoma. The positive expression rate of HPV16 E6 was 15.38% (6/39), 36.17% (17/47), 46.30% (25/54), 100.00% (24/24) in the above four groups. The positive rates in cervical cancer group and high grade CIN group were significantly higher than these in low level of CIN group and chronic cervicitis group (P<0.05). (2) The expression of Daxx was stronger in HPV16 E6 high positive group than that in HPV16 E6 low positive group (P<0.05). (3) There was no significant relationship between the expression of Daxx and pathological classification, clinical stage, situation of lymph node metastasis or patients' age in cervical squamous cell carcinoma (P>0.05). (4) There was significant positive relationship between Daxx and HPV16 E6 expression (r=0.695, P<0.05)., Conclusions: The expression of Daxx and HPV16 E6 gradually increases with cervical lesion degree aggravating. Here might be synergy between them, and both could promote the development of cervical lesions.

Death domain-associated protein (DAXX) as a multifunctional nuclear protein widely resides in nucleolus, nucleoplasm, chromatin, promyelocytic leukaemia nuclear bodies (PML-NBs) and cytoplasm. It plays significant roles in transcriptional regulation, apoptosis, cell cycle and other biological activities. Small ubiquitin-like modifier (SUMO) is required for SUMOylation which is a highly conserved post-translational modification in a wide variety of cellular processes. Numerous studies demonstrated that SUMOylation has a great effect on the subcellular localization and functional regulation of DAXX. This review will provide a summary for SUMOylation of DAXX.

Background: TLE1 is an important protein in regulating Wnt, Notch and EGFR signaling pathways. The TLE1 N-terminal Q domain regulates the pathways by mediating its oligomerization and interaction with LEF1. The aim of this study is to construct the human TLE1 N-terminal Q domain fragment in prokaryotic expression system, express and purify protein TLE1 N-terminal Q domain and prepare its polyclonal antibody., Methods: The sequence of TLE1 N-terminal Q domain obtained by PCR from human lung adenocarcinoma cDNA, was cloned into the prokaryotic expression vector pGEX-4T-1 containing Glutathione S-transferase (GST). Vector pGEX-4T1-TLE1-Q was transformed into E.coli BL21 condon plus. The GST-TLE1-Q(1-136) fusion protein was induced by IPTG, digested by Thrombin, purified with glutathione-sepharose beads and FPLC, identified by SDS-PAGE. Then rabbits were immunized with the purified protein TLE1-Q(1-136) for obtaining the antiserum. The titers and specificity of antibodies were measured by ELISA and Western blot., Results: The PCR identification and the sequencing of recombinant plasmid demonstrated that vector pGEX-4T1-TLE1-Q was successfully constructed. The SDS-PAGE shows target protein (14,000 Da) is the interest protein TLE1-Q(1-136). The TLE1 N-terminal Q domain fragment TLE1-Q(1-136) and its polyclonal antibody have been acquired, with an antibody titer of 1:20,000., Conclusions: Expression vector pGEX-4T1-TLE1-Q is correctly constructed. The TLE1 N-terminal Q domain fragment TLE1-Q(1-136) and its polyclonal antibody have been acquired. These work established the foundation for further biological study between TLE1 and lung cancers.

As a highly conserved nuclear protein, death domain-associated protein(Daxx) plays an important role in transcriptional control, carcinogenesis, resistance to virus infection, and so on. Daxx can be localized in promyleocytic leukaemia protein oncogenic domains, nucleoplasm, nucleolus, cytoplasm, and heterochromatin. The subcellular localization of Daxx can be changed by modification or interacting with other proteins. Under cellular stress, Daxx can interact with many kinds of molecules, and thus affect its downstream signaling pathway. The purpose of this review was to discuss Daxx's subcellular localization in different conditions and its translocation between subcellular compartments.

Objective: To investigate Daxx expression and its clinical significance in children with acute leukemia., Methods: The expression of Daxx protein was detected by immunohistochemical assay in 50 children with newly diagnosed acute leukemia (34 cases of acute lymphocytic leukemia and 16 cases of acute non-lymphocytic leukemia). Twenty children with normal bone marrow were used as the control group., Results: Daxx protein was expressed in 38.0% of 50 children with acute leukemia, which was significantly higher than that of the control group (5.0%) (P < 0.05). The children with acute non-lymphocytic leukemia had significantly higher Daxx expression levels (62.5%) than those with acute lymphocytic leukemia (26.5%; P < 0.05) as well as the control group (P < 0.05). There were no significant differences in the Daxx expression between acute lymphocytic leukemia children and the control group. Daxx protein was expressed in 55.6% of high risk group of acute lymphocytic leukemia but it was not expressed in standard risk group of acute lymphocytic leukemia (P < 0.05)., Conclusions: Daxx expression is abnormal in children with acute leukemia and associated with some clinical features of acute leukemia, suggesting that it may play an important role in the genesis and development of acute leukemia.

The study was aimed to investigate the action of Panax Notoginosides (PNS, extracted from notoginseng herb) on the expression of the apoptosis-related proteins (Daxx, Fas) and transcription factors (NFkB, c-Rel) in the hematopoietic cells and to explore the mechanisms of supporting cells to survive. The colony formation of CFU-GM and CFU-E in human bone marrow was assayed in the presence of various concentrations of PNS. The viability of cells was assayed by trypan blue and the changes of cell morphology were observed with microscope. The Annexin-V positive cells were detected by FCM. Three lineages of human myeloid HL-60, erythroid K562, megakaryoid CHRF-288 and Meg-01 cells were incubated in addition of PNS (10 mg/L) for 14 days. The nuclear or cytoplasm protein of cells was extracted and analyzed by Western blot with monoclonal antibodies against Daxx, Fas or NFkB, c-Rel. The results showed: (1) the proliferation on hematopoietic progenitor cells (CFU-GM and CFU-E) and four cell lines was promoted by PNS; (2) after the four cell lines were promoted by PNS and hungered through wiping off the sera, the viability of the four cell lines was high without significant morphological change and neither the detection of Annexin-V positive cells; (3) the expression of Daxx and Fas protein could be inhibited by PNS. Western Blot showed that Daxx in four cell lines treated by PNS were 33.3-61.5% lower than that in untreated controls. The Fas protein was also descended in three cell lines of K562, CHRF-288 and Meg-01 by 33.3-71.4% respectively, while Fas protein in HL-60 cells was no detectable difference after PNS treatment. (4) The transcription factors NFkB and c-Rel protein could be increased by PNS. The NFkB, c-Rel protein were also enhanced in three cell lines of K562, CHRF-288 and Meg-01 by (2.0-2.7) and (1.5-2.3)-fold respectively, while there were also no detectable difference in HL-60 cells after PNS treatment. It is concluded that PNS inhibites the expression of Daxx and Fas proteins, may decrease the apoptosis of the hematopoietic cells. The level of NFkB and c-Rel proteins can be enhanced by PNS, which not only stimulates the proliferation of cells, but also inhibits the activity of the waterfall of caspase and apoptosis of the hematopoietic cells. PNS may treat the disease with over-apoptosis of hematopoietic cells, as aplastic anemia.