Search

Rationale: Renal cysts in patients with autosomal dominant polycystic kidney disease (ADPKD) can originate from any nephron segments, including proximal tubules (PT), the loop of Henle (LOH), distal tubules (DT), and collecting ducts (CD). Previous studies mostly used limited cell markers and failed to identify cells negative for these markers. Therefore, the cell composition and origin of ADPKD cyst are still unclear, and mechanisms of cystogenesis of different origins await further exploration. Methods: We performed single-cell RNA sequencing for the normal kidney tissue and seven cysts derived from superficial or deep layers of the polycystic kidney from an ADPKD patient. Results: Twelve cell types were identified and analyzed. We found that a renal cyst could be derived either from CD or both PT and LOH. Gene set variation analysis (GSVA) showed that epithelial mesenchymal transition (EMT), TNFA signaling via the NFKB pathways, and xenobiotic metabolism were significantly activated in PT-derived cyst epithelial cells while robust expression of genes involved in G2M Checkpoint, mTORC1 signaling, E2F Targets, MYC Targets V1, MYC Targets V2 were observed in CD-derived cells. Conclusion: Our results revealed that a single cyst could originate from CD or both PT and LOH, suggesting heterogeneity of polycystic composition and origin. Furthermore, cyst epithelial cells with different origins have different gene set activation.

Twins with Duchenne muscular dystrophy (DMD) have been widely studied. We report the first rare case of monozygotic triplets with DMD who shared consistent phenotypes, including delayed motor and language milestones, muscle wasting and weakness, joint contracture, and lumbar lordosis. Muscle magnetic resonance imaging and biopsy revealed the similar muscle injury characteristics and dystrophin absence. Short tandem repeat analysis confirmed monozygosity. A de novo mutation (exon 49-52 deletion) was found in the triplets but not in their mother. Treatment included prednisone, idebenone, and rehabilitation management. At the 2-year follow-up, motor function had deteriorated, and muscle fatty infiltration was more extensive and severe. Our case offers a unique opportunity for genetic and therapeutic research. Furthermore, it highlights the critical role of genetic factors in DMD phenotypes and provides a potential choice for treatment observations.

Autosomal-dominant hypocalcification amelogenesis imperfecta (ADHCAI) is a hereditary disease characterized by enamel defects. ADHCAI is mainly caused by nonsense mutations in a gene called family with sequence similarity 83 member H (FAM83H). To study the pathogenesis of ADHCAI, a Chinese ADHCAI family was investigated. The ultrastructure of enamel was analyzed by micro-CT and scanning electron microscopy. Whole-exome sequencing (WES) was performed to identify the pathogenic gene. The function of the mutant FAM83H was studied by real-time PCR, western blotting, subcellular localization, and protein degradation pathway analyses. WES identified a known nonsense mutation (c.1915A > T) in exon 5 of the FAM83H gene, causing a truncated protein (p.Lys639*). However, the cases reported herein exhibited significant differences in the clinical phenotype compared with that the previously reported case. An abnormal enamel rod head structure was observed in affected teeth. In vitro functional studies showed altered protein localization and a decreased protein degradation rate for mutant FAM83H. We verified the FAM83H p.Lys639* protein as a gain-of-function variant causing ADHCAI. Abnormal enamel rod head structure was observed in teeth with mutant FAM83H proteins. We also investigated the molecular pathogenesis and presented data on the abnormal degradation of mutant FAM83H proteins. This study helped the family members to understand the disease progression and provided new insights into the pathogenesis of ADHCAI. Due to the large heterogeneity of ADHCAI, this study also provided a genetic basis for individuals who exhibit similar clinical phenotypes.

Inflammation-modulating nutrients and inflammatory markers are established cancer risk factors, however, evidence regarding the association between post-diagnosis diet-associated inflammation and breast cancer survival is relatively sparse. We aimed to examine the association between post-diagnosis dietary inflammatory index (DII®) and risks of all-cause and breast cancer-specific mortality. A total of 1064 female breast cancer survivors in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening (PLCO) Trial prospective cohort, were included in this analysis if they had completed the diet history questionnaire (DHQ). Energy-adjusted DII (E-DIITM) scores were calculated based on food and supplement intake. Cox regression and competing risk models were used to estimate multivariable-adjusted hazards ratios (HRs) and 95% confidence intervals (95% CIs) by E-DII tertile (T) for all-cause and breast cancer-specific mortality. With median follow-up of 14.6 years, there were 296 (27.8%) deaths from all causes and 100 (9.4%) breast cancer-specific death. The E-DII was associated with all-cause mortality (HR T3 vs T1, 1.34; 95% CI, 1.01–1.81; Ptrend, 0.049, Table 2) and breast cancer mortality (HR T3 vs T1, 1.47; 95% CI, 0.89–2.43; Ptrend, 0.13; multivariable-adjusted HR for 1-unit increment: 1.10; 95% CI: 1.00–1.22). Non-linear positive dose–response associations with mortality from all causes were identified for E-DII scores (Pnon-linearity

Liver transplantation has been widely accepted as an effective intervention for end-stage liver diseases and early hepatocellular carcinomas. However, a variety of postoperative complications and adverse reactions have baffled medical staff and patients. Currently, transplantation monitoring relies primarily on nonspecific biochemical tests, whereas diagnosis of multiple complications depends on invasive pathological examination. Therefore, a noninvasive monitoring method with high selectivity and specificity is desperately needed. This review summarized the potential of endogenous small-molecule metabolites as biomarkers for assessing graft function, ischemia-reperfusion injury and liver rejection. Exogenous metabolites, mainly those immunosuppressive agents with high intra- and inter-individual variability, were also discussed for transplantation monitoring.

OBJECTIVES The aim is to evaluate the efficacy and complications of percutaneous transluminal angioplasty (PTA)/stenting in the treatment of transplant renal artery stenosis (TRAS). BACKGROUND TRAS is a relatively rare condition, and currently, there is not enough study about interventional therapy for TRAS. METHODS Between April 2011 and July 2018, 33 patients with TRAS underwent interventional therapy. Analysis of parameters was as follows: technical success, pretreatment and posttreatment serum creatinine, and blood pressure, and vessel patency via ultrasound at 1, 6, and 12 months posttreatment and once a year thereafter. RESULTS One procedure failed. The success rate of PTA/stenting placement was 97.0%. Fourteen PTAs with 16 stents were primary interventions, with 2 stent procedures performed subsequently due to restenosis; the restenosis rate was 6.3%. During the follow-up period, two patients progressed to graft renal failure and three patients were lost to follow-up. The rest of the patients still had stable graft function and blood pressure. Compared with preoperative conditions, blood pressure and serum creatinine significantly decreased (p

Background: Wolf-Hirschhorn Syndrome (WHS), a well-known contiguous microdeletion syndrome, is caused by deletions on chromosome 4p. There have been many related studies on it. The clinical symptoms and the critical region for this clinical disorder has been narrowed based on the genotype-phenotype correlations. However, duplications in this region have been reported infrequently. Case presentation: We report on a family showing a set of dysmorphic facial features, attention deficit hyperactivity disorders, learning difficulties, speech and cognitive delays, overgrowth/developmental delay, and musculoskeletal anomalies. Through karyotype analysis and chromosome microarray analysis combined with PCR Results, we found that patients in this family had translocations on chromosomes 2q37 and 4p16. Patients with 2q duplication and 4p deletion showed typical clinical phenotype of WHS syndrome, and family members with 2q deletion and 4p duplication also had obvious clinical phenotype. Conclusions: Our case report shows that not only deletions but also duplications of the Wolf-Hirshhorn critical region cause mental retardation and multiple congenital anomalies.

Background: FERM domain-containing protein 4A (FRMD4A) is a scaffolding protein previously proposed to be critical in the regulation of cell polarity in neurons and implicated in human intellectual development.Case Presentation: We report a case of a 3-year-old boy with corpus callosum anomaly, relative macrocephaly, ataxia, and unexplained global developmental delay. Here, compound heterozygous missense mutations in the FRMD4A gene [c.1830G>A, p.(Met610Ile) and c.2973G>C, p.(Gln991His)] were identified in the proband, and subsequent familial segregation showed that each parent had transmitted a mutation.Conclusions: Our results have confirmed the associations of mutations in the FRMD4A gene with intellectual development and indicated that for patients with unexplained global developmental delay, the FRMD4A gene should be included in the analysis of whole exome sequencing data, which can contribute to the identification of more patients affected by this severe phenotypic spectrum.

A full-term female baby born with an Apgar score of 10 exhibited a continual decrease in blood glucose and acid-base imbalance until she died at 40 h postpartum despite emergency rescue efforts. Autopsy showed a highly oedematous brain and a swollen liver. The cells of the myocardium, liver and kidney exhibited extensive microscopic vacuolar degeneration. The cardiac muscle and liver and kidney tissues were positive for Sudan III staining. Tandem mass spectrometry analysis revealed that the deceased was deficient in free carnitine, accompanied by an increase in multiple acylcarnitines. Genetic testing showed the parents of the deceased to be ETFDH mutation carriers, and the deceased carried a complex heterozygous mutation in ETFDH. Therefore, based on the results of autopsy, specific staining, genetic metabolic disease testing and genetic testing, we speculated that glutaric aciduria type IIc was the cause of death.

Background: Osteogenesis imperfecta (OI) is a clinical and genetic disorder that results in bone fragility, blue sclerae and dentineogenesis imperfecta (DGI), which is mainly caused by a mutation in the COL1A1 or COL1A2 genes, which encode type I procollagen.Case Report: A missense mutation (c.1463G > C) in exon 22 of the COL1A1 gene was found using whole-exome sequencing. However, the cases reported herein only exhibited a clinical DGI-I phenotype. There were no cases of bone disease or any other common abnormal symptom caused by a COL1A1 mutation. In addition, the ultrastructural analysis of the tooth affected with non-syndromic DGI-I showed that the abnormal dentine was accompanied by the disruption of odontoblast polarization, a reduced number of odontoblasts, a reduction in hardness and elasticity, and the loss of dentinal tubules, suggesting a severe developmental disorder. We also investigated the odontoblast differentiation ability using dental pulp stem cells (DPSCs) that were isolated from a patient with DGI-I and cultured. Stem cells isolated from patients with DGI-I are important to elucidate their pathogenesis and underlying mechanisms to develop regenerative therapies.Conclusion: This study can provide new insights into the phenotype-genotype association in collagen-associated diseases and improve the clinical diagnosis of OI/DGI-I.

Our previous studies have revealed that a dominant mutation in vacuolar protein sorting 4B (VPS4B), a member of the AAA ATPase family, causes dentin dysplasia type I. The purpose of the present study was to investigate the roles of VPS4B in human dental pulp stem cells (hDPSCs) and to elucidate the underlying molecular mechanisms. In this study, we found that VPS4B was highly expressed in the dental pulp cells of the mouse molar tooth germ, and the expression of VPS4B increased significantly during the odontoblastic differentiation of hDPSCs. VPS4B downregulation inhibited the proliferation, migration, and odontoblastic differentiation of hDPSCs. Moreover, treatment with lithium chloride, an agonist of the Wnt-β-catenin signalling pathway, partially reversed the VPS4B knockdown-driven suppression of proliferation and of odontoblastic differentiation of hDPSCs. Collectively, our findings indicate that VPS4B, via Wnt-β-catenin signalling, acts as a regulator of the proliferation and differentiation of hDPSCs. Our results suggest potential therapeutic avenues for dentin formation and regenerative endodontics in patients with dentin dysplasia type I.

Purpose Accumulating evidence indicated that triple‐negative breast cancer (TNBC) can stimulate stronger immune responses than other subtypes of breast cancer. We hypothesized that integrating immune‐related genomic signatures with clinicopathologic factors may yield a predictive accuracy exceeding that of the currently available system. Methods Ten signatures that reflect specific immunogenic or immune microenvironmental features of TNBC were identified and re‐analyzed using bioinformatic methods. Then, clinically annotated TNBC (n = 711) with the corresponding expression profiles, which predicted a patient's probability of disease‐free survival (DFS) and overall survival (OS), was pooled to evaluate their prognostic values and establish a clinicopathologic‐genomic nomogram. Three and two immune features were, respectively, selected out of 10 immune features to construct nomogram for DFS and OS prediction based on multivariate backward stepwise Cox regression analyses. Results By integrating the above immune expression signatures with prognostic clinicopathologic features, clinicopathologic‐genomic nomograms were cautiously constructed, which showed reasonable prediction accuracies (DFS: HR, 1.79; 95% CI, 1.46‐2.18, P

Hongsen Zhang, Fu Xiong, Kun Qian, Yiming Liu, Bin Liang, Bin Xiong, Fan Yang, Chuansheng Zheng Department of Radiology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China Purpose: Transcatheter arterial embolization or transcatheter arterial chemoembolization has become a critical therapy for unresectable hepatocarcinoma. Although hypoxia caused by embolization can induce apoptosis and necrosis of the majority of tumor cells, a small proportion of cells can survive with hypoxia and chemotherapy resistance. HIF-1α induced by hypoxia is the key factor rendering surviving tumor cells invasive and metastatic properties. Thus, we generated a synthetic hypoxia-replicative oncolytic adenovirus (HYAD) expecting to further eliminate the surviving tumor cells, which expressed HIF-1α.Materials and methods: In our study, we detected protein expression, proliferation, apoptosis, and necrosis of hepatic tumor cell line when infected with HYAD under hypoxia and normoxia. And we constructed VX2 hepatic cancer rabbit models to explore the therapeutic effect of transcatheter arterial embolization combined with HYAD perfusion under digital subtraction angiography. Inhibition of tumor growth and invasion was detected by histopathological examination and contrast-enhanced CT scan.Results: Experiments in vitro verified that HYAD expressed and replicated along with HIF-1α expression or hypoxia. Compared with wild adenovirus type 5 (WT), HYAD expressed much more under hypoxia, which was the main principle of HYAD killing surviving tumor cells posttransarterial embolization. In vivo experiment of VX2 models, HYAD perfusion combined with polyvinyl alcohol (PVA) embolization achieved the highest expression quantity and the longest expression duration compared with simple HYAD perfusion, WT perfusion combined with PVA embolization, and simple WT perfusion. Because adenovirus expression protein E1A had the properties of promoting apoptosis, inhibiting invasion, and inhibiting metastasis, HYAD perfusion combined with PVA embolization group efficiently repressed tumor growth and intrahepatic metastases compared to other processing groups.Conclusion: HYAD can overcome the hypoxic tumor microenvironment postembolization and target the surviving tumor cells with specificity. In turn, HYAD perfusion combined with PVA embolization can bring out the best effect in each other. Keywords: transcatheter arterial embolization, hepatocarcinoma, hypoxia, oncolytic adenovirus

Background：Osteogenesis imperfecta (OI) is a genetic disorder characterized by bone fragility, blue sclerae and dentinogenesis imperfecta (DGI), which are mainly caused by a mutation of the COL1A1 or COL1A2 genes that encode type I procollagen.Methods: The ultrastructure of dentin was analyzed by micro-CT, scanning electron microscopy, energy-dispersive spectroscopy analysis, nanoindentation test and Toluidine Blue Staining. Whole-exome sequencing (WES) was performed to identify the pathogenic gene. The function of the mutant COL1A1 was studied by real-time PCR, western blotting, subcellular localization. Functional analysis in dental pulp stem cells (DPSCs) was also performed to explore the impact of the identified mutation on this phenotype. Results: WES identified a missense mutation (c.1463G > C) in exon 22 of the COL1A1 gene. However, the cases reported herein only exhibited DGI-I in the clinical phenotype, there is no bone disease and any other common abnormal symptom caused by COL1A1 mutation. In addition, ultrastructural analysis of the tooth affected with non-syndromic DGI-I showed that the abnormal dentin was accompanied by disruption of odontoblast polarization, reduced numbers of odontoblasts, loss of dentinal tubules, and reduction in hardness and elasticity, suggesting severe developmental disturbance. What’s more, the odontoblast differentiation ability based on DPSCs that were isolated and cultured from the DGI-I patient was enhanced compared with those from an age-matched, healthy control.Conclusion: This study helped the family members to understand the disease progression and provided new insights into the phenotype-genotype association in collagen-associated diseases and improve clinical diagnosis of OI/DGI-I.

VPS4B (vacuolar protein sorting 4B), a member of the ATPase associated with diverse cellular activities (AAA) protein family, is a component of the endosomal sorting complexes required for transport machinery which regulates the internalization and lysosomal degradation of membrane proteins. We previously reported that VPS4B is one of the pathogenic genes related to dentin dysplasia type I, although its function was largely unknown. To investigate the role of VPS4B in tooth development, we deleted the Vps4b gene in mice. We found that heterozygous knockout mice (Vps4b+/- ) developed normally and were fertile. However, homozygous deletion of the Vps4b gene resulted in early embryonic lethality of Vps4b-/- mice at approximately embryonic day 9.5 (E9.5). To investigate the underlying molecular mechanisms, we examined the molecular functions of VPS4B in vivo and in vitro. Cell experiments showed that VPS4B influenced the proliferation, apoptosis, and cell cycle of transfected human neuroblastoma cells (IMR-32 cells) with over-expression or knockdown of VPS4B. Moreover, qRT-PCR detection showed that the mRNA expression levels of apoptosis-, cell cycle-, and endocytosis-related genes was significantly down or up-regulated in RNA interference-mediated knockdown of VPS4B in IMR-32 cells and Vps4b+/- E12.5 embryos. We accordingly speculated that signal transduction disorders of cell endocytosis are a contributing factor to the prenatal lethality of Vps4b-/- mice.

To investigate the efficacy of sequential apatinib treatment in patients with unresectable hepatocellular carcinoma (HCC) refractory to sorafenib–transarterial chemoembolization (TACE). This respective study was conducted on 95 consecutive patients with sorafenib–TACE-refractory unresectable HCC in our center from January 2014 to December 2019. According to the response to sorafenib–TACE treatment and the subsequent management, the eligible patients were assigned into three groups: sorafenib–TACE (ST) group, sorafenib–TACE–apatinib (STA) group, and sorafenib–TACE–supportive care (STS) group. The differences in overall survival (OS) and tumor response were compared among the three groups. Risk factors related to prognosis were analyzed. A total of 58 patients were enrolled in the study, with 11, 15, and 28 patients in ST, STA, and STS group, respectively. The median OS of the STA group was significantly improved when compared with the STS group, either from the start of sorafenib–TACE resistance or the initiation of sorafenib–TACE therapy (14.0 versus 4.0 months, p = 0.003; 19.0 versus 9.0 months, p

Background The ABO blood group system is clinically important in kidney transplantation, but ABO genotyping fails to attract sufficient attention in some countries and regions. We identified one case of early graft dysfunction due to an ABO genotype mismatch. Here, we performed ABO genotyping in blood samples, analyzed grouping discrepancies, and investigated the weak A subgroup frequency in kidney transplantation candidates. Methods Blood samples from 302 uremic patients with grouping discrepancies and 356 uremic patients with type A blood were analyzed using standard serologic serotyping techniques. The ABO genotypes and alleles were analyzed by polymerase chain reaction sequence-specific primer (PCR-SSP) and sequence-based typing (PCR-SBT). Results All 302 uremic patients with grouping discrepancies carried weak ABO subgroup alleles and 77.48% carried irregular ABO antibodies. The discrepancy rate between serotyping and genotyping was 42.38%, and the mismatching rate of donor selection according to serotype reached 88.74%. And 2.53% of 356 uremic patients with type A blood were determined to be in the weak A subgroup, which was a higher percentage than that observed in the healthy Chinese population (0.53%) by serological screening, but much lower than that observed in Caucasians (20%). Conclusion We revealed the high risk of blood type misjudgment and genetically ABO-mismatched transplantation if serological test was performed only in blood-group typing. Improved precision of ABO genotyping is crucial for successful kidney transplantation and reasonable organ allocation.

Hypophosphatasia (HPP) is a rare hereditary systemic disease that is characterized by defective bone and/or dental mineralization, and is caused by mutations in the alkaline phosphatase gene (ALPL). The present study investigated the ALPL mutation in a Chinese Han family with HPP and studied the pathogenesis of the mutations of the ALPL gene. DNA was extracted from peripheral venous blood of the family members. Sanger sequencing was used to screen the mutations. Associations between pathogenesis for both mutations were analyzed by bioinformatics, subcellular localization, measurement of enzyme activity and western blotting. Sanger sequencing revealed the compound heterozygous mutations c.203C>T (p.T68M) and c.571G>A (p.E191K). The mutations were located at exon 4 and 6 of the ALPL gene and were predicted by Polyphen-2 analysis to be harmful. Protein analysis indicated a decrease in mature protein production and lower enzyme activity in 293T cells transfected with plasmids carrying the mutations. The ALPL gene was cloned into the pcDNA3.1(+) vector and mutant plasmids ALPL-pT68M and ALPL-pE191K were constructed. Immunofluorescence observed in cells transfected with the ALPL-pE191K mutant plasmid was mainly located in the cell membrane. However, staining in the cytoplasm was increased compared with the wild type, and almost no fluorescence was identified in 293T cells transfected with the ALPL-pT68M mutant plasmid. The present findings demonstrated that the compound heterozygous c.571G>A and c.203C>T mutations may contribute to childhood HPP by resulting in mislocalization, decreased protein expression and loss of enzyme activity in a Han Chinese family. The results of the current study may provide insights into the potential molecular mechanism of HPP.

A splicing mutation in VPS4B can cause dentin dysplasia type I (DD-I), a hereditary autosomal-dominant disorder characterized by rootless teeth, the etiology of which is genetically heterogeneous. In our study, dental follicle cells (DFCs) were isolated and cultured from a patient with DD-I and compared with those from an age-matched, healthy control. In a previous study, this DD-I patient was confirmed to have a loss-of-function splicing mutation in VPS4B (IVS7 + 46C > G). The results from this study showed that the isolated DFCs were vimentin-positive and CK14-negative, indicating that the isolated cells were derived from the mesenchyme. DFCs harboring the VPS4B mutation had a significantly higher proliferation rate from day 3 to day 8 than control DFCs, indicating that VPS4B is involved in cell proliferation. The cells were then replenished with osteogenic medium to investigate how the VPS4B mutation affected osteogenic differentiation. Induction of osteogenesis, detected by alizarin red and alkaline phosphatase staining in vitro, was decreased in the DFCs from the DD-I patient compared to the control DFCs. Furthermore, we also found that the VPS4B mutation in the DD-I patient downregulated the expression of osteoblast-related genes, such as ALP, BSP, OCN, RUNX2, and their encoded proteins. These outcomes confirmed that the DD-I-associated VPS4B mutation could decrease the capacity of DFCs to differentiate during the mineralization process and may also impair physiological root formation and bone remodeling. This might provide valuable insights and implications for exploring the pathological mechanisms underlying DD-I root development.

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are both caused by mutations in DMD gene effecting the expression of dystrophin. Generally female carriers are asymptomatic; however, it has been suggested that carriers may exhibit symptoms. We investigated a 6-year-old Chinese girl exhibiting a suspected BMD phenotype, including persistently elevated creatine kinase and creatine kinase isoenzyme levels. The proband harbored a novel heterozygous mutation, c.3458_3459insAA, within exon 26 of the DMD gene inherited from her mother who had a completely normal phenotype and presented with mosaicism in her lymphocytes with 45, X [17%]/46, XX [83%]. In addition, X-chromosome inactivation (XCI) patterns in the peripheral blood of the child were slightly skewed: proband with 62% (mutant allele)/38% (normal allele) when compared with her mother with 32/68%. Amplification of regions of the cDNA revealed different ratios for the expression of these alleles: proband with 50/50% and her mother with 20/80%. Real-time PCR showed that mRNA expression was significantly decreased in both. We proposed that a frameshift or nonsense mutation may contribute to the development of symptoms in carriers. These phenotypes correlate with nonrandom XCI patterns and are compounded by the locus of the mutation. For incompletely skewed XCI patterns, although the mutant allele could suppress the expression of a normal allele, carriers would remain asymptomatic as long as there was adequate compensation from the normal allele. We also proposed a mechanism where mRNA from the mutant allele may be unstable and easily degraded, allowing for phenotypic compensation by the wildtype allele.

BackgroundChest computed tomography (CT) has been widely used to assess pulmonary involvement in COVID-19. We aimed to investigate the correlation between chest CT and clinical features in COVID-19 suspected patients with or without fever.MethodsWe retrospectively enrolled 211 COVID-19 suspected patients who underwent both chest CT and reverse transcription polymerase chain reaction in Wuhan, China. The performance of CT in patients with relevant onset of symptoms, with fever (n = 141) and without fever (n = 70), was assessed respectively.ResultsThe sensitivity of CT for COVID-19 was 97.3%, with area under the curve (AUC) of 0.71 (95% confidence interval [CI], 0.66–0.76). There were 141 suspected patients with fever and 70 without fever. In the fever group, 4 variables were screened to establish the basic model: age, monocyte, red blood cell, and hypertension. The AUC of the basic model was 0.72 (95% CI, 0.63–0.81), while the AUC of the CT-aided model was 0.77 (95% CI, 0.68–0.85), a significant difference (P < .05). In the nonfever group, only dry cough was screened out to establish the basic model. The AUC was 0.76 (95% CI, 0.64–0.88), which was not significantly different than the CT-aided model (P = .08).ConclusionsChest CT has a high sensitivity in patients with COVID-19, and it can improve diagnostic accuracy for COVID-19 suspected patients with fever during the initial screen, whereas its value for nonfever patients remains questionable.

Charcot–Marie–Tooth (CMT) disease is the most common inherited peripheral neuropathy and shows clinical and genetic heterogeneity. Mutations in C1orf194 encoding a Ca2+ regulator in neurons and Schwann cells have been reported previously by us to cause CMT disease. In here, we further investigated the function and pathogenic mechanism of C1or194 by generating C1orf194 knockout (KO) mice. Homozygous mutants of C1orf194 mice exhibited incomplete embryonic lethality, characterized by differentiation abnormalities and stillbirth on embryonic days 7.5–15.5. Heterozygous and surviving homozygous C1orf194 KO mice developed motor and sensory defects at the age of 4 months. Electrophysiologic recordings showed decreased compound muscle action potential and motor nerve conduction velocity in the sciatic nerve of C1orf194-deficient mice as a pathologic feature of dominant intermediate-type CMT. Transmission electron microscopy analysis revealed demyelination and axonal atrophy in the sciatic nerve as well as swelling and loss of mitochondrial matrix and other abnormalities in axons and Schwann cells. A histopathologic examination showed a loss of motor neurons in the anterior horn of the spinal cord and muscle atrophy. Shorter internodal length between nodes of Ranvier and Schmidt–Lanterman incisures was detected in the sciatic nerve of affected animals. These results indicate that C1orf194 KO mice can serve as an animal model of CMT with a severe dominant intermediate CMT phenotype that can be used to investigate the molecular mechanisms of the disease and evaluate the efficacy of therapeutic strategies.

To evaluate the predictive value of preoperative biochemical marker [platelet-to-lymphocyte ratio (PLR)] in patients with advanced hepatocellular cancer receiving transarterial chemoembolization (TACE) plus targeted molecular therapy (apatinib) treatment. Clinical records of 134 patients receiving the treatment of TACE + apatinib (TACE-A) and the treatment of TACE alone were compared in a single-center study. Time to progression (TTP) and overall survival (OS) were compared between TACE-A and TACE alone groups in patients with PLR 150 and PLR ≤ 150, respectively. The area under the receiver operating characteristic (ROC) curve was used to determine the prediction power of PLR. The median TTP and OS in the TACE-A group were significantly longer than those in the TACE alone group (P 0.001). The median TTP and OS in the TACE-A (PLR ≤ 150) group were longer than those in the TACE-A (PLR 150) group (P 0.05). There was no significant difference between TACE-A (PLR 150) and TACE alone (P = 0.232) groups in OS, but the median TTP in the TACE-A (PLR 150) group was longer than that in the TACE alone group (P = 0.001). ROC analysis showed that the area under the curve was 0.643 and 0.623 for 6- and 12-month survival, respectively. PLR might predict the results of patients with advanced hepatocellular carcinoma received TACE-A treatment.

Low-level BK polyomavirus (BKPyV) shedding is seen in at least 10% of seropositive immunocompetent adults. Moreover, BKPyV infection is highly prevalent amongst immunocompromised populations, yet little is known on its relationship with malignancy. We studied a female patient with BKPyV-associated and donor-derived de novo high-grade sarcomatoid urothelial carcinoma developed 8 years after kidney transplantation from a male donor. Through whole-genome sequencing, we discovered integration of genotype IV BKPyV genome into the non-coding RNA (ncRNA) intronic region of human chromosome 18. The two breakpoints in the virus genome were located at the non-coding control region (NCCR) and large T antigen (TAg) coding region, respectively. Nevertheless, the TAg was overexpressed. We, therefore, inferred that the BKPyV was clonally integrated into the human genome in the form of concatemers, facilitating the expression of the TAg. The patient presented with multiorgan metastases, which were reduced in size and number throughout the body after removal of the graft and cessation of immunosuppressants. The few remaining lesions located in the liver were identified, through biopsy to be necrotic tumor tissue with TAg detected; additionally, genomic sequencing of the liver mass found Y chromosome. In conclusion, we propose that integration of the BKPyV genome is closely related to oncogenesis in this patient; while oncogenesis occurred when host immunity was impaired, recovery of the patient’s native immunity effectively curbed viral replication and eliminated the metastatic lesions.

PURPOSE: To validate the feasibility and efficacy of intratumoral radiofrequency hyperthermia (RFH)-enhanced herpes simplex virus (HSV) thymidine kinase (TK) and ganciclovir (GCV) (hereafter, HSV-TK/GCV) gene therapy for non–small-cell lung cancer (NSCLC). MATERIALS AND METHODS: This study was performed from November 11, 2015, to April 14, 2017, and included (a) in vitro experiments with human NSCLC cells to establish the proof of principle, (b) in vivo experiments using mice with subcutaneous NSCLC to further demonstrate the principle, and (c) in vivo experiments using rats with orthotopic NSCLC to validate the technical feasibility. Cells, nude mice, and nude rats were randomly divided into four groups (six animals per group): (a) combination therapy (HSV-TK/GCV combined with RFH), (b) RFH, (c) HSV-TK/GCV, and (d) phosphate-buffered saline. Data were analyzed by using the Dunnett t test or Kruskal–Wallis test. RESULTS: For in vitro experiments, the cell proliferation assay showed significantly diminished viable cells with combination therapy (mean, 0.56; 95% confidence interval [CI]: 0.44, 0.68) versus RFH (mean, 0.89; 95% CI: 0.82, 0.97), HSV-TK/GCV (mean, 0.71; 95% CI: 0.56, 0.86), and phosphate-buffered saline (mean, 1; 95% CI: 1, 1) (P < .05 for all). For in vivo experiments, optical imaging showed significantly decreased relative bioluminescence signal with combination therapy (mean, 0.71 [95% CI: 0.03, 1.39] in mice; 1.29 [95% CI: 0.51, 2.06] in rats) compared with RFH (mean, 2.66 [95% CI: 1.73, 3.59] in mice; 2.26 [95% CI: 1.51, 3.01] in rats), HSV-TK/GCV (mean, 1.37 [95% CI: 0.65, 2.08] in mice; 1.76 [95% CI: 1.20, 2.31] in rats), and phosphate-buffered saline (mean, 3.07 [95% CI: 2.50, 3.65] in mice; 2.94 [95% CI: 2.29, 3.58] in rats) (P < .001 for all). US showed that the smallest relative tumor volumes occurred with combination therapy (mean, 0.60; 95% CI: 0.15, 1.05) versus RFH (mean, 2.43; 95% CI: 1.80, 3.06), HSV-TK/GCV (mean, 1.32; 95% CI: 0.75, 1.89), and phosphate-buffered saline (mean, 2.56; 95% CI: 1.75, 3.38) (P < .05 for all) in the mouse subcutaneous model. CONCLUSION: Intratumoral radiofrequency hyperthermia–enhanced herpes simplex virus thymidine kinase and ganciclovir gene therapy for non–small-cell lung cancer is feasible and can be guided by molecular imaging. © RSNA, 2018

// Fu Xiong 1, 2, * , Feng Zhang 2, * , Yin Jin 2 , Qiaoyou Weng 2 , Jingjing Song 2 , Guofeng Zhou 1 , David Shin 2 , Chuansheng Zheng 1 and Xiaoming Yang 2 1 Department of Radiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province 430022, China 2 Image-Guided Bio-Molecular Intervention Research and Section of Vascular & Interventional Radiology, Department of Radiology, University of Washington School of Medicine, Seattle, WA 98109, USA * These authors have contributed equally to this work Correspondence to: Xiaoming Yang, email: xmyang@uw.edu Chuansheng Zheng, email: hqzcsxh@sina.com Keywords: radiofrequency hyperthermia; gene therapy; hepatocellular carcinoma; molecular imaging Received: July 04, 2017 Accepted: September 08, 2017 Epub: December 22, 2017 Published: March 06, 2018 ABSTRACT Purpose: To validate the feasibility of using interventional radiofrequency hyperthermia(RFH) to enhance herpes simplex virus-thymidine kinase (HSV-TK)/ganciclovir (GCV) gene therapy of rat orthotopic hepatic cancer. Material and Methods : Rat hepatocellular carcinoma cells (MCA-RH-7777) were transduced with lentivirus/luciferase gene for optical imaging. In-vitro experiments with the luciferase cells and in-vivo experiments on rats with orthotopic hepatic tumors were divided into four treatment groups: (i) HSV-TK/GCV-mediated gene therapy combined with RFH; (ii) gene therapy alone; (iii) RFH alone; and (iv) phosphate buffered saline (PBS). Cell viability was evaluated by MTS assay and confocal microscopy, and HSV-TK gene expression in cells and tumors was quantified by western blotting. Bioluminescent optical imaging and ultrasound imaging were used to monitor and compare the photon signal and tumor size changes among different treatment groups overtime, respectively. The imaging findings were correlated with histology. Results: For in-vitro experiments, the combination therapy group (gene therapy + RFH) demonstrated the lowest cell proliferation by MTS assay, compared to the gene therapy alone, RFH alone, and PBS (26.1±3.2% vs 50.4±4.6% vs 82.9±6.3% vs 100%, p

Fu Xiong,1,2,* Yanyan Cao,1,2,* Xiaopeng Guo,1,2 Hongsen Zhang,1,2 Jihua Wang,1,2 Bin Xiong,1,2 Bin Liang,1,2 Chuansheng Zheng1,2 1Department of Radiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, People’s Republic of China; 2Hubei Province Key Laboratory of Molecular Imaging, Wuhan, Hubei 430022, People’s Republic of China*These authors contributed equally to this workCorrespondence: Chuansheng Zheng Email hqzcsxh@sina.comPurpose: To evaluate the adriamycin (ADM) pervasion distance within tumor stroma after relaxin (RLX) infusion through tumor feeding artery and further investigate the therapeutic effects of RLX infusion combined with transcatheter chemoembolization (TACE) on the rabbit VX2 liver cancer, since the chemotherapy impaired due to limited drug distribution hindered by stiffened tumor stroma.Materials and Methods: In the first part, rabbits received normal saline (NS), RLX or combined with TACE, and the penetration distance of ADM was measured by immunofluorescence and the matrix metalloproteinases (MMPs) were evaluated by gelatin substrate zymography in each group. In the second part, the rabbits received NS, TACE and RLX combined with TACE, respectively. The tumor growth rates, necrosis rates and intrahepatic metastasis were measured, andhematoxylin-eosin (HE), transferase-mediated dUTP-biotin nick end labelling (TUNEL) and Ki67 staining were conducted in each group.Results: In the first part, the expression of MMP-9 was increased in groups treated by RLX compared with NS group, especially three days after RLX infusion (p=0.001). The ADM penetration distance was significantly increased in groups treated by RLX compared with NS group (p< 0.05), and it was farthest three days after RLX infusion. In the second part, compared with the NS and TACE groups, the tumor growth rates, the positive staining rates of Ki67 and the tumor growth rates were significantly decreased in RLX+TACE group (p< 0.05). However, the positive staining rates of TUNEL and the tumor necrosis rates were significantly increased (p< 0.05), andHE staining also revealed higher necrosis rates. The intrahepatic metastasis indicates no difference between the three groups (p=0.273).Conclusion: An increased penetration distance was obtained by RLX infusion through tumor feeding artery, and better therapeutic effects were achieved by RLX combined with TACE.Keywords: relaxin, liver cancer, transcatheter chemoembolization, MMP-9, penetration distance

Methicillin‑resistant Staphylococcus aureus (S. aureus; MRSA) is one of the most common bacterial pathogens and MRSA infections are characterized by high mortality rates. Antimicrobial peptides are considered one of the most promising drugs for the treatment of resistant strains of S. aureus. The present study aimed to examine the antimicrobial activity of L12 against numerous bacterial species using the broth microdilution method. Furthermore, the synergistic effect of L12 combined with various antibacterial drugs was tested, and its antibacterial mechanism was investigated by a checkerboard assay. The alterations in bacterial morphology were detected by electron microscopy, and biofilm formation and removal were tested by crystal violet staining. The present results suggested that L12 affected the growth of gram‑positive strains, particularly S. aureus. Electron microscopy analysis suggested that L12 may target the cell membrane, and L12 increased the antibacterial activity of vancomycin and levofloxacin, exerting a synergistic effect. However, the minimal inhibitory concentrations (MICs) of L12 were not correlated with antibiotic resistance, the strains resistant to more antibiotics were not more resistant to L12. A sub‑MIC of L12 was able to inhibit biofilm formation in a dose‑dependent manner; however, concentrations of L12 ≤10 times the MIC were not sufficient to degrade previously formed biofilm. Collectively, the present study suggested that L12 may represent a novel potential therapeutic molecule for the treatment of S. aureus infections.

Background Vacuolar protein sorting-associated protein 4B (VPS4B) is a member of the ATP enzyme AAA protein family, and is mainly involved in protein degradation and cell membrane fusion. Recently, a dominant mutation in this gene was identified in human dentin dysplasia type I (DD-I). Herein, we report the generation of Vps4b knockout (Vps4b KO) mice; however, the homozygous Vps4b KO mutation was embryonic lethal at the early stages of embryo development, and we therefore report the results of heterozygous mutant mice. Results Mice heterozygous for Vps4b did not develop tooth defects replicating human DD-I. Immunohistochemistry showed that gene KO was successful, as there was decreased expression of Vps4b in heterozygous mice; hematoxylin and eosin (H&E) staining also showed that the width of the pre-dentin zone was increased in heterozygous mice, although the arrangement of the odontoblasts was not significantly different from wild-type (WT) mice. However, H&E staining showed no obvious abnormalities in the bones of heterozygous mice. Moreover, stereomicroscopic and X-ray radiography results indicated no abnormal manifestations in teeth or bones. Furthermore, statistical analysis of the volume and density of dentin and enamel, as well as skeletal analysis, including the volume and separation of trabecular bone analyzed by micro-CT, all showed no differences between Vps4b heterozygotes and WT mice. In addition, there also were no significant differences in bone or cartilage mineralization as evaluated by Alcian blue–Alizarin red staining. Conclusions The heterozygous Vps4b KO mice do not develop tooth defects that replicate human DD-I and this is likely to be due to differences in tooth development between the two species. Consequently, further studies are needed to determine whether mice are an appropriate animal model for human tooth diseases.

Charcot-Marie-Tooth disease is a hereditary motor and sensory neuropathy exhibiting great clinical and genetic heterogeneity. Here, the identification of two heterozygous missense mutations in the C1orf194 gene at 1p21.2-p13.2 with Charcot-Marie-Tooth disease are reported. Specifically, the p.I122N mutation was the cause of an intermediate form of Charcot-Marie-Tooth disease, and the p.K28I missense mutation predominately led to the demyelinating form. Functional studies demonstrated that the p.K28I variant significantly reduced expression of the protein, but the p.I122N variant increased. In addition, the p.I122N mutant protein exhibited the aggregation in neuroblastoma cell lines and the patient’s peroneal nerve. Either gain-of-function or partial loss-of-function mutations to C1ORF194 can specify different causal mechanisms responsible for Charcot-Marie-Tooth disease with a wide range of clinical severity. Moreover, a knock-in mouse model confirmed that the C1orf194 missense mutation p.I121N led to impairments in motor and neuromuscular functions, and aberrant myelination and axonal phenotypes. The loss of normal C1ORF194 protein altered intracellular Ca2+ homeostasis and upregulated Ca2+ handling regulatory proteins. These findings describe a novel protein with vital functions in peripheral nervous systems and broaden the causes of Charcot-Marie-Tooth disease, which open new avenues for the diagnosis and treatment of related neuropathies.

A novel polysaccharide (MFP1P) was isolated from Fructus Mori, followed by purification via DEAE-52 cellulose and 27 % ethanol fraction. The MFP1P had the molecular weight of 56.78 kDa and the total sugar content of 93.32±0.54 %. And the MFP1P is mainly composed of glucose, galactose, galacturonic acid and mannose with molar ratio of 66.62 %, 13.94 %, 18.24 % and 1.20 %, respectively. MFP1P was mainly composed of →3)-α-D-Gal (1→, β-D-Man-(1→ and →6)-α-D-Glc (1→ glycosidic bond and showed a spherical chain conformation with uniform distribution in solution. The MFP1P exhibited great antioxidant activity with oxygen-free radical absorption capacity (ORAC) values of 291.63±6.81 μmol TE/g and MDA IC50 of 0.289±0.022 mg/mL.

Dentin dysplasia type I (DDI) is an autosomal-dominant genetic disorder resulting from dentin defects. The molecular basis of DDI remains unclear. DDI exhibits unique characteristics with phenotypes featuring obliteration of pulp chambers and diminutive root, thus providing a useful model for understanding the genetics of tooth formation. Using a large Chinese family with 14 DDI patients, we mapped the gene locus responsible for DDI to 3p26.1-3p24.3 and further identified a missense mutation, c.353C>A (p.P118Q) in the SSUH2 gene on 3p26.1, which co-segregated with DDI. We showed that SSUH2 (p.P118Q) perturbed the structure and significantly reduced levels of mutant (MT) protein and mRNA compared with wild-type SSUH2. Furthermore, MT P141Q knock-in mice (+/- and -/-) had a unique partial obliteration of the pulp cavity and upregulation or downregulation of six major genes involved in odontogenesis: Dspp, Dmp1, Runx2, Pax9, Bmp2, and Dlx2. The phenotype of missing teeth was determined in zebrafish with morpholino gene knockdowns and rescued by injection of normal human mRNA. Taken together, our observations demonstrate that SSUH2 disrupts dental formation and that this novel gene, together with other odontogenesis genes, is involved in tooth development.

Duchenne muscular dystrophy (DMD) is an X-linked genetic muscular disorder with no effective treatment at present. Mesenchymal stem cell (MSC) transplantation has been used to treat DMD, but the efficiency is low. Our previous studies show that activation of Wnt3a signaling promotes myogenic differentiation of MSCs in vitro. Here we report an effective MSC transplantation therapy in mdx mice by activation of Wnt3a signaling. MSCs were isolated from mouse bone marrow, and pretreated with Wnt3a-conditioned medium (Wnt3a-CM), then transplanted into mdx mice. The recipient mice were euthanized at 4, 8, 12, 16 weeks after the transplantation, and muscle pathological changes were examined. The expression of dystrophin in muscle was detected using immunofluorescence staining, RT-PCR and Western blotting. Sixteen weeks later, transplantation of Wnt3a-pretreated MSCs in mdx mice improved the characteristics of dystrophic muscles evidenced by significant reductions in centrally nucleated myofibers, the variability range of cross-sectional area (CSA) and the connective tissue area of myofibers. Furthermore, transplantation of Wnt3a-pretreated MSCs in mdx mice gradually and markedly increased the expression of dystrophin in muscle, and improved the efficiency of myogenic differentiation. Transplantation of Wnt3a-pretreated MSCs in mdx mice results in long-term amelioration of the dystrophic phenotype and restores dystrophin expression in muscle. The results suggest that Wnt3a may be a promising candidate for the treatment of DMD.

Limited predictable long noncoding RNA (lncRNA) signature was reported in tamoxifen resistance among estrogen receptor (ER)-positive breast cancer (BC) patients. The aim of this study was to identify and assess prognostic lncRNA signature to predict recurrence among ER-positive BC patients treated with tamoxifen. Cohorts from Gene Expression Omnibus (GEO) (n = 298) and The Cancer Genome Atlas (TCGA) (n = 160) were defined as training and validation cohort, respectively. BC relapse associated lnRNAs was identify within training cohort, and the predictable value of recurrence was assessed in both cohorts. A total of 11lncRNAs were recognized to be associated with relapse free survival (RFS) of ER-positive BC patients receiving tamoxifen, who were divided into low-risk and high-risk group on basis of relapse risk scores (RRS). Multivariate cox regression analyses revealed that the RRS is an independent prognostic biomarker in the prediction of ER-positive BC patients’ survival. GSEA indicated that high-risk group was associated with several signaling pathways in processing of BC recurrence and metastasis such as PI3K-Akt and Wnt signaling. Our 11-lncRNA based classifier is a reliable prognostic and predictive tool for disease relapse in BC patients receiving tamoxifen.

We aimed to compare the overall survival (OS) and standardized mortality rate (SMR) of the male breast cancer (MBC) with female breast cancer (FBC) after propensity score matching. Based on the Surveillance, Epidemiology, and End Results (SEER), the early breast cancer patients (T1–2N0–2M0) were extracted from 1998–2007. This study included 1,111 and 2,151 patients with early MBC and FBC, respectively, whose clinicopathological characteristics were well balanced. At a mean follow-up time of 97 months, 10-year OS rate was 58.3% in the MBC group and 68.7% in the FBC (log-rank test, P

Objectives: To investigate the variations in the TNNI3 gene in a Chinese Han family affected by hypertrophic cardiomyopathy (HCM) and the potential molecular mechanism linking these mutations with disease. Methods: Peripheral venous blood was acquired from family members, and TNNI3 mutations were identified by DNA sequencing. The pathophysiology of TNNI3 mutations was investigated using bioinformatics, subcellular localization determination and Western blotting. Results: Sanger sequencing revealed that the proband possessed 2 heterozygous mutations, c.235C>T and c.470C>T, located at exons 4 and 6 of the TNNI3 gene. The proband (II-2) and her brother (II-1), who had been previously diagnosed with HCM, harbored both mutations whereas their healthy parents harbored only 1. Alignment of the TNNI3 amino acid sequence indicated that the two Pro residues were highly conserved across species. Subcellular localization showed that both wild-type (WT) and mutant TNNI3 proteins were localized at the cell nucleus. Western blot analysis of expression in human embryonic kidney 293T cells showed that the intracellular levels of the mutant proteins were significantly decreased compared to WT TNNI3 (p < 0.01). Conclusions: Our findings showed that a double heterozygous mutation in the TNNI3 gene is involved in the pathogenesis of HCM via haploinsufficiency. These results will inspire further studies to investigating the link between the TNNI3 gene and HCM.

Krüppel-like factor 1 (KLF1) regulates erythroid lineage commitment, globin switching, and the terminal maturation of red blood cells. Variants in human KLF1 have been identified as an important causative factor in a wide spectrum of phenotypes. This study investigated two unrelated male children in China who had refractory anemia associated with poikilocythemia. These were accompanied by an upregulation of biochemical markers of hemolysis, along with abnormal hemoglobin (Hb) level and elevated reticulocyte counts. Next-generation sequencing revealed that the patients were compound heterozygotes for a KLF1 frameshift mutation c.525_526insCGGCGCC (p.(Gly176ArgfsTer179)) and one of two missense variants, c.892 G>C (p.(Ala298Pro)) and c.1012C>T (p.(Pro338Ser)). The subjects had microcytic hypochromic anemia, and their healthy parents had single mutation. The two missense mutations affected a highly conserved codon in the zinc finger DNA-binding domain of KLF1, but the protein stability was unaffected in K-562 cells. A KLF1-targeted promoter–reporter assay showed that the two mutations reduce the expression of the HBB, BCL11A, and CD44 genes involved in erythropoiesis, with consequent dyserythropoiesis and an α/non-α chain imbalance. A systematic analysis was performed of the phenotypes associated with the KLF1 mutations in the two families, and the clinical characteristics and differential diagnoses of the disease are presented. This is the first report of an autosomal recessive anemia presenting with microcytic hypochromia, abnormal Hb profile, and other distinctive erythrocyte phenotypes, and provides insight into the multiple roles of KLF1 during erythropoiesis.

Hereditary dentin disorders include dentinogenesis imperfecta (DGI) and dentin dysplasia (DD), which are autosomal dominant diseases characterized by altered dentin structure such as abnormality in dentin mineralization and the absence of root dentin. Shields classified DGI into three subgroups and DD into two subtypes. Although they are all hereditary dentin diseases, they do not share the same causative genes. To date, the pathogenic genes of DGI type I, which is considered a clinical manifestation of syndrome osteogenesis imperfecta, include COL1A1 and COL1A2. Mutations of the DSPP gene, which encodes the dentin sialophosphoprotein, a major non-collagenous protein, are responsible for three isolated dentinal diseases: DGI-II, DGI-III, and DD-II. However, DD-I appears to be special in that researchers have found three pathogenicity genes-VPS4B, SSUH2, and SMOC2-in three affected families from different countries. It is believed that DD-I is a genetically heterogeneous disease and is distinguished from other types of dentin disorders. This review summarizes the DD-I literature in the context of clinical appearances, radiographic characteristics, and functions of its pathogenic genes and aims to serve clinicians in further understanding and diagnosing this disease.

Hemoglobinopathies are among the most common autosomal-recessive disorders worldwide. A comprehensive next-generation sequencing (NGS) test would greatly facilitate screening and diagnosis of these disorders. An NGS panel targeting the coding regions of hemoglobin genes and four modifier genes was designed. We validated the assay by using 2522 subjects affected with hemoglobinopathies and applied it to carrier testing in a cohort of 10,111 couples who were also screened through traditional methods. In the clinical genotyping analysis of 1182 β-thalassemia subjects, we identified a group of additional variants that can be used for accurate diagnosis. In the molecular screening analysis of the 10,111 couples, we detected 4180 individuals in total who carried 4840 mutant alleles, and identified 186 couples at risk of having affected offspring. 12.1% of the pathogenic or likely pathogenic variants identified by our NGS assay, which were undetectable by traditional methods. Compared with the traditional methods, our assay identified an additional at-risk 35 couples. We describe a comprehensive NGS-based test that offers advantages over the traditional screening/molecular testing methods. To our knowledge, this is among the first large-scale population study to systematically evaluate the application of an NGS technique in carrier screening and molecular diagnosis of hemoglobinopathies., Highlights • In 10,111 couples in prevention programs for thalassemia, 186 at-risk couples were identified by NGS and 151 by traditional routine method. • NGS assay had better performance than traditional routine screening approach in carrier screening for hemoglobinopathies. • NGS assay can characterize patients with Thalassemia Major or Thalassemia Intermedia more accurately. Hemoglobinopathies represent a major cause of birth defects in China. Traditional carrier screening strategy is phenotype-based that is associated with risk of miss phenotype-negative carriers. In this study, we validated an effective method for molecular screening and clinical genotyping of hemoglobinopathies using NGS assay. Considering the broad mutation spectrum and high prevalence of hemoglobinopathy-causing mutations across ethnic groups, this technology could make it technically feasible to offer more effective preconception screening and diagnosis in large populations worldwide. Therefore, the current phenotype-based screening guideline might be modified to a better genotype-based screening guideline.

Polycystic kidney disease (PKD) and Alport syndrome (AS) are serious inherited disorders associated with renal disease, and thalassemia is a hereditary blood disease with a high prevalence in south China. Here, we report an exceptional PKD coincidence of thalassemia minor and AS (diagnosed genetically) in a large Chinese family. Whole genome next-generation sequencing (NGS) was performed on the proband, and all family members underwent clinical evaluation. Sanger sequencing was used to validate the mutations distinguished by NGS. The pathogenic potential of the variants were evaluated by Polymorphism Phenotyping v2 (PolyPhen-2), Sorting Intolerant From Tolerant (SIFT) algorithm, and MutationTaster. Immunohistochemical, Western blot, immunofluorescent, and TdT-mediated dUTP nick-end labeling (TUNEL) analyses were performed to investigate polycystin 1 (PC1) expression, and cell proliferation and apoptosis in kidney tissues from the proband and normal control. A novel frameshift polycystic kidney disease 1 (PKD1) mutation (c.3903delC, p.A1302Pfs) was identified to be responsible for renal disease in this family. PC1 expression, and cell proliferation and apoptosis were significantly increased in the kidney tissues of the proband. Moreover, a deletion of approximately 19.3 kb of DNA with α-globin genes (_ _SEA) was associated with thalassemia minor in the family. In addition, a collagen type IV α 5 chain (COL4A5) variant (c.2858G>T, rs78972735), annotated as a pathogenic mutation in dbSNP and human gene mutation database (HGMD), was found in four family members with no clinical traits of AS. A novel pathogenic PKD1 mutation (c.3903delC) and (_ _SEA) thalassemia deletion were found to be responsible for the clinical symptoms in this family. The reported pathogenic COL4a5 variant (c.2858G>T, rs78972735) was not pathogenic alone.

Background Haploinsufficiency of the runt-related transcription factor 2 (RUNX2) gene is known to cause cleidocranial dysplasia (CCD). Here, we investigated a complex, heterozygous RUNX2 gene mutation in a Chinese family with CCD and the pathogenesis associated with the variations. Methods Genomic DNA extracted from peripheral venous blood was taken from the proband, her parents and 3 siblings, and 150 normal controls. Analysis of their respective RUNX2 gene sequences was performed by PCR amplification and Sanger sequencing. Pathogenesis associated with RUNX2 mutations was investigated by performing bioinformatics, real-time PCR, western blot analysis, and subcellular localization studies. Results We identified 2 complex heterozygous mutations involving a c.398–399 insACAGCAGCAGCAGCA insertion and a c.411–412 insG frameshift mutation in exon 3 of the RUNX2 gene. The frameshift mutation changed the structure of the RUNX2 protein while did not affect its expression at the mRNA level. Transfection of HEK293T cells with a plasmid expressing the RUNX2 variant decreased the molecular weight of the variant RUNX2 protein, compared with that of the wild-type protein. Subcellular localization assays showed both nuclear and cytoplasmic localization for the mutant protein, while the wild-type protein localized to the nucleus. Conclusions Our findings demonstrated that the novel c.398–399insACAGCAGCAGCAGCA mutation occurred alongside the c.411–412insG frameshift mutation, which resulted in RUNX2 truncation. RUNX2 haploinsufficiency was associated with CCD pathogenesis. These results extend the known mutational spectrum of the RUNX2 gene and suggest a functional role of the novel mutation in CCD pathogenesis.

We aimed to evaluate the interference of 50 Hz extremely low frequency electromagnetic field (ELF-EMF) occupational exposure on the neurobehavior tests of workers performing tour-inspection close to transformers and distribution power lines. Occupational short-term "spot" measurements were carried out. 310 inspection workers and 300 logistics staff were selected as exposure and control. The neurobehavior tests were performed through computer-based neurobehavior evaluation system, including mental arithmetic, curve coincide, simple visual reaction time, visual retention, auditory digit span and pursuit aiming. In 500 kV areas electric field intensity at 71.98% of total measured 590 spots were above 5 kV/m (national occupational standard), while in 220 kV areas electric field intensity at 15.69% of total 701 spots were above 5 kV/m. Magnetic field flux density at all the spots was below 1,000 μT (ICNIRP occupational standard). The neurobehavior score changes showed no statistical significance. Results of neurobehavior tests among different age, seniority groups showed no significant changes. Neurobehavior changes caused by daily repeated ELF-EMF exposure were not observed in the current study.

Background Congenital heart disease is the most common birth defect in newborns in southern China. The germline mutations in GATA4, NKX2.5, and TFAP2B genes have been identified to be responsible for congenital heart disease. The frequency of GATA4, NKX2.5, and TFAP2B mutations in subjects with congenital heart disease in southern China and the correlation between their genotype and congenital heart disease phenotype are not known. Methods We screened germline mutations in the coding exons and the flanking intron sequences of the GATA4, NKX2.5, and TFAP2B genes in 224 congenital heart disease patients located in southern China by denaturing high-performance liquid chromatography and DNA sequencing. Results Fifteen heterozygous mutations in the GATA4 gene were identified in 30 congenital heart disease patients, including a novel heterozygous missense mutation (c.788 C>G) of GATA4 in one patient with ventricular septal defect. A novel TFAP2B mutation (c.31 A>G) in a patient with endocardial cushion defect and an unreported novel TFAP2B variant (c.1006 G>A) in six patients suffering from tetralogy of Fallot (one patient), persistent truncus arteriosus (two patients) and patent ductus arteriosus (three patients) was found. There were no reported NKX2.5 mutations except for several single nucleotide polymorphisms in the patients. Conclusion These results suggest that genomic GATA4 and TFAP2B missense mutations may be associated with nonfamilial congenital heart disease with diverse clinical phenotypes in patients with congenital heart disease from southern China. They also revealed that the variation of the NKX2.5 gene may not be a risk factor for sporadic patients with congenital heart disease in this population.

Statement of problem The introduction of polymer-infiltrated ceramic network (PICN) materials may provide more options for dentists in restoring short clinical crowns and extensively damaged posterior teeth, but clinical data for their performance are lacking. Purpose The purpose of this clinical study was to compare the 3-year performance and survival rates of PICN material with those of conservative ceramic onlay restorations for endodontically treated posterior teeth using the CEREC AC chair-side system. Material and methods A total of 101 onlay restorations of endodontically treated posterior teeth using the CEREC AC chair-side system were provided in 93 participants. The 101 teeth were divided into 2 groups: Vita Enamic group and Vitablocs Mark II group. Using the modified US Public Health Service quality evaluation system, 2 calibrated evaluators examined the performance of the onlay restorations over 3 years. The Kaplan-Meier method was adopted to analyze the survival rate of restorations (α=.05). The log rank test was used to compare the survival rates of the 2 groups. The Fisher exact test was performed to detect differences in the success rates for extensively damaged teeth and short clinical crown restorations between the 2 groups. The Silness and Loe gingival index was also recorded. Results The restoration survival rates in the 2 groups were 97.0% (Vita Enamic) and 90.7% (Vitablocs Mark II) (P>.05). Five failures were recorded (4.95%). These failures were caused by restoration debonding (60%), ceramic fractures (20%), and tooth fractures (20%). There were no significant differences between the success rates of restoring extensively damaged teeth and short clinical crowns between the 2 groups (P>.05). The periodontal condition of 25% of participants was improved 3 years after the onlay restorations. Conclusions Onlay restorations of endodontically treated posterior teeth with Vita Enamic using the CEREC AC chair-side system are clinically promising prosthodontic alternatives, with a survival rate of 97.0% after 3 years. More research is needed to verify the results of this study.