Dynamic readjustment of parental methylation patterns of the 5'-flank of the mouse H19 gene during in vitro organogenesis

Gametic marks are stably propagated in order to manifest parent of origin-specific expression patterns of imprinted genes in the developing conceptus. Although the character of the imprint has not yet been fully elucidated, there is compelling evidence that it involves a methylation mark. This is exemplified by a region upstream of the H19 gene, which is not only methylated in a parent of origin-specific manner, but also regulates the silencing of the maternal Igf2 and paternal H19 alleles, respectively. We show here that the parental-specific methylation patterns within the differentially methylated domain (DMD) are perturbed in the soma during in vitro organogenesis. Under these conditions, the paternal DMD allele becomes partially demethylated, whereas the maternal DMD allele gains methylation. Despite these effects, there were no changes in allelic Igf2 or H19 expression patterns in the embryo. Finally, we show that although TSA derepresses the paternal H19 allele in ectoplacental cone when in vitro developed, there is no discernible effect on the methylation status of the paternally inherited 5'-flank in comparison to control samples. Collectively, this data demonstrates that the parental mark is sensitive to a subset of environmental cues and that a certain degree of plasticity of the gametic mark is tolerated without affecting the manifestation of the imprinted state.     

Body Height,Estimated Cerebrospinal Fluid Pressure and Open-Angle Glaucoma. The Beijing Eye Study 2011

Purpose

To examine potential associations between body height, cerebrospinal fluid pressure (CSFP), trans-lamina cribrosa pressure difference (TLCPD) and prevalence of open-angle glaucoma (OAG) in a population-based setting.

Methods

The population-based Beijing Eye Study 2011 included 3468 individuals with a mean age of 64.6±9.8 years (range:50–93 years). A detailed ophthalmic examination was performed. Based on a previous study with lumbar cerebrospinal fluid pressure (CSFP) measurements, CSFP was calculated as CSFP[mmHg] = 0.44×Body Mass Index[kg/m²]+0.16×Diastolic Blood Pressure[mmHg]-0.18×Age[Years]-1.91

Results

Data of IOP and CSFP were available for 3353 (96.7%) subjects. Taller body height was associated with higher CSFP (P<0.001; standardized correlation coefficient beta:0.13; regression coefficient B:0.29; 95% confidence interval (CI):0.25,0.33) after adjusting for male gender, urban region of habitation, higher educational level, and pulse rate. If TLCPD instead of CSFP was added, taller body height was associated with lower TLCPD (P<0.001;beta:−0.10;B:−0.20;95%CI:−0.25,−0.15). Correspondingly, higher CSFP was associated with taller body height (P = 0.003;beta:0.02;B:0.01;95%CI:0.00,0.02), after adjusting for age, gender, body mass index, pulse, systolic blood pressure, and blood concentration of cholesterol. If IOP was added to the model, higher CSFP was associated with higher IOP (P<0.001;beta:0.02;B:0.02;95%CI:0.01,0.03). TLCPD was associated with lower body height (P = 0.003;beta:−0.04;B −0.02,95%CI:−0.04,−0.01) after adjusting for age, body mass index, systolic blood pressure, pulse, blood concentrations of triglycerides, axial length, central corneal thickness, corneal curvature radius, and anterior chamber depth. Adding the prevalence of OAG to the multivariate analysis revealed, that taller body height was associated with a lower OAG prevalence (P = 0.03;beta:−0.03;B:−1.20;95%CI:−2.28,−0.12) after adjusting for educational level and gender.

Conclusions

Taller body height was associated with higher CSFP and lower TLCPD (and vice versa), after adjusting for systemic and ocular parameters. Parallel to the associations between a higher prevalence of glaucoma with a lower CSFP or higher TLCPD, taller body height was associated with a lower prevalence of OAG.     

R环的形成及对基因组稳定性的影响

R-环是由一个RNA:DNA杂交体和一条单链状态的DNA分子共同组成的三链核酸结构。其中, RNA:DNA杂交体的形成起因于基因转录所合成的RNA分子不能与模板分开, 或RNA分子重新与一段双链DNA分子中的一条链杂交。在基因转录过程中, 当转录泡遇到富含G碱基的非模板链区或位于某些与人类疾病有关的三核苷酸卫星DNA时, 转录泡后方累积的负超螺旋可促进R环形成。同时, 新生RNA分子未被及时加工、成熟或未被快速转运到细胞质等因素也会催生R环。研究表明, 细胞拥有多种管理R环的方法, 可以有效地管理R环的形成和处理已经形成的R环, 以尽量避免R环对DNA复制、基因突变和同源重组产生不利影响。文章重点分析了R-环的形成机制及R环对DNA复制、基因突变和同源重组的影响, 并针对R-环诱导的DNA复制在某些三核苷酸重复扩增有关的神经肌肉退行性疾病发生过程中的作用进行了分析和讨论。     

重组猪源胰蛋白酶原的构建、表达与活性分析

目的：构建、表达重组猪源胰蛋白酶原,并对其进行分离纯化和酶活性分析。方法：利用大肠埃希菌表达系统（pET-28a,宿主菌BL21 DE3）优化天然猪源胰蛋白酶原基因,经乳糖诱导表达重组胰蛋白酶原蛋白,采用阴离子交换色谱法分离纯化,以SDS-PAGE 电泳鉴定目标蛋白,以紫外分光光度法检测重组蛋白的酶活力。结果：测序结果表明,重组猪源胰蛋白酶原基因工程菌构建成功,SDS-PAGE检测显示目的蛋白条带位置与标准猪源胰蛋白酶条带位置一致,且酶活力可达513.09 U?mg-1。结论：成功获得了重组猪源胰蛋白酶原,且酶活力较好,为进一步研究生产提供了基础。     

Seroprevalence of Merkel Cell Polyomavirus in the General Rural Population of Anyang,China

Background

Despite the probably causal link between Merkel cell polyomavirus (MCPyV) infection and Merkel cell carcinoma (MCC), a rare but aggressive skin malignancy, little is known about the seroepidemiology of MCPyV among healthy adults in China.

Methods

Serum antibodies against MCPyV were evaluated by multiplex serology in a population-based study of 5548 adults (including 1587 heterosexual couples) aged 25–65 years who were enrolled from rural Anyang, China in 2007–2009. Univariate and multivariate logistic regression analyses were performed to assess the risk factors for the seropositivity of MCPyV.

Results

The seroprevalence for MCPyV was 61.0%. MCPyV seropositivity was significantly higher in males than in females (64.5% vs. 57.7%, P<0.001), and for both genders, showed a trend of increase with age (Male: P _trend<0.001; Female: P _trend<0.001). Furthermore, among antibody positives, antibody levels of MCPyV increased with advancing age (P _trend = 0.017). MCPyV seropositivity of one spouse was significantly associated with that of the other partner (Adjusted OR = 1.32, 95% CI: 1.07–1.62). However, there was no association between sexual behaviors and the seropositivity of MCPyV.

Conclusions

High seroprevalence of MCPyV was observed in healthy Chinese individuals. Serological evidence suggests that nonsexual horizontal spread of MCPyV can occur among family members, and further research in this regard is needed.     

Prognostic Impact of Renal Dysfunction Does Not Differ According to the Clinical Profiles of Patients: Insight from the Acute Decompensated Heart Failure Syndromes (ATTEND) Registry

Background

Renal dysfunction associated with acute decompensated heart failure (ADHF) is associated with impaired outcomes. Its mechanism is attributed to renal arterial hypoperfusion or venous congestion, but its prognostic impact based on each of these clinical profiles requires elucidation.

Methods and Results

ADHF syndromes registry subjects were evaluated (N = 4,321). Logistic regression modeling calculated adjusted odds ratios (OR) for in-hospital mortality for patients with and without renal dysfunction. Renal dysfunction risk was calculated for subgroups with hypoperfusion-dominant (eg. cold extremities, a low mean blood pressure or a low proportional pulse pressure) or congestion-dominant clinical profiles (eg. peripheral edema, jugular venous distension, or elevated brain natriuretic peptide) to evaluate renal dysfunction''s prognostic impact in the context of the two underlying mechanisms. On admission, 2,150 (49.8%) patients aged 73.3±13.6 years had renal dysfunction. Compared with patients without renal dysfunction, those with renal dysfunction were older and had dominant ischemic etiology jugular venous distension, more frequent cold extremities, and higher brain natriuretic peptide levels. Renal dysfunction was associated with in-hospital mortality (OR 2.36; 95% confidence interval 1.75–3.18, p<0.001), and the prognostic impact of renal dysfunction was similar in subgroup of patients with hypoperfusion- or congestion-dominant clinical profiles (p-value for the interaction ranged from 0.104–0.924, and was always >0.05).

Conclusions

Baseline renal dysfunction was significantly associated with in-hospital mortality in ADHF patients. The prognostic impact of renal dysfunction was the same, regardless of its underlying etiologic mechanism.     

Phytantriol-Based In Situ Liquid Crystals with Long-Term Release for Intra-articular Administration

The purpose of this study was to develop an injectable in situ liquid crystal formulation for intra-articular (IA) administration, and in situ forming a viscous liquid crystalline gel with long-term release of sinomenine hydrochloride (SMH) upon water absorption. The pseudo-ternary phase diagram of phytantriol (PT)-ethanol (ET)-water was constructed, and isotropic solutions were chosen for further optimization. The physicochemical properties of isotropic solutions were evaluated, and the phase structures of liquid crystalline gels formed by isotropic solutions in excess water were confirmed by crossed polarized light microscopy (CPLM) and small-angle X-ray scattering (SAXS). In vitro drug release studies were conducted by using a dialysis membrane diffusion method. The optimal in situ cubic liquid crystal (ISV₂) (PT/ET/water, 64:16:20, w/w/w) loaded with 6 mg/g of SMH showed a suitable pH, showed to be injectable, and formed a cubic liquid crystalline gel in situ with minimum water absorption within the shortest time. The optimal ISV₂ was able to sustain the drug release for 6 days. An in situ hexagonal liquid crystal (ISH₂) system was prepared by addition of 5% vitamin E acetate (VitEA) into PT in the optimal ISV₂ system to improve the sustained release of SMH. This ISH₂ (PT/VitEA/ET/water, 60.8:3.2:16:20, w/w/w/w) was an injectable isotropic solution with a suitable pH range. The developed ISH₂ was found to be able to sustain the drug release for more than 10 days and was suitable for IA injection for the treatment of rheumatoid arthritis (RA).KEY WORDS: in situ cubic liquid crystal, in situ hexagonal liquid crystal, phytantriol, sinomenine hydrochloride, sustained drug release     

Proteinase-activated Receptor 2 Promotes Cancer Cell Migration through RNA Methylation-mediated Repression of miR-125b

Proteinase activated-receptor 2 (PAR₂) participates in cancer metastasis promoted by serine proteinases. The current study aimed to test the molecular mechanism by which PAR₂ promotes cancer cell migration. In different cancer cells, activation of PAR₂ by activating peptide (PAR₂-AP) dramatically increased cell migration, whereas knock down of PAR₂ inhibited cellular motility. The PAR₂ activation also repressed miR-125b expression while miR-125b mimic successfully blocked PAR₂-induced cell migration. Moreover, Grb associated-binding protein 2 (Gab2) was identified as a novel target gene of miR-125b and it mediated PAR₂-induced cell migration. The correlation of PAR₂ with miR-125b and Gab2 was further supported by the findings obtained from human colorectal carcinoma specimens. Remarkably, knock down of NOP2/Sun domain family, member 2 (NSun2), a RNA methyltransferase, blocked the reduction in miR-125b induced by PAR₂. Furthermore, PAR₂ activation increased the level of N⁶-methyladenosine (m⁶A)-containing pre-miR-125b in NSun2-dependent manner. Taken together, our results demonstrated that miR-125b mediates PAR₂-induced cancer cell migration by targeting Gab2 and that NSun2-dependent RNA methylation contributes to the down-regulation of miR-125b by PAR₂ signaling. These findings suggest a novel epigenetic mechanism by which microenvironment regulates cancer cell migration by altering miRNA expression.