Preparation of Tetrandrine Nanocrystals by Microfluidic Method and Its In Vitro and In Vivo Evaluation.

The anti-hepatocellular carcinoma effects of TET are acknowledged, but its application is hindered by its poor water solubility and low bioavailability. Conventional methods for nanocrystal preparation are laborious and lack control. To address these limitations, we propose employing the microfluidic method in the preparation of TET nanocrystals, aiming to enhance the aforementioned constraints. The objectives of this study were to prepare TET nanocrystals (TET-NC@GL) using a Y-microfluidic method with glycyrrhetinic acid (GL) as a stabilizer. The optimal preparation prescription was determined through a single-factor test and Box-Behnken response surface method. Additionally, the nanocrystals prepared with the commonly used stabilizer polyvinylpyrrolidone K30 (PVPK30), known as TET-NC@PVPK30, were characterized and evaluated for their toxicity to HepG2 cells. Hybridized nanocrystals (TET-HNC@GL and TET-HNC@PVPK30) were synthesized using a water-soluble aggregation-induced emission (AIE) fluorescent probe (TVP). Qualitative and quantitative cellular uptake experiments were conducted using these hybridized nanocrystals. Conducting in vivo pharmacokinetic assays evaluates the relative bioavailability of nanocrystals. The results indicated that TET-NC@GL, optimized using the response surface method, had a particle size of 136.47 ± 3.31 nm and a PDI of 0.219 ± 0.002. The administration of TET-NC@GL significantly enhanced the cell inhibition rate compared to the TET group and the TET-NC@PVPK30 group (P < 0.01). Moreover, the qualitative and quantitative cellular uptake results revealed a significant enhancement in cellular uptake in the TET-HNC@GL administration group compared to the TET-HNC@PVPK30 group (P < 0.01). In vivo pharmacokinetic results showed that the bioavailability of TET-NC@GL group was 3.5 times higher than that of the TET group. The results demonstrate the successful preparation of TET-NC@GL nanocrystals.

EGR2 is a hub-gene in myocardial infarction and aggravates inflammation and apoptosis in hypoxia-induced cardiomyocytes.

BACKGROUND: Myocardial infarction (MI) is characterized by coronary artery occlusion, ischemia and hypoxia of myocardial cells, leading to irreversible myocardial damage. Therefore, it is urgent to explore the potential mechanism of myocardial injury during the MI process to develop effective therapies for myocardial cell rescue. METHODS: We downloaded the GSE71906 dataset from GEO DataSets, and the R software was used to identify the differentially expressed genes (DEGs) in mouse heart tissues of MI and sham controls. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment were performed to understand the significantly activated signaling pathways in MI. Protein-protein interaction (PPI) network was constructed to highlight the hub genes in DEGs. The Western Blot, qRT-PCR and TUNEL staining were used to explore the function of hub gene in hypoxia-induced cardiomyocytes in vitro. RESULTS: A total of 235 DEGs were identified in GSE71906 dataset. Functional enrichment analysis revealed that the upregulated genes were primarily associated with the inflammatory response and apoptosis. 20 hub genes were identified in PPI network, and the early growth response 2 (EGR2) was highlighted. In vitro. We confirmed the EGR2 was upregulated induced by hypoxia and revealed the upregulated EGR2 aggravates pro-inflammation and pro-apoptotic genes expression. In addition, EGR2 knockout mitigates hypoxia-induced inflammation and apoptosis in cardiomyocytes. CONCLUSION: The present study identified the EGR2 was a hub gene in myocardial damage during MI process, the excessive EGR2 aggravates hypoxia-induced myocardial damage by accelerating inflammation and apoptosis in vitro. Therefore, targeting EGR2 offers a potential pharmacological strategy for myocardial cell rescue in MI.

Optimizing genomic prediction model given causal genes in a dairy cattle population.

As genotypic data are moving from SNP chip toward whole-genome sequence, the accuracy of genomic prediction (GP) exhibits a marginal gain, although all genetic variation, including causal genes, are contained in whole-genome sequence data. Meanwhile, genetic analyses on complex traits, such as genome-wide association studies, have identified an increasing number of genomic regions, including potential causal genes, which would be reliable prior knowledge for GP. Many studies have tried to improve the performance of GP by modifying the prediction model to incorporate prior knowledge. Although several plausible results have been obtained from model modification or strategy optimization, most of them were validated in a specific empirical population with a limited variety of genetic architecture for complex traits. An alternative approach is to use simulated genetic architecture with known causal genes (e.g., simulated causative SNP) to evaluate different GP models with given causal genes. Our objectives were to (1) evaluate the performance of GP under a variety of genetic architectures with a subset of known causal genes and (2) compare different GP models modified by highlighting causal genes and different strategies to weight causal genes. In this study, we simulated pseudo-phenotypes under a variety of genetic architectures based on the real genotypes and phenotypes of a dairy cattle population. Besides classical genomic best linear unbiased prediction, we evaluated 3 modified GP models that highlight causal genes as follows: (1) by treating them as fixed effects, (2) by treating them as a separate random component, and (3) by combining them into the genomic relationship matrix as random effects. Our results showed that highlighting the known causal genes, which explained a considerable proportion of genetic variance in the GP models, increased the predictive accuracy. Combining all given causal genes into the genomic relationship matrix was the optimal strategy under all the scenarios validated, and treating causal genes as a separate random component is also recommended, when more than 20% of genetic variance was explained by known causal genes. Moreover, assigning differential weights to each causal gene further improved the predictive accuracy.

Dynamic DNA methylation of ovaries during pubertal transition in gilts.

BACKGROUND: In female mammals, the initiation of puberty, coupling with the dramatically morphological changes in ovaries, indicates the sexual and follicular maturation. Previous studies have suggested that the disrupted DNA methylation results in the delayed puberty. However, to date, the changes in ovarian methylomes during pubertal transition have not been investigated. In this study, using gilts as a pubertal model, the genome-wide DNA methylation were profiled to explore their dynamics during pubertal transition across Pre-, In- and Post-puberty. RESULTS: During pubertal transition, the follicles underwent maturation and luteinization, coupled with the significant changes in the mRNA expression of DNMT1 and DNMT3a. DNA methylation levels of In-puberty were higher than that of Pre- and Post-puberty at the locations of genes and CpG islands (CGIs). Analysis of the DNA methylation changes identified 12,313, 20,960 and 17,694 differentially methylated CpGs (DMCs) for the comparisons of Pre- vs. In-, In vs. Post-, and Pre- vs. Post-puberty, respectively. Moreover, the CGIs, upstream and exonic regions showed a significant underrepresentation of DMCs, but the CGI shores, CGI shelves, intronic, downstream and intergenic regions showed a significant overrepresentation of DMCs. Furthermore, biological functions of these methylation changes enriched in PI3K-Akt signaling pathway, GnRH signaling pathway, and Insulin secretion, and the mRNA expressions of several genes of these signaling pathway, including MMP2, ESR1, GSK3B, FGF21, IGF1R, and TAC3, were significantly changed across Pre-, In- and Post-puberty in ovaries. CONCLUSIONS: During pubertal transition in gilts, the DNA methylation changes of ovaries were likely to affect the transcription of genes related to PI3K-Akt signaling pathway, GnRH signaling pathway, and Insulin secretion. These observations can provide new insight into the epigenetic mechanism of follicular and sexual maturation during pubertal transition in mammals.

Genome wide association study on feed conversion ratio using imputed sequence data in chickens.

OBJECTIVE: Feed consumption contributes a large percentage for total production costs in the poultry industry. Detecting genes associated with feeding traits will be of benefit to improve our understanding of the molecular determinants for feed efficiency. The objective of this study was to identify candidate genes associated with feed conversion ratio (FCR) via genome-wide association study (GWAS) using sequence data imputed from single nucleotide polymorphism (SNP) panel in a Chinese indigenous chicken population. METHODS: A total of 435 Chinese indigenous chickens were phenotyped for FCR and were genotyped using a 600K SNP genotyping array. Twenty-four birds were selected for sequencing, and the 600K SNP panel data were imputed to whole sequence data with the 24 birds as the reference. The GWAS were performed with GEMMA software. RESULTS: After quality control, 8,626,020 SNPs were used for sequence based GWAS, in which ten significant genomic regions were detected to be associated with FCR. Ten candidate genes, ubiquitin specific peptidase 44, leukotriene A4 hydrolase, ETS transcription factor, R-spondin 2, inhibitor of apoptosis protein 3, sosondowah ankyrin repeat domain family member D, calmodulin regulated spectrin associated protein family member 2, zinc finger and BTB domain containing 41, potassium sodium-activated channel subfamily T member 2, and member of RAS oncogene family were annotated. Several of them were within or near the reported FCR quantitative trait loci, and others were newly reported. CONCLUSION: Results from this study provide valuable prior information on chicken genomic breeding programs, and potentially improve our understanding of the molecular mechanism for feeding traits.

Incomplete penetrance, variable expressivity, or dosage insensitivity in four families with directly transmitted unbalanced chromosome abnormalities.

The direct transmission of microscopically visible unbalanced chromosome abnormalities (UBCAs) is rare and usually has phenotypic consequences. Here we report four families in which a normal phenotype was initially found in one or more family members. Each UBCA was interpreted with regard to overlapping examples and factors previously associated with transmitted imbalances including incidental ascertainment, low gene density, benign copy number variation (CNV) content, and gene relatedness. A 4.56 Mb deletion of 8p23.1-p23.2 was thought to be causal in the affected proband but showed incomplete penetrance in her mother and sibling (Family 1). Incomplete penetrance was also associated with a 10.88 Mb duplication of 13q21.31-q22.1 (Family 3) and dosage insensitivity with a 17.6 Mb deletion of 22pter-q11.21 (Family 4) that were both ascertained at prenatal diagnosis and each found in 4 unaffected family members. The 22pter-q11.21 deletion is part of a region with high benign CNV content and supports the mapping of cat eye syndrome to a 600 kb interval of 22q11.1-q11.21. Low gene densities of less than 2.0 genes/Mb were found in each of these three families but only after segmentally duplicated genes were excluded from the deletions of 8p and 22q. In contrast, gene density was average and variable expressivity associated with a 3.59 Mb duplication of 8p23.1 incidentally ascertained for paternal infertility (Family 2). Our results indicate that a greater degree of direct parental transmission, incomplete penetrance, and variable expression are features of both sub-microscopic CNVs and UBCAs with relatively low gene and high benign CNV content.

Pallister-Killian syndrome: a study of 22 British patients.

BACKGROUND: Pallister-Killian syndrome is a rare, sporadic condition caused by mosaic tetrasomy of the short arm of chromosome 12 (12p). The main features are intellectual disability, seizures, dysmorphic features and a variety of congenital malformations. Most available information comes from individual case reports. We report the results of a British study into Pallister-Killian syndrome, which is the first to provide comprehensive data on a population-based sample. METHOD: A detailed phenotypical study was carried out in Great Britain. All individuals with Pallister-Killian syndrome were eligible to participate. Each participant underwent a structured history, developmental assessment and clinical examination. Buccal mucosal samples were analysed by interphase fluorescence in situ hybridization (FISH) and blood samples by array comparative genomic hybridization (CGH). Genotype-phenotype correlations were sought in these tissues and existing skin biopsy reports. RESULTS: Twenty-two patients with Pallister-Killian syndrome, ranging from 4 months to 31 years were recruited and comprehensive data on each obtained. The birth incidence was 5.1 per million live births. Array CGH only suggested the diagnosis in 15.8% but buccal FISH could have made the diagnosis in 75.0%. There was no genotype-phenotype correlation in any of the tissues studied. This study shows that the high birth weights and profound intellectual disability classically described in Pallister-Killian syndrome are not universal. Mild or moderate intellectual disability was present in 27.6% of this cohort and all birth weights were within 2.67SD of the mean. New features which have not previously been recognised as part of Pallister-Killian syndrome include anhydrosis/hypohydrosis and episodic hyperventilation, suggesting involvement of the autonomic system.

A reliable method for the detection of BRCA1 and BRCA2 mutations in fixed tumour tissue utilising multiplex PCR-based targeted next generation sequencing.

BACKGROUND: Germline mutations in BRCA1 or BRCA2 lead to a high lifetime probability of developing ovarian or breast cancer. These genes can also be involved in the development of non-hereditary tumours as somatic BRCA1/2 pathogenic variants are found in some of these cancers. Since patients with somatic BRCA pathogenic variants may benefit from treatment with poly ADP ribose polymerase inhibitors, it is important to be able to test for somatic changes in routinely available tumour samples. Such samples are typically formalin-fixed paraffin-embedded (FFPE) tissue, where the extracted DNA tends to be highly fragmented and of limited quantity, making analysis of large genes such as BRCA1 and BRCA2 challenging. This is made more difficult as somatic changes may be evident in only part of the sample, due to the presence of normal tissue. METHODS: We examined the feasibility of analysing DNA extracted from FFPE ovarian and breast tumour tissue to identify significant DNA variants in BRCA1/ BRCA2 using next generation sequencing methods that were sensitive enough to detect low level mutations, multiplexed to reduce the amount of DNA required and had short amplicon design. The utility of two GeneRead DNAseq Targeted Exon Enrichment Panels with different designs targeting only BRCA1/2 exons, and the Ion AmpliSeq BRCA community panel, followed by library preparation and adaptor ligation using the TruSeq DNA PCR-Free HT Sample Preparation Kit and NGS analysis on the MiSeq were investigated. RESULTS: Using the GeneRead method, we successfully analysed over 76% of samples, with >95% coverage of BRCA1/2 coding regions and a mean average read depth of >1000-fold. All mutations identified were confirmed where possible by Sanger sequencing or replication to eliminate the risk of false positive results due to artefacts within FFPE material. Admixture experiments demonstrated that BRCA1/2 variants could be detected if present in >10% of the sample. A sample subset was evaluated using the Ion AmpliSeq BRCA panel, achieving >99% coverage and sufficient read depth for a proportion of the samples. CONCLUSIONS: Detection of BRCA1/2 variants in fixed tissue is feasible, and could be performed prospectively to facilitate optimum treatment decisions for ovarian or breast cancer patients.

Spexin inhibits excessive autophagy-induced ferroptosis to alleviate doxorubicin-induced cardiotoxicity by upregulating Beclin 1.

BACKGROUND AND PURPOSE: Doxorubicin is widely used in the treatment of malignant tumours, but doxorubicin-induced cardiotoxicity severely limits its clinical application. Spexin is a neuropeptide that acts as a novel biomarker in cardiovascular disease. However, the effects of spexin on doxorubicin-induced cardiotoxicity is unclear. EXPERIMENTAL APPROACH: We established a model of doxorubicin-induced cardiotoxicity both in vivo and in vitro. Levels of cardiac damage in mice was assessed through cardiac function assessment, determination of serum cardiac troponin T and CKMB levels and histological examination. CCK8 and PI staining were used to assess the doxorubicin-induced toxicity in cultures of cardiomyocytes in vitro. Ferroptosis was assessed using FerroOrange staining, determination of MDA and 4-HNE content and ferroptosis-associated proteins SLC7A11 and GPX4. Mitochondrial membrane potential and lipid peroxidation levels were measured using TMRE and C11-BODIPY 581/591 probes, respectively. Myocardial autophagy was assessed by expression of P62 and Beclin1. KEY RESULTS: Spexin treatment improved heart function of mice with doxorubicin-induced cardiotoxicity, and attenuated doxorubicin-induced cardiotoxicity by decreasing iron accumulation, abnormal lipid metabolism and inhibiting ferroptosis. Interestingly, doxorubicin caused excessive autophagy in cardiomyocyte in culture, which could be alleviated by treatment with spexin. Knockdown of Beclin 1 eliminated the protective effects of spexin in mice with DIC. CONCLUSION AND IMPLICATIONS: Spexin ameliorated doxorubicin-induced cardiotoxicity by inhibiting excessive autophagy-induced ferroptosis, suggesting that spexin could be a drug candidate against doxorubicin-induced cardiotoxicity. Beclin 1 might be critical in mediating the protective effect of spexin against doxorubicin-induced cardiotoxicity.

Deletion 17q12 is a recurrent copy number variant that confers high risk of autism and schizophrenia.

Autism spectrum disorders (ASD) and schizophrenia are neurodevelopmental disorders for which recent evidence indicates an important etiologic role for rare copy number variants (CNVs) and suggests common genetic mechanisms. We performed cytogenomic array analysis in a discovery sample of patients with neurodevelopmental disorders referred for clinical testing. We detected a recurrent 1.4 Mb deletion at 17q12, which harbors HNF1B, the gene responsible for renal cysts and diabetes syndrome (RCAD), in 18/15,749 patients, including several with ASD, but 0/4,519 controls. We identified additional shared phenotypic features among nine patients available for clinical assessment, including macrocephaly, characteristic facial features, renal anomalies, and neurocognitive impairments. In a large follow-up sample, the same deletion was identified in 2/1,182 ASD/neurocognitive impairment and in 4/6,340 schizophrenia patients, but in 0/47,929 controls (corrected p = 7.37 × 10⁻5). These data demonstrate that deletion 17q12 is a recurrent, pathogenic CNV that confers a very high risk for ASD and schizophrenia and show that one or more of the 15 genes in the deleted interval is dosage sensitive and essential for normal brain development and function. In addition, the phenotypic features of patients with this CNV are consistent with a contiguous gene syndrome that extends beyond RCAD, which is caused by HNF1B mutations only.

Identification of a diagnostic model and molecular subtypes of major depressive disorder based on endoplasmic reticulum stress-related genes.

Subject: Major depressive disorder (MDD) negatively affects patients' behaviours and daily lives. Due to the high heterogeneity and complex pathological features of MDD, its diagnosis remains challenging. Evidence suggests that endoplasmic reticulum stress (ERS) is involved in the pathogenesis of MDD; however, relevant diagnostic markers have not been well studied. This study aimed to screen for ERS genes with potential diagnostic value in MDD. Methods: Gene expression data on MDD samples were downloaded from the GEO database, and ERS-related genes were obtained from the GeneCards and MSigDB databases. Differentially expressed genes (DEGs) in MDD patients and healthy subjects were identified and then integrated with ERS genes. ERS diagnostic model and nomogram were developed based on biomarkers screened using the LASSO method. The diagnostic performance of this model was evaluated. ERS-associated subtypes were identified. CIBERSORT and GSEA were used to explore the differences between the different subtypes. Finally, WGCNA was performed to identify hub genes related to the subtypes. Results: A diagnostic model was developed based on seven ERS genes: KCNE1, PDIA4, STAU1, TMED4, MGST1, RCN1, and SHC1. The validation analysis showed that this model had a good diagnostic performance. KCNE1 expression was positively correlated with M0 macrophages and negatively correlated with resting CD4+ memory T cells. Two subtypes (SubA and SubB) were identified, and these two subtypes showed different ER score. The SubB group showed higher immune infiltration than the SubA group. Finally, NCF4, NCF2, CSF3R, and FPR2 were identified as hub genes associated with ERS molecular subtypes. Conclusion: Our current study provides novel diagnostic biomarkers for MDD from an ERS perspective, and these findings further facilitate the use of precision medicine in MDD.

Bibliometric and visual analysis in the field of traditional Chinese medicine in cancer from 2002 to 2022.

Objective: Traditional Chinese medicine (TCM) has been used as a complementary treatment for cancer patients, but there has been no quantitative comprehensive analysis of TCM's efficacy. The purpose of this paper is to explore the current status and hotspots of TCM in cancer research from 2002 to 2022 and to provide a reference for future research. Methods: We retrieved articles published between 2002 and 2022 from the Web of Science database and analyzed them using R software, VOSviewer, and CiteSpace software. Results: A total of 7,129 articles were included in this study. The publication rate of TCM cancer research increased steadily from 2002 to 2022, with a rapid increase from 2010 to 2021. China was the country with the most published articles, followed by the United States, Republic of Korea, Germany, and Japan. China was also the country with the most international collaborations, and China Medical University and Shanghai University of Traditional Chinese Medicine were the most representative cooperation centers. The Journal of Ethnopharmacology was the most published and cited journal. Apoptosis, expression, in vitro, activation, and other related keywords were commonly used in these articles. Breast cancer, colorectal cancer, gastric cancer, liver cancer, and lung cancer were the most studied cancer types in TCM research. Pathway-related apoptosis, anti-inflammation, and oxidative stress were the hotspots and trends of TCM's anti-cancer mechanism. Metabolomics combined with network pharmacology was the main research method. Conclusion: Traditional Chinese medicine as an anti-cancer drug has received increasing attention from researchers worldwide, and it is expected to be a hotspot for developing new anti-cancer drugs in the future. Our study provides a comprehensive analysis of the current status and hotspots of TCM cancer research, which could serve as a valuable reference for future studies.

Using the spring constant method to analyze arterial elasticity in type 2 diabetic patients.

BACKGROUND: This study tests the validity of a newly-proposed spring constant method to analyze arterial elasticity in type 2 diabetic patients. METHODS: The experimental group comprised 66 participants (36 men and 30 women) ranging between 46 and 86 years of age, all with diabetes mellitus. In the experimental group, 21 participants suffered from atherosclerosis. All were subjected to the measurements of both the carotid-femoral pulse wave velocity (cfPWV) and the spring constant method. The comparison (control) group comprised 66 normal participants (37 men and 29 women) with an age range of 40 to 80 years who did not have diabetes mellitus. All control group members were subjected to measurement by the spring constant method. RESULTS: Statistical analysis of the experimental and control groups indicated a significant negative correlation between the spring constant and the cfPWV (P < .001; r = - 0.824 and - 0.71). Multivariate analysis similarly indicated a close relationship. The Student's t test was used to examine the difference in the spring constant parameter between the experimental and control groups. A P-value less than .05 confirmed that the difference between the 2 groups was statistically significant. In receiver operating characteristic curve (ROC), the Area Under Curve (AUC, = 0.85) indicates good discrimination. These findings imply that the spring constant method can effectively identify normal versus abnormal characteristics of elasticity in normal and diabetic participants. CONCLUSIONS: This study verifies the use of the spring constant method to assess arterial elasticity, and found it to be efficient and simple to use. The spring constant method should prove useful not only for improving clinical diagnoses, but also for screening diabetic patients who display early evidence of vascular disease.

Analysis of using the tongue deviation angle as a warning sign of a stroke.

BACKGROUND: The symptom of tongue deviation is observed in a stroke or transient ischemic attack. Nevertheless, there is much room for the interpretation of the tongue deviation test. The crucial factor is the lack of an effective quantification method of tongue deviation. If we can quantify the features of the tongue deviation and scientifically verify the relationship between the deviation angle and a stroke, the information provided by the tongue will be helpful in recognizing a warning of a stroke. METHODS: In this study, a quantification method of the tongue deviation angle was proposed for the first time to characterize stroke patients. We captured the tongue images of stroke patients (15 males and 10 females, ranging between 55 and 82 years of age); transient ischemic attack (TIA) patients (16 males and 9 females, ranging between 53 and 79 years of age); and normal subjects (14 males and 11 females, ranging between 52 and 80 years of age) to analyze whether the method is effective. In addition, we used the receiver operating characteristic curve (ROC) for the sensitivity analysis, and determined the threshold value of the tongue deviation angle for the warning sign of a stroke. RESULTS: The means and standard deviations of the tongue deviation angles of the stroke, TIA, and normal groups were: 6.9 ± 3.1, 4.9 ± 2.1 and 1.4 ± 0.8 degrees, respectively. Analyzed by the unpaired Student's t-test, the p-value between the stroke group and the TIA group was 0.015 (>0.01), indicating no significant difference in the tongue deviation angle. The p-values between the stroke group and the normal group, as well as between the TIA group and the normal group were both less than 0.01. These results show the significant differences in the tongue deviation angle between the patient groups (stroke and TIA patients) and the normal group. These results also imply that the tongue deviation angle can effectively identify the patient group (stroke and TIA patients) and the normal group. With respect to the visual examination, 40% and 32% of stroke patients, 24% and 16% of TIA patients, and 4% and 0% of normal subjects were found to have tongue deviations when physicians "A" and "B" examined them. The variation showed the essentiality of the quantification method in a clinical setting. In the receiver operating characteristic curve (ROC), the Area Under Curve (AUC, = 0.96) indicates good discrimination. The tongue deviation angle more than the optimum threshold value (= 3.2°) predicts a risk of stroke. CONCLUSIONS: In summary, we developed an effective quantification method to characterize the tongue deviation angle, and we confirmed the feasibility of recognizing the tongue deviation angle as an early warning sign of an impending stroke.

Recurrent rearrangements of chromosome 1q21.1 and variable pediatric phenotypes.

BACKGROUND: Duplications and deletions in the human genome can cause disease or predispose persons to disease. Advances in technologies to detect these changes allow for the routine identification of submicroscopic imbalances in large numbers of patients. METHODS: We tested for the presence of microdeletions and microduplications at a specific region of chromosome 1q21.1 in two groups of patients with unexplained mental retardation, autism, or congenital anomalies and in unaffected persons. RESULTS: We identified 25 persons with a recurrent 1.35-Mb deletion within 1q21.1 from screening 5218 patients. The microdeletions had arisen de novo in eight patients, were inherited from a mildly affected parent in three patients, were inherited from an apparently unaffected parent in six patients, and were of unknown inheritance in eight patients. The deletion was absent in a series of 4737 control persons (P=1.1x10(-7)). We found considerable variability in the level of phenotypic expression of the microdeletion; phenotypes included mild-to-moderate mental retardation, microcephaly, cardiac abnormalities, and cataracts. The reciprocal duplication was enriched in nine children with mental retardation or autism spectrum disorder and other variable features (P=0.02). We identified three deletions and three duplications of the 1q21.1 region in an independent sample of 788 patients with mental retardation and congenital anomalies. CONCLUSIONS: We have identified recurrent molecular lesions that elude syndromic classification and whose disease manifestations must be considered in a broader context of development as opposed to being assigned to a specific disease. Clinical diagnosis in patients with these lesions may be most readily achieved on the basis of genotype rather than phenotype.

An evidence-based approach to establish the functional and clinical significance of copy number variants in intellectual and developmental disabilities.

PURPOSE: Copy number variants have emerged as a major cause of human disease such as autism and intellectual disabilities. Because copy number variants are common in normal individuals, determining the functional and clinical significance of rare copy number variants in patients remains challenging. The adoption of whole-genome chromosomal microarray analysis as a first-tier diagnostic test for individuals with unexplained developmental disabilities provides a unique opportunity to obtain large copy number variant datasets generated through routine patient care. METHODS: A consortium of diagnostic laboratories was established (the International Standards for Cytogenomic Arrays consortium) to share copy number variant and phenotypic data in a central, public database. We present the largest copy number variant case-control study to date comprising 15,749 International Standards for Cytogenomic Arrays cases and 10,118 published controls, focusing our initial analysis on recurrent deletions and duplications involving 14 copy number variant regions. RESULTS: Compared with controls, 14 deletions and seven duplications were significantly overrepresented in cases, providing a clinical diagnosis as pathogenic. CONCLUSION: Given the rapid expansion of clinical chromosomal microarray analysis testing, very large datasets will be available to determine the functional significance of increasingly rare copy number variants. This data will provide an evidence-based guide to clinicians across many disciplines involved in the diagnosis, management, and care of these patients and their families.

Transmitted deletions of medial 5p and learning difficulties; does the cadherin cluster only become penetrant when flanking genes are deleted?

The central portion of the short arm of chromosome 5 is unusual in that large, cytogenetically visible interstitial deletions segregate in families with and without phenotypic consequences. Here we present a family in which a transmitted interstitial deletion of 5p13.3 to 5p14.3 co-segregated with learning and/or behavioral difficulties in six family members. Facial dysmorphism was not striking but a father and daughter both had lacrimal fistulae. The deletion was 12.23 Mb in size (chr5:20,352,535-32,825,775) and contained fifteen known protein coding genes. Five of these (GOLPH3; MTMR12; ZFR; SUB1; and NPR3) and an ultra-conserved microRNA (hsa-miR-579) were present in an 883 kb candidate gene region in 5p13.3 that was deleted in the present family but not in previously reported overlapping benign deletions. Members of the cadherin precursor gene cluster, with brain specific expression, were deleted in both affected and benign deletion families. The candidate genes in 5p13.3 may be sufficient to account for the consistent presence or absence of phenotype in medial 5p deletions. However, we consider the possibility of position effects in which CDH6, and/or other cadherin genes, become penetrant when adjacent genes, or modifiers of gene expression, are also deleted. This could account for the absence of intellectual disability in benign deletions of the cadherin cluster, the cognitive phenotype in medial 5p deletion syndrome and the greater severity of intellectual disability in patients with cri-du-chat syndrome and deletions of 5p15 that extend into the region deleted in the present family.

Germline FFPE inherited cancer panel testing in deceased family members: implications for clinical management of unaffected relatives.

Where previously, germline genetic testing in deceased affected relatives was not possible due to the absence of lymphocytic DNA, the North-West-Genomic-Laboratory Hub (NWGLH) has developed and validated next-generation sequencing based gene panels utilising formalin-fixed-paraffin-embedded (FFPE) tissue DNA from deceased individuals. This technology has been utilised in the clinical setting for the management of unaffected relatives seen in the Clinical Genetics Service (CGS). Here we assess the clinical impact. At the time of data collection, the NWGLH had analysed 180 FFPE tissue samples from deceased affected individuals: 134 from breast and/or ovarian cancer cases for germline variants in the BRCA1/BRCA2 genes and 46 from colorectal, gastric, ovarian and endometrial cancer cases for germline variants in a panel of 13 genes implicated in inherited colorectal cancer and gastric cancer conditions. Successful analysis was achieved in 140/180 cases (78%). In total, 29 germline pathogenic/likely pathogenic variants were identified in autosomal dominant cancer predisposition genes where the gene was pertinent to the cancer family history (including BRCA1/BRCA2, the mismatch-repair genes and APC). Of the 180 cases, the impact of the result on clinical management of unaffected relatives was known in 143 cases. Of these, the results in 54 cases (38%) directly impacted the clinical management of relatives seen by the CGS. This included changes to risk assessments, screening recommendations and the availability of predictive genetic testing to unaffected relatives. Our data demonstrate how FFPE testing in deceased relatives is an accurate and informative tool in the clinical management of patients referred to the CGS.

The Improvement Effect of Different Doses of Gamma Globulin on the Disease Condition of Infants with Hemolytic Disease of Newborn and Their Effects on Immune Factors in Serum.

BACKGROUND: To explore the effect of different doses of Gamma Globulin (GG) on the condition of children with Hemolytic Disease of Newborn (HDN) and the influence of immune factors in serum. METHODS: Overall, 180 infants with hemolytic disease of newborn in the People's Hospital of Zhangqiu Area, Jinan, China from April 2016 to August 2018 were divided into group A (88 cases) and group B (92 cases). Group A was given intravenous low-dose GG on the basis of phototherapy, and group B was given intravenous high-dose GG on the basis of phototherapy. The level of serum total bilirubin of the infants, the levels of CD3+, CD4+, CD8+, IgA, IgG and IgM of the infants, the time of jaundice disappearance and the length of hospital stay, hemoglobin and reticulocyte levels were recorded before treatment and after treatment. The number and condition of adverse reactions were recorded. RESULTS: After treatment, the levels of TBiL, hemoglobin and reticulocyte, the time of jaundice disappearance and hospital stay in group B were significantly lower than those in group A. The level of immune cells in group B was significantly higher than that in group A after 7 days of treatment, and the levels of IgA / IgG / IgM in group B were significantly higher than those in group A after 28 days of treatment. CONCLUSION: Intravenous high-dose GG has a better effect on the condition of neonatal hemolytic disease patients, and more effectively improve the immune function of children.

Breakpoint mapping and haplotype analysis of three reciprocal translocations identify a novel recurrent translocation in two unrelated families: t(4;11)(p16.2;p15.4).

The majority of constitutional reciprocal translocations appear to be unique rearrangements arising from independent events. However, a small number of translocations are recurrent, most significantly the t(11;22)(q23;q11). Among large series of translocations there may be multiple independently ascertained cases with the same cytogenetic breakpoints. Some of these could represent additional recurrent rearrangements, alternatively they could be identical by descent (IBD) or have subtly different breakpoints when examined under higher resolution. We have used molecular breakpoint mapping and haplotyping to determine the origin of three pairs of reciprocal constitutional translocations, each with the same cytogenetic breakpoints. FISH mapping showed one pair to have different breakpoints and thus to be distinct rearrangements. Another pair of translocations were IBD with identical breakpoint intervals and highly conserved haplotypes on the derived chromosomes. The third pair, t(4;11)(p16.2;p15.4), had the same breakpoint intervals by aCGH and fosmid mapping but had very different haplotypes, therefore they represent a novel recurrent translocation. Unlike the t(11;22)(q23;q11), the formation of the t(4;11)(p16.2;p15.4) may have involved segmental duplications and sequence homology at the breakpoints. Additional examples of recurrent translocations could be identified if the resources were available to study more translocations using the approaches described here. However, like the t(4;11)(p16.2;p15.4), such translocations are likely to be rare with the t(11;22) remaining the only common recurrent constitutional reciprocal translocation.