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Evidence suggests that abnormal DNA methylation patterns play a crucial role in the etiology and pathogenesis of colon adenocarcinoma (COAD). In this study, we identified a total of 97 methylation-driven genes (MDGs) through a comprehensive analysis of the Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Univariate Cox regression analysis identified four MDGs (CBLN2, RBM47, SLCO4C1, and TMEM220) associated with overall survival (OS) in COAD patients. A risk prediction model was then developed based on these four MDGs to predict the prognosis of COAD patients. We also created a nomogram that incorporated risk scores, age, and TNM stage to promote a personalized prediction of OS in COAD patients. Compared with the traditional TNM staging system, our new nomogram was better at predicting the OS of COAD patients. In cell experiments, we confirmed that the mRNA expression levels of CLBN2 and TMEM220 were regulated by the methylation of their promoter regions. Moreover, immunohistochemistry showed that CBLN2 and TMEM220 were potential prognostic biomarkers for COAD patients. In summary, we have established a risk prediction model and nomogram that might be effectively utilized to promote the prediction of OS in COAD patients.

Effective therapeutic system to periodontitis was designed using cross-linked cyclodextrin metal-organic framework (COF) as carrier for iodine and further suspended in hydroxyethyl cellulose gel as I2@COF-HEC hydrogel. Inclusion of iodine within the COF was demonstrated by SR-FTIR spectral and characteristic DSC and TGA changes. Molecular modelling identified the interaction of iodine with both COF central cavity and individual cyclodextrin moieties of COF. In vitro results of study demonstrated that iodine release in artificial saliva from I2@COF-HEC hydrogel could be extended up to 5 days, which was slower than I2@COF particles. Using an in vivo rat model of periodontitis, micro-computed tomography of alveolar bone morphology demonstrated that I2@COF-HEC hydrogel showed similar effects in decreasing periodontal pocket depth and alveolar bone resorption to minocycline ointment, a periodontitis antibiotic. The I2@COF-HEC hydrogel is a novel local delivery device of iodine as a broad spectrum antimicrobial use for treatment of periodontitis.

We had previously demonstrated that the calcitonin gene‐related peptide (CGRP) suppresses the oxidative stress and vascular smooth muscle cell (VSMC) proliferation induced by vascular injury. A recent study also indicated that CGRP protects against the onset and development of angiotensin II (Ang II)‐induced hypertension, vascular hypertrophy and oxidative stress. However, the mechanism behind the effects of CGRP on Ang II‐induced oxidative stress is unclear. CGRP significantly suppressed the level of reactive oxygen species (ROS) generated by NADPH oxidase in Ang II‐induced VSMCs. The Ang II‐stimulated activation of both Src and the downstream transcription factor, STAT3, was abrogated by CGRP. However, the antioxidative effect of CGRP was lost following the expression of constitutively activated Src or STAT3. Pre‐treatment with H‐89 or CGRP8–37 also blocked the CGRP inhibitory effects against Ang II‐induced oxidative stress. Additionally, both in vitro and in vivo analyses show that CGRP treatment inhibited Ang II‐induced VSMC proliferation and hypertrophy, accompanied by a reduction in ROS generation. Collectively, these results demonstrate that CGRP exhibits its antioxidative effect by blocking the Src/STAT3 signalling pathway that is associated with Ang II‐induced VSMC hypertrophy and hyperplasia.

Progression of acute pancreatitis (AP) into a severe form usually results in a life-threatening condition with multiple organ dysfunction, and in particular acute lung injury (ALI), often contributes to the majority of AP-associated deaths. Increasing evidence has shown that uncontrolled activation of the immune system with rapid production of inflammatory cytokines play a dominant role in this process. As an intracellular inflammatory signaling platform, the NOD-like receptor protein 3 (NLRP3) inflammasome, is recently reported to be involved in the pathogenesis of AP progression, however, the relationship between NLRP3 inflammasome activation and AP-associated lung injury remains unclear yet. Here, we show that NLRP3 inflammasome activation and subsequent pyroptosis in alveolar macrophages (AMs) is responsible for the lung injury secondary to AP. In addition, plasma-derived exosomes from AP mice is capable of triggering NLRP3-dependent pyroptosis in AMs. Inhibition of exosome release or uptake in vivo by inhibitors substantially suppresses AMs pyroptosis and thereby alleviates AP-induced pulmonary lesion. Collectively, the current work reveals for the first time the involvement of NLRP3-dependent pyroptosis induced by plasma exosomes in the pathogenesis of AP-induced ALI, suggesting that the exosome-mediated NLRP3 inflammatory pathway is a potential therapeutic target for the treatment of lung injury during AP.

The study aims to investigate the changes and regulation of androgen receptor and insulin-like growth factor-1 in the PC3 prostate cells treated with 5α-dihydrotestosterone, estrone, and flutamide. The PC3 cells were cultured and treated with 5α-dihydrotestosterone, estrone, and flutamide. Immunocytochemistry and Western blot were used to detect the expression of androgen receptor and insulin-like growth factor-1. The androgen receptor expression was analyzed by Western blot and optic density scan in the presence or absence of various kinase inhibitors. The statistical calculations were performed with the statistics-analyzing software package SPSS 13.0. A P

Recent studies suggest that endoplasmic reticulum stress (ERS) is the key process in ischemic brain injury. The JNK pathway is also involved in the process of ischemic brain injury.We established a middle cerebral artery occlusion/reperfusion (MCAO/R) model in rats; detected the changes in c-Jun N-terminal kinase (JNK), GADD153 and caspase-12 at different reperfusion time points by immunohistochemistry, Western blot and double-label immunofluorescence; and observed the effect of JNK inhibitor SP600125 on the expression of JNK, GADD153 and caspase-12 to explore the relationship between the endoplasmic reticulum road and JNK pathway.The expression of the two hallmarks of ERS-GADD153 and caspase-12-significantly increased, and the activation of JNK also obviously increased. After interference by SP600125, the expression of p-JNk and caspase-12 obviously decreased, whereas the decrease of GADD153 occurred only after 24 h reperfusion.Both ERS and JNK pathways are involved in the pathological process of ischemic brain injury. The JNK pathway may be involved in the process of ERS, but perhaps has more effect on the caspase-12 pathway.

The current prospective randomized study was designed to evaluate the safety and efficacy of combined intrapleural cisplatin and OK-432 (picibanil) plus hyperthermotherapy in patients with malignant pleural effusion (MPE).A total of 358 patients with MPE due to end-stage malignancies were enrolled and randomly divided into two groups, A and B: the intrapleural combination of cisplatin and OK-432 with hyperthermotherapy (n = 179) or without hyperthermotherapy (n = 179), respectively. Mild toxicities such as nausea, vomiting or anorexia, bone marrow depression, and pyrexia were similar in both groups.Patients in Group A (with hyperthermotherapy) showed a significantly higher overall response (93.4%) compared to those in Group B (79.8%, χ(2) = 43.11, p.05). The median survival time for patients in Group A and Group B were 8.9 and 6.2 months, respectively (p.05). After treatment, the quality of life scores were significantly increased in both groups as compared to prior treatment (p.05).In conclusion, our study suggests that combined intrapleural cisplatin and OK-432 followed by hyperthermotherapy are more effective in the control of MPE and improve patients' quality of life.

Knowledge of allele and genotype frequencies is an essential prerequisite to the use of any human polymorphism in forensic work.To study the genetic polymorphism and evaluate the application value of nine STR loci.Genotyping of nine STR loci, including D11S2368, D12S391, D13S325, D18S1364, D22-GATA198B05, D6S1043, D2S1772, D7S3048 and D8S1132, of 1050 unrelated individuals was performed with the STR_Typer_10_v1 kit and Genetic Analyzer 3100 and analyzed with PowerState V12.xls and Arlequin ver 3.11 analyzing software.Allele frequency distribution was statistically analyzed and Hardy-Weinberg equilibrium determined. Several common parameters used in forensic sciences were found: the heterozygosity (H) ranged from 0.827 to 0.892; the matching probability (MP) ranged from 0.029 to 0.074; the power of discrimination (PD) ranged from 0.926 to 0.971; the power of exclusion (PE) ranged from 0.649 to 0.779; the polymorphic information content (PIC) ranged from 0.77 to 0.86; and the typical paternity index (TPI) ranged from 2.88 to 4.62.The results indicate that nine STR loci are high polymorphic among the Han population in Southern China. This set of polymorphic STR loci is a useful tool in forensic paternity testing and anthropological study.

To explore the mutation of Y-STR loci in meiotic allelic transmission in a large pedigree.The oral swabs of 163 male individuals were collected from a Lin pedigree. Twenty-two Y-STR genetic markers were typed with AGCU Y24 fluorescent detection kit (AGCU Y24 system), which also contained 16 Y-STR markers included in Yfiler multiple amplification kit (Yfiler system). The genotyping results of Y-STR loci were compared between each two males in the pedigree.There were 20 and 30 kinds of haplotypes obtained with Yfiler and AGCU Y24 systems in 163 male individuals from the Lin pedigree, respectively. The rates referred to haplotype differences (RRHD) of these two typing systems between male pairs were 0.910 5 and 0.922 7, respectively. The average number of marker differences were 6.582 1 and 9.824 8, respectively. The RRHD increased along with the incidents of meiosis.Y-STR mutation leads to different Y-STR haplotypes among the male members in a paternal pedigree and the rate of difference increases along with the incidents of meiosis.

To investigate genetic diversity in Chinese populations, 706 unrelated male individuals from five ethnic groups (Han, Korean, Hui, Mongolian, and Tibetan, respectively) were analyzed with 17 Y-chromosomal STRs. The haplotype diversity was 0.99985 in the combined data. A total of 675 distinct haplotypes were observed, of which 649 were unique. Y-chromosome haplogroups in the five groups were also predicted with Y-STR haplotypes. Genetic distance between the five studied ethnic groups and other published groups was analyzed by analysis of molecular variance and visualized in a multidimensional scaling plot. In conclusion, the 17 Y-STR loci are highly polymorphic markers in the five groups and hence are very useful in forensic application, population genetics, and human evolution studies.

To identify potential mutation in a Chinese family featuring X-linked alpha thalassemia/mental retardation syndrome (ATR-X).Based on clinical symptoms and inheritance pattern, linkage analysis of X chromosome short tandem repeats (X-STR) loci was carried out to locate the candidate gene. Subsequently, sequences of exons and exon-intron boundaries of the candidate gene were amplified with polymerase chain reaction (PCR). Potential mutations were detected by direct DNA sequencing. All patients were also analyzed for the trait of thalassemia.Linkage analysis indicated the candidate gene to be ATRX. Subsequently, a homozygous missense mutation c.736CT (p.R246C) was found in exon 9 of ATRX in all of the 3 patients. And a heterozygous mutation c.736CT (p.R246C) was also identified in the patient's mother and grandmother. Similar mutations were not detected in other members of the family. Alpha thalassemia was detected in the proband and another patient, whose genotypes were determined as -α(3.7)/αα and --(sea)/αα, respectively.Missense mutation of c.736CT in ATRX gene is a mutation hotspot, and p.R246C may disturb the function of ATRX-DNMT3-DNMT3L domain (ADD), which may be responsible for the disease in this family.

Y chromosome is a male-specific paternal inherited chromosome. The STR markers on Y chromosome have been widely used in forensic practices. This article summarizes the characteristics of Y-STR and some factors are considered of selecting appropriate Y-STR markers for Chinese population. The prospects of existing and potential forensic applications of Y-STR profiles are discussed including familial excluding, familial searching, crowd source deducing, mixture sample testing, and kinship identifying. The research, development, verification of Y-STR kit, Y-STR mutation rate, and search software are explored and some suggestions are given.

To deduce the calculation formulae of likelihood ratio for kinship testing with X-short tandem repeat (X-STR) typing.With the identity by decent coefficient of different relationships, the joint genotypic probability of two individuals with specific genotypes was calculated as X value, and then the joint genotypic probability of two unrelated individuals was calculated as Y value; therefore, the likelihood ratio value (X/Y) was obtained.The calculation formulae of the likelihood ratio for different genotype combinations of the female-female, female-male and male-male genetic relationships were derived and verified by real cases.The calculation formulae are simple and accurate to evaluate the likelihood ratio for two individuals' genetic relationship with X-STR typing. The formulae provide the basic potential value for the difficult kinship testing with X-STR loci.

Excessive K(+) efflux promotes central neuronal apoptosis; however, the type of potassium channel that mediates K(+) efflux in response to different apoptosis-inducing stimuli is still unknown. It is hypothesized that the activation of large-conductance Ca(2+)-activated K(+) channels (BKCa) mediates hypoxia/reoxygenation (H/R)- and ischemia/reperfusion (I/R)-induced neuronal apoptosis. Rat hippocampal neuronal cultures underwent apoptosis after reoxygenation, as assessed by morphologic observation, terminal deoxynucleotidyl transferase dUTP nick end labeling staining, and caspase-3 activation. Single-channel recordings revealed upregulation of BKCa channel activity 6 h after reoxygenation, which might be caused by elevated cytosolic Ca(2+). The K(+) ionophore valinomycin and the BKCa channel opener NS1619 induced neuronal apoptosis. Transfection of the BKCa channel α subunit into Chinese hamster ovary (CHO-K1) cells, which do not express endogenous K(+) channels, or into neurons will induce cell apoptosis, indicating that the opening of the BKCa channel serves as a pivotal event in mediating cell apoptosis. The specific BKCa channel blockers charybdotoxin and iberiotoxin and the nonselective K(+) channel blocker tetraethylammonium at concentrations more specific to the BKCa channel were neuroprotective. The A-type potassium channel blocker 4-aminopyridine and apamin, a small-conductance Ca(2+)-activated K(+) channel blocker, were not protective. This result suggests the involvement of the BKCa channel in H/R-induced apoptosis. Similarly, specific BKCa channel blockers also showed neuroprotection in neurons subjected to oxygen-glucose deprivation/reoxygenation or animals subjected to forebrain ischemia-reperfusion. These results demonstrate that the over-activity of BKCa channels mediates hippocampal neuronal damage induced by H/R in vitro and I/R in vivo.

To study the roles of βIII-tubulin and survivin in the development of gastric cancer, and see for any relationship between the expression levels of these two genes and docetaxel chemoresistance in advanced gastric cancer.Patients with advanced gastric cancer treated with docetaxel-based chemotherapy were enrolled and their tumor samples were collected retrospectively for analysis (study group). The control group consisted of patients with benign gastric mucosa lesions. Expression levels of βIII-tubulin and survivin were evaluated with immunohistochemistry.The expression levels of βIII-tubulin and survivin in the study group were significantly higher compared with those in the control group (p0.01). Spearman's correlation analysis suggested that the expression of βIII-tubulin was also correlated with that of survivin (p0.05), but no correlation existed between the expression levels of βIII-tubulin and survivin with age, gender and pathological tumor type. The complete response (CR) + partial response (PR) rates were 54.10%. Patients with βIII-tubulin and/or survivin overexpression were less responsive to docetaxel-based therapy (p0.05) and also had shorter median time to progression and 1- and 2-year survival rates (p0.05).Overexpression of βIII-tubulin and survivin in gastric cancer cells was associated with resistance to docetaxel-based chemotherapy in patients with advanced gastric cancer.

To explore the cause for allelic drop-out at short tandem repeat (STR) loci upon paternity testing with a PowerPlex® 16 kit.A total of 10 642 DNA confirmed paternity testing cases (18 314 parent/child allelic transfers) were analyzed with the PowerPlex® 16 kit. Samples suspected for having allelic drop-out were verified with an Identifiler™ kit and/or locus-specific singleplex amplification systems. PCR products of null alleles were separated and directly sequenced.Eight cases of allelic drop-out were found. The overall rate of null allele in the PowerPlex® 16 system was 0.437 × 10(-3). DNA sequencing has confirmed single base variations within the binding region of published primers, in which 4 cases involved the D18S51 locus (2 cases with GA transitions at 79 bp upstream of the repeats, 1 case with GT transversion at 162 bp downstream of the repeats and 1 case with GC transversion at 74 bp upstream of the repeats), 2 cases involved the D21S11 locus (1 case with CA transversion at 17 bp upstream of the repeats and 1 case with AG transition at 12 bp upstream of the repeats). One case involved the FGA locus (1 case with GA transition at 142 bp downstream of the repeats) and 1 case involved TPOX locus (1 case with GA transition at 198 bp downstream of the repeats).Base variation in the primer binding region may cause failed PCR and result in null allele reports. Alternative primer sets should be used to verify the suspected allelic drop-out. Attention should be paid to this during paternity testing and data exchange for personal identification.

Conjoined twins are rarely observed, but heteropagus conjoined twins occur even less frequently. Heteropagus conjoined twins are asymmetrically conjoined twins, in which one twin is nearly anatomically normal (autosite), and the other twin remains anatomically incomplete (parasite) but physically attached to the autosite. Epigastric heteropagus conjoined twins (EHTs), referring to a parasitic twin who is attached to the epigastrium of the autosite, are extremely rare (1). Due to the rare incidence of EHTs, there are few publications on this subject, which has resulted in a general lack of knowledge among surgeons with respect to how to approach the management of EHTs. Furthermore, EHTs generally manifest as more medically complex situations than typical conjoined twins. Therefore, the etiology, manifestation, and therapeutic outcomes of EHTs remain controversial. In this article, we report two cases of EHTs encountered in our department between May 2007 and October 2007. One EHT case involved a large ventral hernia near the omphalocele, which has not been previously reported. Moreover, DNA analysis of case number 2 suggested a monozygotic origin of the EHTs.

To introduce the method of avuncular index (AI) calculation.Identity by decent coefficient, coancestry coefficient and AI law were employed in identification of uncle-niece relationship, when autosomal STR loci were detected to determine controversial uncle-niece relationship.The results of AI calculation were coincidental using identity by descent coefficien, coancestry coefficient and AI law.The results are coincidental using three methods in the different situations. AI index is higher with participation of children's mother.

To investigate the genetic polymorphisms of 12 X chromosome short tandem repeat (X-STR) loci of Investigator Argus X-12 amplification kit in Guangdong Han population.DNA samples from 200 unrelated individuals (100 males and 100 females) and 103 families (59 father-mother-daughter trios and 44 mother-son duos) were extracted and amplified with fluorescence labeled multiplex PCR system. PCR products were separated and genotyped with capillary array electrophoresis.One hundred and thirty-seven alleles,including 9 off ladder alleles (OL allele) were observed at the 12 X-STR loci in the population. Six mutations were observed in 162 meioses. The combined power of discrimination (DP) was 0.999 999 997 in males and 0.999 999 999 in females, and the combined mean exclusion chance (MEC) was 0.999 999 988 in the trio cases and 0.999 998 013 in the duo cases.Investigator-Argus X-12 amplification system is highly polymorphic in Guangdong Han population and it is powerful for personal identification and paternity testing.

Changes of androgen receptor (AR) and insulin-like growth factor-1 (IGF-1) were investigated in LNCaP cells treated with 5α-dihydrotestosterone (DHT), estrone and flutamide. Real-time PCR, immunocytochemistry and western blotting were used to detect the expression of AR and IGF-1 in the presence or absence of various kinase inhibitors. Low concentrations of DHT, estrone and flutamide increased the expression of AR and IGF-1, especially estrone, with concentration and time dependence. With DHT and flutamide, there was a significant alteration in AR expression (p0.001). The results indicated expression of AR and IGF-1 genes to be influenced by DHT, estrone and flutamide in LNCaP cells, regulated by multiple signal pathways.

A fluorescent multiplex PCR that simultaneously amplifies five X-chromosomal short tandem repeat (X-STR) loci (DXS6803, DXS981, DXS6809, DXS6789 and DXS7132)was set up to study their polymorphic nature and to determine its use in kinship tests for forensic cases. PCR products were analyzed using capillary electrophoresis and ABI prism 3100 Genetic Analyzer with GeneMapper ID 3.1 Analysis Software. The pentaplex system gave satisfactory results as its sensitivity, reproducibility and unambiguous genotyping. About 20 ng DNA was routinely used, although 0.25 ng DNA was sufficient for allele typing. The results demonstrate that the multiplex system of the five X-STR loci provides a fast technology for forensic identification and paternity testing. The X-STR pentaplex system can complement the analysis of AS-STR and Y-STR efficiently, especially in complex cases of kinship testing.

To develop single-cell multiplex nested polymerase chain reaction (PCR) assays for preimplantation genetic diagnosis (PGD) in couples at risk of having child with beta-thalassemia.Primers were designed and synthesized according to the documented mutation sites common among Chinese. Venous blood was collected from 4 pairs of husband and wife, all heterozygotes for beta-thalassemia, and underwent multiple nested PCR. Intraooplasmic sperm injection and mechanical bio psy was used to obtain single blastomere. Multiplex nested PCR was used to detect the CD41-42 mutation and the closely linked polymorphic marker, HumTHO1 gene or CD41-42, CD41-28, IVSII654 mutation and HumTHO1 gene in the single blastomeres from four clinical PGD cycles. The normal embryos with high scores capable of continuing to divide were transplanted into the uteri. The process of gestation was observed.200 lymphocytes were amplified by nested PCR. The average amplification rate of the most common 16 beta-thalassemia mutations in Chinese population was 91.3% and the average rate of allele drop out for different sites was 17.0% without differences between any 2 sites. During the 4 PGD cycles 33 embryos underwent bioassay with a success rate of 100%. 33 blastomeres were obtained to undergo PCR, of which 30 were successfully amplified with an amplification rate of 90.9%. Explicit diagnosis was obtained in 26 of the 30 embryos: 7 normal homozygotes, 11 heterozygotes, and 8 abnormal or complex heterozygotes. One or more embryos were transferred back into the uteri of the 4 women and clinical pregnancy occurred in one woman. Five weeks after the implantation B-mode ultrasonography showed monocyesis, and in the 17th week of gestational period paracentesis of cord blood showed normal homozygote. At last a normal female infant confirming the PGD result had been born, which was the first reported unaffected pregnancy resulting from PGD using multiplex nested PCR for couples as beta-thalassemia gene carriers. The results of diagnosis for embryo all corresponded to those for blastomere. The average ADO rate of blastomere was 13.3% (4/30).PGD using multiplex nested PCR, as an alternative to prenatal diagnosis, is a reliable and effective way to help couples-carriers of pathogenetic genes to get a healthy baby.

To understand the forensic practice of DNA profiling from minimal amount of DNA.Serial dilutions of DNA were amplified with the PowerPlex 16 System Kit, then the genotyping of short tandem repeat(STR) was performed by ABI 377 DNA automated Sequencer.When the mount of DNA template was less than 250 pg, allelic drop-out apparently occurred at several loci. Other disturbed peaks, such as artefact bands and imbalanced heterozygote, also presented.The anomalous results may result in incorrect genotyping. Careful and comprehensive considerations are needed to interpret the STR profile of minute DNA.