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Study Question: Does embryo vitrification affect placental histopathology pattern and perinatal outcome in singleton live births?, Summary Answer: Embryo vitrification has a significant effect on the placental histopathology pattern and is associated with a higher prevalence of dysfunctional labor., What Is Known Already: Obstetrical and perinatal outcomes differ between live births resulting from fresh and frozen embryo transfers. The effect of embryo vitrification on the placental histopathology features associated with the development of perinatal complications remains unclear., Study Design, Size, Duration: Retrospective cohort study evaluating data of all live births from one academic tertiary hospital resulting from IVF treatment with autologous oocytes during the period from 2009 to 2017., Participants/materials, Setting, Methods: All patients had placentas sent for pathological evaluation irrelevant of maternal or fetal complications status. Placental, obstetric and perinatal outcomes of pregnancies resulting from hormone replacement vitrified embryo transfers were compared with those after fresh embryo transfers. A multivariate analysis was conducted to adjust the results for determinants potentially associated with the development of placental histopathology abnormalities., Main Results and the Role of Chance: A total of 1014 singleton live births were included in the final analysis and were allocated to the group of pregnancies resulting from fresh (n = 660) and hormone replacement frozen (n = 354) embryo transfers. After the adjustment for confounding factors the frozen embryo transfers were found to be significantly associated with chorioamnionitis with maternal (odds ratio (OR) 2.0; 95% CI 1.2-3.3) and fetal response (OR 2.6; 95% CI 1.2-5.7), fetal vascular malperfusion (OR 3.9; 95% CI 1.4-9.2), furcate cord insertion (OR 2.3 95% CI 1.2-5.3), villitis of unknown etiology (OR 2.1; 95% CI 1.1-4.2), intervillous thrombi (OR 2.1; 95% CI 1.3-3.7), subchorionic thrombi (OR 3.4; 95% CI 1.6-7.0), as well as with failure of labor progress (OR 2.5; 95% CI 1.5-4.2)., Limitations, Reasons for Caution: Since the live births resulted from frozen-thawed embryos included treatment cycles with previously failed embryo transfers, the factors over embryo vitrification may affect implantation and placental histopathology., Wider Implications of the Findings: The study results contribute to the understanding of the perinatal future of fresh and vitrified embryos. Our findings may have an implication for the clinical decision to perform fresh or frozen-thawed embryo transfer., Study Funding/competing Interest(s): Authors have not received any funding to support this study. There are no conflicts of interest to declare., Trial Registration Number: N/A., (© The Author(s) 2022. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Robins (1998) introduced marginal structural models, a general class of counterfactual models for the joint effects of time-varying treatments in complex longitudinal studies subject to time-varying confounding. Robins (1998) established the identification of marginal structural model parameters under a sequential randomization assumption, which rules out unmeasured confounding of treatment assignment over time. The marginal structural Cox model is one of the most popular marginal structural models for evaluating the causal effect of time-varying treatments on a censored failure time outcome. In this paper, we establish sufficient conditions for identification of marginal structural Cox model parameters with the aid of a time-varying instrumental variable, in the case where sequential randomization fails to hold due to unmeasured confounding. Our instrumental variable identification condition rules out any interaction between an unmeasured confounder and the instrumental variable in its additive effects on the treatment process, the longitudinal generalization of the identifying condition of Wang & Tchetgen Tchetgen (2018). We describe a large class of weighted estimating equations that give rise to consistent and asymptotically normal estimators of the marginal structural Cox model, thereby extending the standard inverse probability of treatment weighted estimation of marginal structural models to the instrumental variable setting. Our approach is illustrated via extensive simulation studies and an application to estimating the effect of community antiretroviral therapy coverage on HIV incidence., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Biometrika Trust. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Emerging evidence supports the physical health and social benefits of dog ownership. This study examined the longitudinal effect of dog ownership and dog walking on mental health., Methods: Data from a cohort of 1023 participants taking part in the RESIDential Environments project, in Perth, Western Australia were collected over a 2 year period (baseline and follow-up). Self-report survey items measured mental health (stress and depression), dog ownership status and weekly minutes of dog walking. Logistic regression models accounted for potential confounding factors including socio-demographic, self-rated health and baseline mental health., Results: Overall, no statistically significant effects were observed over time between dog ownership and stress (adjusted OR: 1.20; 95% CI: 0.79, 1.81) or depression (adjusted OR: 1.51; 95% CI: 0.72, 3.16). There was a small inverse but non-significant association between weekly minutes of dog walking and stress over time (adjusted OR: 0.85; 95% CI: 0.60, 1.22)., Conclusion: There was little evidence of prospective associations between dog ownership or dog walking and mental health. Further research is required to confirm longitudinal relationships between dog ownership and dog walking and mental health and investigate dog-related factors, such as a person's attachment to their dog., (© The Author(s) 2019. Published by Oxford University Press on behalf of Faculty of Public Health. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Aims: Because the effect of Cd 2+ -immobilized microbe on Cd uptake of plants in Cd-contaminated soil remains underexplored, this study focuses on the effect of Cd 2+ -immobilized rhizobia on Cd uptake of soybean., Methods and Results: Strain KG2 from soybean nodule was identified as Rhizobium pusense KG2 by phylogenetic analysis. Rhizobium pusense KG2 showed the 120 mg l -1 of minimal lethal concentration for Cd 2+ . In 50 and 100 mg l -1 of Cd 2+ liquid, approximately 2 × 10 10 cells removed 56·71 and 22·11% of Cd 2+ , respectively. In pot soil containing 50 and 100 mg kg -1 of Cd 2+ , strain KG2 caused a 45·9 and 35·3% decrease in soybean root Cd content, respectively. Meanwhile, KG2 improved the root and shoot length, nitrogen content and biomass of soybean plants and superoxide dismutase activity., Conclusions: The Cd 2+ -immobilized rhizobia could inhibit soybean plants to absorb Cd 2+ from soil, promote plant growth and improve plant's tolerance against Cd. This study is the first time to report that R. pusense is an effective nodulating rhizobium of legume., Significance and Impact of the Study: Some Cd 2+ -immobilized microbe lowering Cd uptake of plant and promoting plant growth should be considered as an effective strategy for producing safety crops in the Cd-contaminated agricultural soil., (© 2018 The Society for Applied Microbiology.)

Background: Psoriasis is a complex disease influenced by both genetic and environmental factors with abnormal gene expression in lesional skin. However, no studies are available on genome-scale gene expression of psoriatic lesions in the Chinese population. In addition, systematic studies on the biological pathways, pathogenicity and interaction networks of psoriasis-related genes with abnormal expression profiles require further investigation., Objectives: To further explore the associated pathways in psoriasis by functional analysis and to identify the key genes by gene pathogenicity analysis., Methods: We performed RNA sequencing on 60 skin biopsy samples from patients with psoriasis and healthy controls to identify the primary differentially expressed genes in psoriatic lesional skin. We retrieved all reported psoriasis-associated genes and performed integrative analyses covering gene expression profiling, pathway analysis, gene pathogenicities and protein-protein interaction networks., Results: We found that internal and external stimuli may activate immunoinflammatory responses to promote the development of psoriasis. Pathways associated with infectious diseases and cancers were identified by functional and pathway analyses. The gene pathogenicity analysis revealed five key genes in psoriasis: PPARD, GATA3, TIMP3, WNT5A and PTTG1., Conclusions: Our analyses showed that genes contributed to the pathogenesis of psoriasis by activating risk pathways with components abnormality in expression. We identified five potentially pathogenic genes for psoriasis that may serve as important biomarkers for the diagnosis and treatment., (© 2017 British Association of Dermatologists.)

Background: A previous study provided evidence for a genetic association between PPP2CA on 5q31.1 and systemic lupus erythematosus (SLE) across multi-ancestral cohorts, but failed to find significant evidence for an association in the Han Chinese population., Objectives: To explore the association between this locus and SLE using data from our previously published genome-wide association study (GWAS)., Methods: Single-nucleotide polymorphisms (SNPs) rs7726414 and rs244689 (near TCF7 and PPP2CA in 5q31.1) were selected as candidate independent associations from a large-scale study in a Han Chinese population consisting of 1047 cases and 1205 controls. Subsequently, 3509 cases and 8246 controls were genotyped in two further replication studies. We then investigated the SNPs' associations with SLE subphenotypes and gene expression in peripheral blood mononuclear cells., Results: Highly significant associations with SLE in the Han Chinese population were detected for SNPs rs7726414 and rs244689 by combining the genotype data from our previous GWAS and two independent replication cohorts. Further conditional analyses indicated that these two SNPs contribute to disease susceptibility independently. A significant association with SLE, age at diagnosis < 20 years, was found for rs7726414 (P = 0·001). The expression levels of TCF7 and PPP2CA messenger RNA in patients with SLE were significantly decreased compared with those in healthy controls., Conclusions: This study found evidence for multiple associations with SLE in 5q31.1 at genome-wide levels of significance for the first time in a Han Chinese population, in a combined genotype dataset. These findings suggest that variants in the 5q31.1 locus not only provide novel insights into the genetic architecture of SLE, but also contribute to the complex subphenotypes of SLE., (© 2017 British Association of Dermatologists.)

Aims: To analyse whether some indoleacetic acid (IAA)-secreting plant growth-promoting bacteria can alleviate cadmium (Cd) stress, the role of an Ochrobactrum sp. MGJ11 from rhizosphere of soybean in promoting plant growth, and to evaluate the counteracting Cd effects on soybean., Methods and Results: Ochrobactrum sp. MGJ11 produced 121·2 mg l -1 of IAA. MGJ11 increased soybean root length, shoot length and biomass by 30·1, 30·8 and 13·4% respectively. In liquid medium, no IAA production was detected in Cd concentration of 100 mg l -1 . In soil with 20-80 mg kg -1 Cd, MGJ11 promoted soybean root elongation (29·4-161·4%) and increased the shoot length (up to 52·7%) and biomass (up to 87·2%). After growing for 38 days, Cd concentrations in the roots of inoculated soybean were lower than in those of noninoculated plants. Only a little Cd (2·6-16·9 μg g -1 ) was translocated from the root to shoot., Conclusions: Ochrobactrum sp. MGJ11 secretes IAA and shows tolerance against Cd. MGJ11 inoculation improves the root length, shoot length and biomass of soybean in both vermiculite and Cd vermiculite, and decreases Cd concentration of soybean root. The characteristics of MGJ11 suggest that it could be used for promoting soybean growth and lowering bioavailability of soil Cd for soybean root., Significance and Impact of the Study: In this study, we isolated a plant growth-promoting Ochrobactrum with the activity of mitigating Cd toxicity to plant roots. The Ochrobactrum can be considered as a potential bioaugmentation agent that promotes plant growth, especially in some agricultural systems, or that helps in the phytoremediation of Cd-contaminated soil., (© 2016 The Society for Applied Microbiology.)

The defect of esophagus after surgical excision in patients is usually replaced by autologous stomach, jejunum, or colon. The operation brings severe trauma and complications. Using artificial esophagus to replace the defect in situ can reduce the operative trauma, simplify the operative procedures, and decrease the influence to digestive function. A variety of experiments have been designed for developing a practical artificial esophagus. Nevertheless, a safe and reliable artificial esophagus is not yet available. The objective is to evaluate the possibility of the artificial esophagus made of non-degradable polyurethane materials being used in reconstruction of the segmental defect of cervical esophagus in beagles, observe the regeneration of esophageal tissue, and gather experience for future study. The cervical esophageal defects in 13 beagles were designed to 2-cm long and were constructed by the artificial esophagus made of non-degradable polyurethane materials. Nutrition supports were given after the operation. The operative mortality, anastomotic leakage, migration of artificial esophagus, and dysphagia were followed up. The regeneration of the esophageal tissues was evaluated by histopathology and immunohistochemical labeled streptavidin-biotin method. The surgical procedures were successfully completed in all beagles, and 12-month follow-ups were done. Only one beagle died of severe infection, and all others survived until being killed. The anastomotic leakage occurred in nine beagles, most of them (8/9) were cured after supportive therapy. The migration of artificial esophagus occurred in all 12 surviving beagles, and one artificial esophagus stayed in situ after migration. All 12 surviving beagles showed dysphagia with taking only fluid or soft food. No beagle died of malnutrition. The neo-esophagus was composed of granulation tissue, and the inner surface was covered by epithelium in 2-3 months completely. But the inner surface of neo-esophagus with artificial esophagus staying in situ after migration was not covered by epithelium, and the granulation tissue was infiltrated by a great deal of inflammatory cells. Antibodies against cytokeratin were positively expressed in epithelium of neo-esophagus. Up to 12 months after operation, antibodies against smooth muscle actin and desmin were both negatively expressed in neo-esophagus. The artificial esophagus made of non-degradable polyurethane reconstructing cervical esophageal defect is practicable. Although there are some problems, including anastomotic leakage, migration, and dysphagia, they are not lethal following good supportive therapy. The esophageal epithelium can regenerate with the supporting role of artificial esophagus. In the future, deformable artificial esophagus should be improved, and a much longer follow-up will be performed to evaluate whether the esophageal gland and skeletal muscle can regenerate., (© 2014 International Society for Diseases of the Esophagus.)

Peroral endoscopic myotomy (POEM) has been developed as a minimally invasive endoscopic treatment for achalasia for years. However, the optimal length of submucosal tunnel and myotomy of muscle bundles during procedure of POEM has not yet been determined, so we aim to assess safety and efficacy of modified POEM with shorter myotomy of muscle bundles in achalasia patients. Consecutive achalasia patients had been performed modified POEM with shorter myotomy, and assessed by symptoms, high-resolution manometry, and barium swallow examinations before and 3 months after POEM for safety and efficacy evaluation. Modified POEM with shorter submucosal tunnel (mean length 6.8 cm) and endoscopic myotomy of muscle bundles (total mean length 5.4 cm) were completed in 46 consecutive achalasia patients. During the 3-month follow up in all cases, significant improvement of symptoms (a significant drop in the Eckardt score 8.4 ± 3.2 vs. 2.7 ± 1.9; P < 0.001), decreased lower esophageal sphincter pressure (39.4 ± 10.1 vs. 24.4 ± 9.1 mmHg; P < 0.001) and integrated relaxation pressure (38.6 ± 10.4 vs. 25.7 ± 9.6 mmHg; P < 0.01), and a drop in height of esophagus barium-contrast column (5.4 ± 3.1 vs. 2.6 ± 1.8 cm; P < 0.001) were observed. The frequencies of adverse events were lower in those under endotracheal anesthesia and CO2 insufflations compared with intravenous anesthesia and air insufflations. Only three patients were found to have gastroesophageal reflux disease on follow up. Modified POEM with shorter myotomy under endotracheal anesthesia and CO2 insufflations shows its good safety and excellent short-term efficacy in the treatment of achalasia. But further studies are warranted to assess the long-term efficacy., (© 2014 International Society for Diseases of the Esophagus.)

Unlabelled: A novel nanoparticle-assisted polymerase chain reaction (nanoPCR) assay to detect porcine parvovirus (PPV) is described here. Primers for this assay were designed based on the conserved region of the nonstructural protein 1 (NS1) gene of PPV, which encodes one of the nonstructural proteins. The sensitivity of the PPV nanoPCR assay was measured by using diluted recombinant plasmids in which the PPV NS1 gene had been inserted. The detection limit was 5.6 × 10(1) copies μl(-1) for the PPV nanoPCR assay vs 5.6 × 10(3) copies μl(-1) for conventional PCR assay. The results showed that the sensitivity of PPV nanoPCR assay was 100 times higher than that of conventional PCR assay. The PPV nanoPCR assay produced 142-bp product as expected when amplifying PPV DNA, while produced nothing when amplifying the DNA or cDNA of the following viruses: swine encephalomyocarditis virus, classical swine fever virus, porcine pseudorabies virus, porcine reproductive and respiratory syndrome virus, porcine teschovirus and porcine circovirus type II. PPV was detected in 108 of 109 clinical swine samples from Heilongjiang, Jilin and Henan provinces using the nanoPCR assay, and the results were confirmed by sequencing., Significance and Impact of the Study: Nanoparticle-assisted polymerase chain reaction (nanoPCR) assay is an improved PCR. NanoPCR is highly sensitive and specific because the nanofluids formed in the nanobuffer have high thermal conductivity, which reduces the time required to reach the target temperature. It is more sensitive than conventional PCR, and it could detect the cases earlier than conventional PCR. This report describes the first application of the highly efficient nanoPCR technology for the detection of porcine parvovirus (PPV). The PPV nanoPCR assay will be useful for the detection and study of PPV and will also be applicable to improve the detection of other viruses., (© 2013 The Society for Applied Microbiology.)

Aims: Natural products have been used as potentially important sources of novel antibacterials in combating pathogenic Staphylococcus aureus isolates, a major problem around the world. In this study, we aimed to investigate the antibacterial effects of pinocembrin (PNCB) against Staph. aureus pneumonia in a murine model and its influence on the production of Staph. aureus α-haemolysin (Hla)., Methods and Results: The in vitro activities of PNCB on α-haemolysin production were determined using haemolysis, Western blot and real-time RT-PCR assays. The viability and cytotoxicity assays were performed to evaluate the influence of PNCB on α-toxin-mediated injury of human alveolar epithelial cells. Moreover, through histopathologic analysis, we further determined the in vivo effects of PNCB on Staph. aureus pneumonia in a mouse model. In vitro, PNCB at low concentrations exhibited inhibitory activity against α-haemolysin production and attenuated α-haemolysin-mediated cell injury. Furthermore, the in vivo findings demonstrated that PNCB protected mice from Staph. aureus pneumonia., Conclusions: We have provided new evidence of the effects of PNCB, which suggest that PNCB attenuated α-haemolysin-mediated cell injury and protected mice from Staph. aureus pneumonia., Significance and Impact of the Study: The findings indicate that PNCB may be used as a basis for anti-Staphylococcus agent., (Journal of Applied Microbiology © 2013 The Society for Applied Microbiology.)

Objectives: The objective of this study was to compare the image quality and radiation dose of chest CT images reconstructed with a blend of adaptive statistical iterative reconstruction (ASIR) and filtered back-projection (FBP) with images generated using conventional FBP., Methods: Patients with chest CT re-examinations were alternately assigned to two scanners with different reconstruction techniques. The study groups included noise index (NI) 11 with 30% ASIR (A30), NI 13 with 40% ASIR (A40), NI 15 with 50% ASIR (A50) and NI 17 with 60% ASIR (A60), sequentially changed every 2 months. The control images were obtained using FBP and NI 11. All acquisitions were performed with automatic dose modulation. Paired t-test and non-parameter test were applied to compare the difference., Results: The radiation doses were significantly lower in the examinations that used ASIR (p<0.001). The mean dose reduction rate was 27.7%, 45.2%, 57.1% and 71.8% for Groups A30, A40, A50 and A60, respectively. The image quality of Groups A30-A50 was not inferior to that of the control examinations. The image noise of Group A60 was greater and subjective image quality was inferior to that of the control., Conclusions: ASIR enabled the use of a higher NI with automatic dose modulation. With 50% ASIR and a NI of 15, the effective radiation dose was reduced by 57%, without compromising image quality.

Objective: To provide evidence for the selection of an optimal cross-sectional reconstruction mode in spectral CT imaging of the abdomen, we compared the monochromatic images with polychromatic images., Methods: Three phase-enhanced CT scans of the abdomen were recorded using the spectral imaging technique on 100 patients. Images were reconstructed using two modes: polychromatic and 70 keV monochromatic. The following variables were then compared: contrast-to-noise ratio (CNR) of the liver, spleen, gallbladder, kidney and pancreas, and the noise. Paired t-tests were used to compare differences between the two sets of images. Three experienced doctors graded the quality of the images with a five-point scale. The image quality scores were compared with a non-parametric rank sum test., Results: Compared with polychromatic images, the 70 keV monochromatic mode images yielded significantly greater tissue-to-fat CNR and lower noise (p<0.001 for all comparisons). The image quality of the 70 keV monochromatic mode showed significantly better results than the polychromatic mode (p<0.001)., Conclusions: In abdominal spectral CT imaging, 70 keV monochromatic mode reconstruction images were better than those reconstructed using the polychromatic mode. The monochromatic mode may become the routine reconstruction mode for cross-sectional images.

Caspase-3 plays an important role as the key effector during apoptosis, but there are very few studies of caspase-3 in esophageal squamous cell carcinoma (ESCC). The purpose of this study was to investigate the expression and prognostic significance of caspase-3 in ESCC from Linzhou City, a high incidence area in northern China. All 64 patients underwent esophagectomy for ESCC between January 2002 and December were enrolled in this study. Caspase-3 expression was assessed by immunohistochemistry (IHC) in primary ESCC and paired normal esophageal epithelium. The positive rate of caspase-3 expression was higher in ESCC than in normal esophageal epithelium (79.7% vs. 50.0%, Chi-square = 12.372, P= 0.001). Caspase-3 expression was correlated with tumor cell differentiation (Phi = 0.717, P < 0.001), tumor infiltration depth (Phi =-0.334, P= 0.008), and pathologic TNM (pTNM) staging (rs =-0.268, P= 0.032). Patients in caspase-3 positive group had a significantly better 5-year overall survival than those in the negative group (77.4% vs. 35.9%, chi(2)= 7.344, P= 0.007). Our results showed that caspase-3 expression was upregulated in ESCC compared with normal esophageal epithelium in population of Chinese high incidence area, and patients with caspase-3 positive expression had better prognosis. Therefore, caspase-3 immunostaining could be a simple and useful tool for predicting survival in ESCC patients.

Aims: Gas chromatography (GC) was utilized to investigate the cellular fatty acids (CFAs) composition of 141 Yersinia pestis isolates from different plague foci of China, and 20 Yersinia pseudotuberculosis strains as well., Methods and Results: The whole cell fatty acid methyl esters (FAMEs) were obtained by saponification, methylation and extraction followed with analysis using a standardized Microbial Identification System (MIS). Y. pestis and Y. pseudotuberculosis strains are quite similar in major CFA profiles, which include 16:0, 17:0 cyclo, 3-OH-14:0, 16:1 omega 7c and 18:1 omega 7c, accounting for more than 80% of the total CFAs., Conclusions: Yersinia pestis could be easily differentiated from Y. pseudotuberculosis by plotting the ratios of some CFA pairs, i.e.,14:0/18:0 vs 18:1 omega 7c/18:0, 3-OH-14:0/18:0 vs 18:1 omega 7c/18:0, 16:1 omega 7c/18:0 vs 18:1 omega 7c/18:0, 12:0/18:0 vs 18:1 omega 7c/18:0 and 12:0 ALDE/18:0 vs 16:1 omega 7c/18:0 fatty acids., Significance and Impact of the Study: In the present study, the normalized Sherlock MIS and Sherlock standard libraries were used to analyse the fatty acid composition of different strains of Y. pestis and Y. pseudotuberculosis. Meanwhile, ratios of certain CFA components were found to serve as chemical markers for differentiating the two closely related bacteria that are difficult to be differentiated by simply comparing CFA profiles based on other researches.

Epidermolysis bullosa simplex (EBS) is a group of inherited skin diseases, characterized by the formation of intraepidermal blisters. We performed genetic analysis of the keratin 5 (KRT5) gene in two Chinese pedigrees. One novel missense mutation was identified in a patient with sporadic EBS (general, non-Dowling-Meara). Sequence analysis showed a heterozygous T > A transition at nucleotide 1730 of KRT5, changing phenylalanine (Phe) to tyrosine (Tyr) at position 577 of the keratin 5 (K5). In addition, two recurrent mutations c.1649delG (p.Gly550AlafsX77) and c.508G > (p.Glu170Lys) in KRT5 were identified in Chinese patients with mottled pigmentation EBS and localized EBS, respectively. None of the mutations were found in any unaffected family members or in an additional 100 unrelated control samples. These results suggest that these mutations are pathogenic and might be one of the potential causes of EBS in these Chinese patients.

Background: Psoriasis is a common inflammatory and hyperproliferative skin disease. The pathogenesis of psoriasis remains obscure. Family and twin studies have suggested a strong genetic susceptibility to psoriasis. Eight linkage loci (PSORS1-7, PSORS9) were identified and accepted by the OMIM and an additional 16 susceptibility loci have been suggested so far., Objectives: To investigate further three suggested psoriasis susceptibility loci at 2p22.3-11.2, 13q21-32 and 17q22-25.3 in a Chinese population. Using an expanded sample of 180 Chinese families with psoriasis and improved marker coverage, we verified whether they were Chinese Han psoriasis susceptibility loci., Methods: In total, 180 Chinese Han families with psoriasis vulgaris (including the 61 families used in the original genome-wide scan and 119 new families) were recruited from the Dermatology Department at the First Hospital Affiliated to Anhui Medical University. Two-point and multipoint parametric and nonparametric linkage (NPL) analyses were performed at 2p, 13q and 17q in the total 180 families as well as the 61 original and 119 new families separately., Results: At the region 2p, a maximum multipoint NPL score of 4.11 was identified at locus D2S337 (P=0.000003), and a maximum multipoint heterogeneity LOD (HLOD) score of 4.93 (alpha=54%) was identified at the same locus in the analysis of the 180 families. However, the analysis of the 180 families did not identify any significant linkage evidence at the region 13q21-32 [a maximum multipoint HLOD score of 0.10 (alpha=7%) and NPL score of 0.95 (P=0.14)] or the region 17q22-25.3 [a maximum multipoint HLOD score of 0.08 (alpha=6%) and NPL value of 0.94 (P=0.14)]. For these two regions, the LOD scores from the 180 families as well as the 119 new families were much smaller than the ones obtained from the original 61 families., Conclusions: Our study indicates that 2p22.3-11.2 is a novel psoriasis susceptibility locus in the Chinese Han population and confirms that psoriasis is a genetically heterogeneous disease.

Aberrant methylation of tumor suppressor genes plays an important role in the development of esophageal squamous cell carcinoma (ESCC). The purpose of the present study was to identify the epigenetic changes in ESCC. Methylation-sensitive arbitrarily primed polymerase chain reaction (MS AP-PCR) analysis was used on 22 matched ESCC tumors and adjacent normal tissues. Through this screen we identified a frequently methylated fragment that showed a high homology to the 5'-CpG island of the gene encoding a transmembrane protein containing epidermal growth factor and follistatin domains (TPEF). The methylation status of the TPEF gene was then detected by bisulfite sequencing and the levels of TPEF mRNA were detected by RT-PCR. In addition, the effects of a methylation inhibitor 5-aza-2'-deoxycytidine on TPEF mRNA expression was determined in cells of ESCC cell lines. Hypermethylation of the 5'-CpG island of TPEF was found in 12 of 22 (54.5%) primary tumors. Reverse transcription PCR analysis demonstrated that TPEF mRNA expression was significantly lower in tumors than in adjacent normal tissues, which is associated with promoter hypermethylation. In addition, treatment of ESCC cell lines with 5-aza-2'-deoxycytidine led to re-expression of the TPEF transcript. In conclusion, we observed promoter of TPEF gene is frequently hpermethylated, and is associated with the loss of TPEF mRNA expression in ESCC samples. Promoter hypermethylation of TPEF gene may play a role in the development of ESCC.

Background: Acrokeratosis verruciformis of Hopf (AKV) is a rare genodermatosis characterized by multiple flat-topped, flesh-coloured papules on the dorsa of hands and feet, and punctuate keratoses on the palms and soles. A mutation in the ATP2A2 gene has been shown to be associated with AKV and with Darier's disease (DD)., Objectives: To explore the molecular aetiology of AKV and DD., Methods: We investigated the clinical and histological information in two families and a sporadic case with AKV and one family and a sporadic case with DD in China. Mutation analysis of ATP2A2 was performed by PCR and direct sequencing, and genotyping and linkage analysis performed using six polymorphic microsatellite markers spanning the locus at 12q23-12q24 containing ATP2A2., Results: Mutational analysis showed no mutation in ATP2A2 among the AKV patients, but we found two novel mutations (p.C318F and p.M719fs) in the DD patients. The genotyping and linkage analysis results revealed no linkage evidence of the locus at 12q23-12q24 in a large AKV family., Conclusions: Our findings provide evidence for the genetic heterogeneity of AKV and demonstrate that mutations in genes other than ATP2A2 are responsible for AKV in a proportion of the Chinese population.

Background: Punctate palmoplantar keratoderma (PPK) is a rare autosomal dominant cutaneous disorder characterized by numerous hyperkeratotic papules distributed on the palms and soles. Two loci for punctate PPK were recently found to be located on 8q24.13-8q24.21 and 15q22-15q24. However, no genes for this disease have been identified to date. Objectives To refine the previously mapped regions and to identify the disease gene locus in a four-generation Chinese family with punctate PPK., Methods: Genetic linkage analysis was carried out in this family using microsatellite markers on chromosomes 8q and 15q. Two-point linkage analysis was performed using Linkage programs version 5.10 and the haplotype was constructed using Cyrillic version 2.02 software., Results: We failed to confirm our previous locus at 8q24.13-8q24.21, but significant evidence for linkage was observed in the region of 15q with a maximum two-point LOD score of 5.38 at D15S153 (theta = 0.00). Haplotype analysis localized the punctate PPK locus within the region defined by D15S651 and D15S988. This region overlaps by 5.06 cM with the previously reported punctate PPK region., Conclusions: This study refines a disease gene causing punctate PPK to a 5.06-cM interval at 15q22.2-15q22.31.

Marie Unna hereditary hypotrichosis (MUHH) is a rare autosomal dominant disorder with progressive hair loss starting in early childhood and aggravating at puberty. Several studies have mapped the MUHH gene to chromosome 8p21. Here we report a Chinese MUHH family with variable phenotypes. All affected individuals have anomalies affecting both hair density and hair shafts. Major clinical characteristics, disease history and histological examination support the diagnosis of MUHH, but the features of scarring in this kindred are modest and none of the patients have vertex hair loss, which is in contrast with typical MUHH. We now report genotyping and linkage analysis using 11 polymorphic microsatellite markers spanning the MUHH locus at 8p. Two-point linkage analysis using these markers revealed significant exclusion of this locus (log of the odds scores < - 2) at Theta = 0 indicating that there is a range of clinical presentations in MUHH, and that more than one genetic locus is responsible for the disorder.

Background: Marie Unna hereditary hypotrichosis (MUHH) is a rare autosomal congenital alopecia with progressive hair loss starting in early childhood and accelerating at puberty. A locus for MUHH has been mapped on chromosome 8p21 but no genes for MUHH have been identified to date., Objectives: To refine the MUHH locus to a narrow chromosome region to facilitate cloning of the gene., Methods: We performed genotyping and linkage analysis in a multigeneration Chinese family with MUHH, using 18 high-density microsatellite markers spanning the previously mapped interval at 8p21., Results: Significant evidence for linkage was observed in this region, with a maximum two-point LOD score of 3.01 (theta = 0). Haplotype analysis localized the MUHH locus within the region defined by D8S282 and D8S1839. This region overlaps by 1.1-cM with the previously reported MUHH region and represents a physical distance of about 380 kb., Conclusions: This study provides a refined map location (1.1 cM) for isolation of the gene causing MUHH. These data also indicate the existence of a common MUHH locus at 8p21.3 between affected caucasian and Chinese families.

Background: Dyschromatosis symmetrica hereditaria (DSH) is an autosomal dominant pigmentary genodermatosis characterized by hyperpigmented and hypopigmented macules on the extremities, which has recently been mapped to an 11.6-cM interval on chromosome 1q11-21. So far, most cases of DSH have been reported in Japan and dermatologists around the world might think this disorder mainly occurs in Japan. In fact, there are 17 DSH families including 136 cases reported in China since 1980, but most of them are described in Chinese., Objectives: To refine the previously mapped region that facilitates the identification of the DSH gene and to delineate the clinical and genetic features of Chinese DSH cases by a literature review of 136 cases reported in China., Methods: We performed genotyping and linkage analysis using polymorphic microsatellite markers at 1q11-22 in two Chinese DSH families, and reviewed all of the DSH cases reported in China since 1980., Results: A cumulative maximum two-point lod score of 3.68 was produced with marker D1S506 at a recombination frequency of theta = 0.00 in these two families. Haplotype analysis refined the DSH locus to a 9.4-cM interval flanked by D1S2343 and D1S2635. The genetic and clinical features of Chinese cases with DSH were summarized. In some Chinese cases, hyperpigmented and hypopigmented macules were scattered on the neck and chest, but among Japanese patients there were no similar skin lesions to be reported on these sites., Conclusions: This study confirms linkage of DSH to a previously mapped region and refines the DSH gene to a 9.4-cM interval at 1q21-22. Likewise, the literature review indicates that DSH is not an uncommon disorder in China and the differences in the distribution of skin lesions could be related to race and environment.

The CCR4-NOT complex is involved in the regulation of gene expression both positively and negatively. The repressive effects of the complex appear to result in part from restricting TBP access to noncanonical TATAA binding sites presumably through interaction with multiple TAF proteins. We provide here genetic evidence that the CCR4-NOT complex also plays a role in transcriptional elongation. First, defects in CCR4-NOT components as well as overexpression of the NOT4 gene elicited 6-azauracil (6AU) and mycophenolic acid sensitivities, hallmarks of transcriptional elongation defects. A number of other transcription initiation factors known to interact with the CCR4-NOT complex did not elicit these phenotypes nor did defects in factors that reduced mRNA degradation and hence the recycling of NTPs. Second, deletion of ccr4 resulted in severe synthetic effects with mutations or deletions in the known elongation factors RPB2, TFIIS, and SPT16. Third, the ccr4 deletion displayed allele-specific interactions with rpb1 alleles that are thought to be important in the control of elongation. Finally, we found that a ccr4 deletion as well as overexpression of the NOT1 gene specifically suppressed the cold-sensitive phenotype associated with the spt5-242 allele. The only other known suppressors of this spt5-242 allele are factors involved in slowing transcriptional elongation. These genetic results are consistent with the model that the CCR4-NOT complex, in addition to its known effects on initiation, plays a role in aiding the elongation process.

Dendritic cells (DCs) are essential for the presentation of antigens in the primary immune response. To examine the generation of DCs from hemopoietic stem cells in the bone marrow (BM), lineage-negative (Lin-)/CD71- bone marrow cells (BMCs) from C57BL/6 mice were separated into major histocompatibility complex (MHC) class Ihigh/ c-kit(low) and MHC class Ihigh/c-kit(low)(phenotypically c-kit-negative, but c-kit message only detected by reverse transcriptase-polymerase chain reaction) populations. A large number of cells with the morphological, phenotypical, and functional characteristics of DCs was generated from both c-kit(low) and c-kit(low) populations when cultured with a combination of cytokines (GM-CSF, tumor necrosis factor-a [TNF-a], interleukin 7 [IL-7], IL-3, stem cell factor [SCF], and flt3 ligand); the cytokine combination studies revealed that SCF and IL-3 in addition to GM-CSF and TNF-a are essential for DCs to be generated from these primitive populations. To our surprise most (>80%) generated cells expressed high levels of DC surface markers such as DEC205 and MHC class II, and they were potent stimulators in the primary allogeneic T cell activation. The development of DCs from c-kit(

Self-incompatibility (SI) in Brassica is controlled by a single locus, termed the S locus. There is evidence that two of the S locus genes, SLG, which encodes a secreted glycoprotein, and SRK, which encodes a putative receptor kinase, are required for SI on the stigma side. The current model postulates that a pollen ligand recognizing the SLG/SRK receptors is encoded in the genomic region defined by the SLG and SRK genes. A fosmid contig of approximately 65 kb spanning the SLG-910 and SRK-910 genes was isolated from the Brassica napus W1 line. A new gene, SLL3, was identified using a novel approach combining cDNA subtraction and direct selection. This gene encodes a putative secreted small peptide and exists as multiple copies in the Brassica genome. Sequencing analysis of the 65-kb contig revealed seven additional genes and a transposon. None of these seven genes exhibited features expected of S genes on the pollen side. An approximately 88-kb contig of the A14 S region also was isolated from the B. napus T2 line and sequenced. Comparison of the two S regions revealed that (1) the gene organization downstream of SLG in both S haplotypes is highly colinear; (2) the distance between SLG-A14 and SRK-A14 genes is much larger than that between SLG-910 and SRK-910, with the intervening region filled with retroelements and haplotype-specific genes; and (3) the gene organization downstream of SRK in the two haplotypes is divergent. These observations lead us to propose that the SLG downstream region might be one border of the S locus and that the accumulation of heteromorphic sequences, such as retroelements as well as haplotype-unique genes, may act as a mechanism to suppress recombination between SLG and SRK.

Using Ly5 congenic mice, we characterized the early differentiation step of pluripotent hemopoietic stem cells. Lineage- (Lin-)/CD71- cells in the bone marrow cells were separated into major histocompatibility complex (MHC) class I(high)/c-kit(low) and MHC class I(high)/c-kit

Esophagectomy with esophagogastric anastomosis is commonly complicated by anastomotic dehiscence. Although this is a major problem in clinical esophageal surgery, laboratory investigation of esophagogastric anastomotic wound healing has been hampered by the lack of a practical rodent model. Problems with aspiration pneumonia and anastomotic strictures hindered our previous studies in the rat. Other researchers have turned to large animal experiments, or used various upper gastrointestinal pseudoanastomotic techniques in rodents. None of these approaches has proved satisfactory. We developed a technique of side-to-side esophagogastric anastomosis in the rat, and then studied normal esophagogastric anastomotic healing in this model. Anastomoses were performed in 24 rats. Anastomotic breaking strength and hydroxyproline concentration were measured 1 and 2 weeks after surgery. Anastomotic breaking strength was 3.78 +/- 1.18 N at 1 week and 4.83 +/- 0.91 N after 2 weeks (p < 0.03). Anastomotic tissue hydroxyproline concentration was 370.6 +/- 31.2 nmol/mg at 1 week and 462.1 +/- 69.7 nmol/mg after 2 weeks (p < 0.001). Many of the problems encountered in esophagogastric anastomotic studies in the rat have been overcome using this new model.

When grown in medium containing ascorbic acid and beta-glycerol phosphate, mouse MC3T3-E1 cells express an osteoblast phenotype and produce a highly mineralized extracellular matrix. The purpose of this study was to independently examine the role of the collagenous matrix and functional osteoblasts on the mineralization process. Cultures with and without an extensive collagenous matrix were prepared by growing MC3T3-E1 cells in the presence and absence of ascorbic acid. Matrix-rich cultures mineralized at much lower calcium phosphate ion products than nonmatrix cultures. At higher ion products, spontaneous precipitation in the medium and cell layers of nonmatrix cultures were observed. In contrast, mineral in matrix-rich cultures was still exclusively associated with collagen fibrils and not with ectopic sites in the cell layer or medium. To examine the effect of cell viability on matrix mineralization, cells were grown 8 or 16 days in the presence of ascorbic acid, then killed and incubated in a mineralizing medium. Significant mineralization was not observed in the collagenous matrix of 8-day killed cultures or age-matched controls. At 16 days mineral was associated with collagen fibrils at specific foci in the matrix of both viable and killed cultures. This observation is consistent with the concept that collagenous matrices must undergo a maturation process before they can support a mineral induction and growth. It further shows that osteoblast-like cells are not required for mineralization of mature matrices, but are required for matrix maturation.

This study was undertaken to examine the mechanisms involved in polymorphonuclear leukocyte superoxide release stimulated by exogenous phosphatidic acid (PA). Unlike the immediate burst of superoxide release affected by membrane-permeable dioctanoylglycerol (DiC8-DAG), dioctanoyl phosphatidic acid (DiC8-PA) induced superoxide release after a lag period of 5-20 min. This period was considerably reduced or eliminated when cells were primed by substimulatory levels of phorbol myristate acetate (PMA). Granule-depleted neutrophil cytoplasts also responded to DiC8-PA with a burst of superoxide generation. Activation of the cytoplast superoxide generating system in response to DiC8-PA was also significantly faster after cells had been preexposed to substimulatory levels of PMA, indicating that at least a portion of the priming mechanism was independent of PMA-induced degranulation. To further examine the potential mechanism of PMA priming of responses to PA, we evaluated the activity of neutrophil ecto-phosphatidic acid phosphohydrolase (ecto-PA phosphohydrolase), which generates diacylglycerol from exogenous PA. PMA priming had no discernable effect on the activity of this enzyme. In addition, propranolol, an inhibitor of PA phosphohydrolase, did not selectively inhibit PMA priming of neutrophil responses to DiC8-PA, indicating that priming did not result from acceleration of DiC8-PA hydrolysis. We therefore investigated the possibility that activation of protein kinase C was the basis of the primed response. Several semiselective protein kinase C inhibitors (calphostin C, H-7, and acylmethylglycerol) inhibited DiC8-DAG- and DiC8-PA-induced superoxide release as well as PMA-primed responses to approximately the same extent. These results are consistent with the hypothesis that neutrophil responses to phosphatidate are mediated by diglyceride generated by the action of ecto-PA phosphohydrolase. PMA priming does not result from increased catalytic activity of ecto-PA phosphohydrolase but rather seems to result from potentiation of an intermediate involved in the cells' response to multiple stimuli.

TGF beta has opposing effects on osteoblasts which are thought to be differentiation stage dependent; however, little is known concerning the effects of TGF beta on osteoblastic characteristics at different stages of maturation. The purpose of this study was to characterize the pattern of mRNA expression for the PTH/PTHrP receptor during normal osteoblastic differentiation in vitro, and evaluate the effects of TGF beta 1 on PTH/PTHrP receptor and osteocalcin (OCN) steady-state mRNA at different stages of osteoblastic differentiation. MC3T3-E1 preosteoblasts were plated at low density and induced to differentiate with ascorbic acid and beta-glycerophosphate. The first group served as a vehicle control and the remaining five groups received a single 48 h TGF beta 1 (3.0 ng/ml)-pulse staggered on a weekly basis for 30 days. Cell cultures were harvested weekly and evaluated for: steady-state PTH/PTHrP receptor and OCN mRNA levels via northern analysis, calcium and phosphorous levels, bone nodules via Von Kossa staining, alkaline phosphatase enzyme levels, and hydroxyproline levels. Group 1 (control) samples followed a normal pattern of proliferation, extracellular matrix deposition, and mineralization. PTH/PTHrP receptor and OCN mRNA expression increased 8-fold and 10-fold respectively, over the collection periods. When TGF beta 1 was administered during the first 48 h period (group 2) while cells were rapidly proliferating, there was a persistent inhibition of PTH/PTHrP receptor expression and a striking reduction in OCN mRNA expression at all time points. There was also a down-regulation of PTH/PTHrP receptor and OCN expression when TGF beta 1 was administered later during osteoblast differentiation (groups 3-6); however, these effects were not persistent. In addition there was a total lack of bone nodule formation in group two cultures, whereas groups 3-6 had increasing bone nodule formation because the TGF beta 1 was administered later in the culture period. These studies indicate that expression of the PTH/PTHrP receptor increases with osteoblastic differentiation and suggest that TGF beta 1 inhibits osteoblastic maturation with more persistent effects found in less differentiated osteoblastic cells.

The uterine endometria of rabbits induced into pseudopregnancy by intramuscular injection of 17 beta-estradiol, followed by intravenous injection of human chorionic gonadotropin, expressed cholesterol sulfate at a significantly high concentration. The highest concentration of cholesterol sulfate was observed 4 days after the injection of gonadotropin for formation of the corpus luteum, being 10 times higher than that in nonpregnant endometria, and 15.2% of the total cholesterol in the endometrium was converted to the sulfated form, whose percentage in nonpregnant endometrium was 3.2%. However, no significant change in the concentration of gangliosides was observed during the period of pseudopregnancy. In the pseudopregnant endometria, the activity of cholesterol sulfotransferase, a cytosolic thiol enzyme, was increased thirtyfold over that in the nonpregnant endometria, whereas cholesterol sulfate sulfatase, a microsomal enzyme, exhibited approximately one-tenth of the activity in nonpregnant endometria. Arylsulfatase C, but not arylsulfatases A and B, exhibited the same change in activity as cholesterol sulfate sulfatase. Thus, the striking increase in cholesterol sulfate after induction of pseudopregnancy was found to be due to the activation of cholesterol sulfotransferase and the simultaneous inhibition of cholesterol sulfate sulfatase.

Treatment of mouse MC3T3-E1 cells with ascorbic acid initiates the formation of a collagenous extracellular matrix and synthesis of several osteoblast-related proteins. We recently showed that ascorbic acid dramatically increases alkaline phosphatase and osteocalcin mRNAs and that this induction is blocked by inhibitors of collagen triple-helix formation (Franceschi and Iyer, J Bone Miner Res 7:235). In the present study, the relationship between collagen matrix formation and osteoblast-specific gene expression is explored in greater detail. Kinetic studies revealed that ascorbic acid increased proline hydroxylation in the intracellular procollagen pool within 1 h and stimulated the cleavage of type I collagen propeptides beginning at 2.5 h. Mature alpha 1(I) and alpha 2(I) collagen components were first detected at 10 h and continued to increase in both cell layer and culture medium for up to 72 h. Ascorbic acid also increased the rate of procollagen secretion from cell layers to culture medium. The secretion of another matrix protein, fibronectin, was only slightly affected. Alkaline phosphatase or its mRNA was first detected 2-3 days after ascorbic acid addition, but osteocalcin mRNA was not seen until day 6. Two inhibitors of collagen triple-helix formation, ethyl-3,4-dihydroxybenzoate and 3,4-dehydroproline, inhibited procollagen hydroxylation and alkaline phosphatase induction. 3,4-Dehydroproline also inhibited the induction of alkaline phosphatase and osteocalcin mRNAs. Surprisingly, induction was not blocked if cells were exposed to ascorbic acid before inhibitor addition. Alkaline phosphatase was also partially inhibited if cells were grown in the presence of purified bacterial collagenase. These results indicate that the induction of osteoblast markers by ascorbic acid does not require the continuous hydroxylation and processing of procollagens and suggest that a stable, possibly matrix-associated signal is generated at early times after ascorbic acid addition that allows subsequent induction of osteoblast-related genes.