Genetic etiology study in a large cohort with congenital insensitivity to pain with anhidrosis.

ABSTRACT: Pathogenic variations in the NTRK1 can cause congenital insensitivity to pain with anhidrosis (CIPA), a rare autosomal recessive inherited neuropathy. The precise diagnosis of CIPA relies on the identification of pathogenic genotypes. Therefore, it is essential to expand the NTRK1 variation spectrum and improve molecular diagnosis methods. In this study, 74 probands with typical manifestations of CIPA but unknown genotypes were recruited. A comprehensive molecular genetic analysis was performed to identify variations in the NTRK1, using techniques including Sanger and next-generation sequencing, bioinformatic analysis, quantitative polymerase chain reaction (qPCR), gap-PCR, short tandem repeat (STR) genotyping, and reverse-transcription PCR. In addition, functional assays were conducted to determine the pathogenicity of variants of uncertain significance (VUS) and further characterized changes in glycosylation and phosphorylation of 14 overexpressed mutant vectors with variants at different domains in the TrkA protein, which is encoded by NTRK1. A total of 48 variations in the NTRK1 were identified, including 22 novel ones. When combined with data from another 53 CIPA patients examined in our previous work, this study establishes the largest genotypic and phenotypic spectra of CIPA worldwide, including 127 CIPA families. Moreover, functional studies indicated that the pathogenicity of VUS mainly affected insufficient glycosylation in the extracellular domain and abnormal phosphorylation in the intracellular domain. This study not only provides important evidence for precise diagnosis of CIPA but also further enriches our understanding of the pathogenesis of this disease.

The pathogenic mechanism of syndactyly type V identified in a Hoxd13Q50R knock-in mice.

Syndactyly type V (SDTY5) is an autosomal dominant extremity malformation characterized by fusion of the fourth and fifth metacarpals. In the previous publication, we first identified a heterozygous missense mutation Q50R in homeobox domain (HD) of HOXD13 in a large Chinese family with SDTY5. In order to substantiate the pathogenicity of the variant and elucidate the underlying pathogenic mechanism causing limb malformation, transcription-activator-like effector nucleases (TALEN) was employed to generate a Hoxd13Q50R mutant mouse. The mutant mice exhibited obvious limb malformations including slight brachydactyly and partial syndactyly between digits 2-4 in the heterozygotes, and severe syndactyly, brachydactyly and polydactyly in homozygotes. Focusing on BMP2 and SHH/GREM1/AER-FGF epithelial mesenchymal (e-m) feedback, a crucial signal pathway for limb development, we found the ectopically expressed Shh, Grem1 and Fgf8 and down-regulated Bmp2 in the embryonic limb bud at E10.5 to E12.5. A transcriptome sequencing analysis was conducted on limb buds (LBs) at E11.5, revealing 31 genes that exhibited notable disparities in mRNA level between the Hoxd13Q50R homozygotes and the wild-type. These genes are known to be involved in various processes such as limb development, cell proliferation, migration, and apoptosis. Our findings indicate that the ectopic expression of Shh and Fgf8, in conjunction with the down-regulation of Bmp2, results in a failure of patterning along both the anterior-posterior and proximal-distal axes, as well as a decrease in interdigital programmed cell death (PCD). This cascade ultimately leads to the development of syndactyly and brachydactyly in heterozygous mice, and severe limb malformations in homozygous mice. These findings suggest that abnormal expression of SHH, FGF8, and BMP2 induced by HOXD13Q50R may be responsible for the manifestation of human SDTY5.

Rapid Screening of 34 Emerging Contaminants in Surface Water by UHPLC-Q-TOF-MS.

OBJECTIVES: To establish a rapid screening method for 34 emerging contaminants in surface water by ultra-high performance liquid chromatography-quadrupole-time of flight mass spectrometry (UHPLC-Q-TOF-MS). METHODS: The pretreatment conditions of solid phase extraction (SPE) were optimized by orthogonal experimental design and the surface water samples were concentrated and extracted by Oasis® HLB and Oasis® MCX SPE columns in series. The extracts were separated by Kinetex® EVO C18 column, with gradient elution of 0.1% formic acid aqueous solution and 0.1% formic acid methanol solution. Q-TOF-MS 'fullscan' and 'targeted MS/MS' modes were used to detect 34 emerging contaminants and to establish a database with 34 emerging contaminants precursor ion, product ion and retention times. RESULTS: The 34 emerging contaminants exhibited good linearity in the concentration range respectively and the correlation coefficients (r) were higher than 0.97. The limit of detection was 0.2-10 ng/L and the recoveries were 81.2%-119.2%. The intra-day precision was 0.78%-18.70%. The method was applied to analyze multiple surface water samples and 6 emerging contaminants were detected, with a concentration range of 1.93-157.71 ng/L. CONCLUSIONS: The method is simple and rapid for screening various emerging contaminants at the trace level in surface water.

The effect of total sleep deprivation on working memory: evidence from diffusion model.

STUDY OBJECTIVES: Working memory is crucial in human daily life and is vulnerable to sleep loss. The current study investigated the impact of sleep deprivation on working memory from the information processing perspective, to explore whether sleep deprivation affects the working memory via impairing information manipulation. METHODS: Thirty-seven healthy adults attended two counterbalanced protocols: a normal sleep night and a total sleep deprivation (TSD). The N-back and the psychomotor vigilance task (PVT) assessed working memory and sustained attention. Response time distribution and drift-diffusion model analyses were applied to explore cognitive process alterations. RESULTS: TSD increased the loading effect of accuracy, but not the loading effect of response time in the N-back task. TSD reduced the speed of information accumulation, increased the variability of the speed of accumulation, and elevated the decision threshold only in 1-back task. Moreover, the slow responses of PVT and N-back were severely impaired after TSD, mainly due to increased information accumulation variability. CONCLUSIONS: The present study provides a new perspective to investigate behavioral performance by using response time distribution and drift-diffusion models, revealing that sleep deprivation affected multicognitive processes underlying working memory, especially information accumulation processes.

Effect of homeostatic pressure and circadian rhythm on the task-switching: Evidence from drift diffusion model and ERP.

The effect of diurnal fluctuations on cognitive functions is widely studied, yet rare research has attempted to separate the role of two crucial processes underlying diurnal fluctuations: homeostatic pressure and circadian rhythm. The present study aimed to dissociate their effects by conducting a task-switching task in the morning, napping afternoon, and no-napping afternoon, respectively. Additionally, DDM and ERP were utilized to explore how these two processes differentially affect cognitive processes involved in task-switching. By a within-participant design, 35 healthy adults (20.03 ± 2.01 year-old, 14 males) with an intermediate-type chronotype were recruited in the current study. The results demonstrated that accumulated homeostatic pressure caused reduced accuracy, drift rate, and decision threshold. In the no-napping afternoon, P1 and P2 amplitudes were also decreased due to homeostatic pressure, whereas an afternoon nap could partially restore performance and neural activity. Conversely, the upward circadian rhythm in the afternoon exerted a compensatory effect, resulting in increases in N2 and P3 amplitudes. The findings highlight the disassociated impacts of homeostatic pressure and circadian rhythm on the cognitive processes involved in task-switching and further underscore the importance of considering diurnal variation in both scientific research and accident prevention.

Nickel-catalyzed direct stereoselective α-allylation of ketones with non-conjugated dienes.

The development of efficient and sustainable methods for the construction of carbon-carbon bonds with the simultaneous stereoselective generation of vicinal stereogenic centers is a longstanding goal in organic chemistry. Low-valent nickel(0) complexes which promote α-functionalization of carbonyls leveraging its pro-nucleophilic character in conjunction with suitable olefin acceptors are scarce. We report a Ni(0)NHC catalyst which selectively converts ketones and non-conjugated dienes to synthetically highly valuable α-allylated products. The catalyst directly activates the α-hydrogen atom of the carbonyl substrate transferring it to the olefin acceptor. The transformation creates adjacent quaternary and tertiary stereogenic centers in a highly diastereoselective and enantioselective manner. Computational studies indicate the ability of the Ni(0)NHC catalyst to trigger a ligand-to-ligand hydrogen transfer process from the ketone α-hydrogen atom to the olefin substrate, setting the selectivity of the process. The shown selective functionalization of the α-C-H bond of carbonyl groups by the Ni(0)NHC catalyst opens up new opportunities to exploit sustainable 3d-metal catalysis for a stereoselective access to valuable chiral building blocks.

Effect of homeostatic pressure and circadian arousal on the storage and executive components of working memory: Evidence from EEG power spectrum.

Diurnal fluctuations in working memory (WM) performance, characterized by task-specific peaks and troughs, are likely attributed to the differential regulation of WM subcomponents by interactions between circadian and homeostatic processes. The current study aimed to investigate the independent effects of circadian and homeostatic processes on the storage and executive subcomponents of WM. We assessed the change in frontal-midline theta (FMT) power supporting WM executive component and posterior alpha/beta power supporting WM storage during N-back tasks in the morning, midafternoon with and without a nap from 31 healthy adults. The results suggested that when the accumulated sleep homeostasis was alleviated in the midafternoon by a daytime nap, higher ACC, less number of omissions, and a stronger increase in FMT power from the no nap to nap conditions. Compared to the morning, a stronger decrease in posterior alpha power, and posterior beta power (only in the 3-back task), was observed in the no-nap condition because of circadian arousal regulation. These findings suggest that the circadian process primarily influences the storage aspect of WM supported by posterior alpha and beta activity, while sleep homeostasis has a greater impact on the execution aspect supported by FMT activity.

[Genetic analysis of two families with Short-rib thoracic dysplasia type 3].

OBJECTIVE: To explore the pathogenic variants and clinical classification of two fetuses with Short-rib thoracic dysplasia with or without polydactyly (SRTD). METHODS: With informed consent obtained, the phenotypic characteristics of the fetuses were comprehensively examined, and genomic DNA was extracted from fetal skin tissue and peripheral blood samples of the parents with conventional phenol-chloroform method. Whole exome sequencing (WES) was carried out on both fetuses, and the candidate variants were validated by Sanger sequencing. The pathogenicity of the candidate variants was analyzed using bioinformatic software VarCards, and the impact of the variants on the protein structure was predicted with Swiss-Pdb-viewer. RESULTS: Both fetuses were found to harbor compound heterozygous variants of the DYNC2H1 gene, including c.515C>A (p.Pro172Gln) and c.5983G>A (p.Ala1995Thr) in fetus 1, and c.5920G>T (pGly1974) and c.9908T>C (p.He3303Thr) in fetus 2. The parents of both fetuses were heterozygous carriers. CONCLUSION: The compound heterozygous variants of the DYNC2H1 gene probably underlay the SRTD3 in the two fetuses.

Soil Nutrients, Enzyme Activities, and Microbial Communities along a Chronosequence of Chinese Fir Plantations in Subtropical China.

Forests undergo a long-term development process from young to mature stages, yet the variations in soil nutrients, enzyme activities, microbial diversity, and community composition related to forest ages are still unclear. In this study, the characteristics of soil bacterial and fungal communities with their corresponding soil environmental factors in the young, middle, and mature stages (7, 15, and 25-year-old) of Chinese fir plantations (CFP) in the subtropical region of China were investigated in 2021. Results showed that the alpha diversity indices (Chao1 and Shannon) of soil bacteria and fungi were higher in 15 and 25-year-old stands than in 7-year-old stand of CFP, while the soil pH, soil water content, soil organic carbon, total nitrogen, total phosphorus, sucrase, urease, acid phosphatase, catalase, and microbial biomass carbon, nitrogen, and phosphorus showed higher in 7-year-old stand than other two stands of CFP. The nonmetric multidimensional scaling analysis revealed that the soil microbial species composition was significantly different in three stand ages of CFP. The redundancy and canonical correspondence analysis indicated that the soil urease and microbial biomass nitrogen were the main factors affecting soil bacterial and fungal species composition. Our findings suggested that soil microbial diversity and community structure were inconsistent with changes in soil nutrients and enzyme activities during CFP development, and enhancing stand nurturing and soil nutrient accumulation in the mid-development stage were beneficial to the sustainable management of CFP.

Identification of FCER1G as a key gene in multiple myeloma based on weighted gene co-expression network analysis.

PURPOSE: Although the prognosis of multiple myeloma (MM) has remarkably improved with the emerge of novel agents, it remains incurable and relapses inevitably. The molecular mechanisms of MM have not been well-studied. Herein, this study aimed to identify key genes in MM. MATERIALS AND METHODS: The GSE39754 dataset was used to screen differentially expressed genes (DEGs) and construct a co-expression network. Hub nodes were identified in the protein and protein interaction (PPI) network. Datasets GSE13591 and GSE2658 were used to validate hub genes. Moreover, function and gene set enrichment analyses were performed to elucidate the molecular pathogenesis of MM. RESULTS: In this study, 11 genes were found to be hub genes in the co-expression network, among which four genes (CD68, FCER1G, PLAUR and LCP2) were also identified as hub nodes. In the test dataset GSE13591, CD68 and FCER1G were significantly downregulated in MM. Besides, the areas under the curve (AUCs) of CD68 and FCER1G were greater than 0.8 in both the training dataset and the test dataset. Our results also confirmed that FCER1G highly expressed patients had remarkably longer survival times in MM. Function and pathway enrichment analyses suggested that hub genes were associated with epithelial mesenchymal transition, TNF-α signaling via NF-κB and inflammatory response. GSEA in our study indicated that FCER1G participated in NK cell mediated cytotoxicity and the NOD-like receptor signaling pathway. CONCLUSION: Our study identified FCER1G as a key gene in MM, providing a novel biomarker and potential molecular mechanisms of MM for further studies.

Clinical and genetic analysis in Chinese families with synpolydactyly, and cellular localization of HOXD13 with different length of polyalanine tract.

Synpolydactyly (SPD) is caused by mutations in the transcription factor gene HOXD13. Such mutations include polyalanine expansion (PAE), but further study is required for the phenotypic spectrum characteristics of HOXD13 PAE. We investigated four unrelated Chinese families with significant limb malformations. Three PAEs were found in the HOXD13 polyalanine coding region: c.172_192dup (p.Ala58_Ala64dup) in Family 1, c.169_192dup (p.Ala57_Ala64dup) in Family 2, and c.183_210dup (p.Ala62_Ala70dup) in Family 3 and Family 4. Interestingly, we identified a new manifestation of preaxial polydactyly in both hands in a pediatric patient with an expansion of seven alanines, a phenotype not previously noted in SPD patients. Comparing with the wild-type cells and mutant cells with polyalanine contractions (PACs), the HOXD13 protein with a PAE of nine-alanine or more was difficult to enter the nucleus, and easy to form inclusion bodies in the cytoplasm, and with the increase of PAE, the more inclusion bodies were formed. This study not only expanded the phenotypic spectrum of SPD, but also enriched our understanding of its pathogenic mechanisms.

CRISPR/Cas9 correction of a dominant cis-double-variant in COL1A1 isolated from a patient with osteogenesis imperfecta increases the osteogenic capacity of induced pluripotent stem cells.

Osteogenesis imperfecta (OI) is a hereditary skeletal disorder that is mainly caused by variants in COL1A1/2. So far, no specific treatment has been developed to correct its underlying etiology. We aimed to gain a better understanding of the pathological mechanisms of OI and develop gene therapies to correct OI-causing variants. A de novel cis-double-variant c.[175C>T; 187T>A] in COL1A1 was identified from a 5-year-old OI patient by whole-exome sequencing (WES). Three peptide nucleic acids (PNAs) were designed and then transfected patient-derived fibroblasts. PNA2 affected the translational strand and induced an optimal interfering effect at 0.25µM concentration, proved by Sanger sequencing, qPCR, Western blot, and immunostaining. Additionally, induced pluripotent stem cells (iPSCs) were cultured from patient-derived fibroblasts. Clones of iPSCs with c.187T>A variant and those with both variants largely restored their osteogenic capacities after CRISPR/Cas9 gene editing, which corrected the variants. Importantly, correcting c.187T>A variant alone in CRISPR-edited iPSCs was sufficient to alleviate OI phenotypes, as indicated by increased levels of COL1A1, COL1A2, ALP mRNAs, and COL1A1 protein. Our findings suggest that c.187T>A is the dominant variant of cis-double-variant in COL1A1 that led to OI, and PNA interference and CRISPR/Cas9 gene editing may be new therapeutic tools for OI treatment. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Identification of six novel variants from nine Chinese families with hypophosphatemic rickets.

BACKGROUND: Hypophosphatemic rickets (HR) is a rare genetic disorder associated with renal phosphate wasting and characterized by bone defects. Inactivating mutations in the phosphate regulating endopeptidase homolog X­linked gene (PHEX) account for most cases of HR. The aim of this study was to identify causative variants in nine unrelated Chinese families associated with HR, and to determine potential pathogenicity of the identified variants. METHODS: Genomic DNA was isolated from the peripheral blood of HR patients and their healthy relatives, followed by next-generation sequencing and/or Sanger sequencing. In silico prediction combined with conservation analysis was performed to assess the effects of the variants, and 3D protein modeling was conducted to predict the functional effects on the encoded protein. RESULTS: All HR patients recruited in this study displayed bone deformities and tooth agenesis, as well as reduced serum phosphate levels and elevated urine phosphate levels. Nine PHEX variants were identified in eight families, including four novel variants (c.1661_1726del, c.980A > G, c.1078A > T, and c.1017_1051dup). Of the nine identified PHEX variants, five caused a truncated protein, two caused an altered amino acid, and the other two were the canonical splicing variants. Novel variants c.1336G > A and c.1364 T > C in SLC34A3 were also found in one family. Conservation analysis showed that all the amino acids corresponding to the missense variants were highly conserved. In silico analysis and 3D protein structure modeling confirmed the pathogenicity of these variants. CONCLUSIONS: This study identified four novel variants in PHEX and two novel variants in SLC34A3 in a Chinese cohort with HR. Our findings highlight the dominant role of PHEX in HR, and expand the genotypic and phenotypic spectra of this disorder.

Identification of variants in ACAN and PAPSS2 leading to spondyloepi(meta)physeal dysplasias in four Chinese families.

BACKGROUND: Spondyloepi(meta)physeal dysplasias (SE[M]D) are a group of inherited skeletal disorders that mainly affect bone and cartilage, and next-generation sequencing has aided the detection of genetic defects of such diseases. In this study, we aimed to identify causative variants in four Chinese families associated with SE(M)D. METHODS: We recruited four unrelated Chinese families all displaying short stature and growth retardation. Clinical manifestations and X-ray imaging were recorded for all patients. Candidate variants were identified by whole-exome sequencing (WES) and verified by Sanger sequencing. Pathogenicity was assessed by conservation analysis, 3D protein modeling and in silico prediction, and was confirmed according to American College of Medical Genetics and Genomics. RESULTS: Three novel SE(M)D-related variants c.1090dupG, c.7168 T > G, and c.2947G > C in ACAN, and one reported variant c.712C > T in PAPSS2 were identified. Among them, c.1090dupG in ACAN and c.712C > T in PAPSS2 caused truncated protein and the other two variants led to amino acid alterations. Conservation analysis revealed sites of the two missense variants were highly conserved, and bioinformatic findings confirmed their pathogenicity. 3D modeling of mutant protein encoded by c.7168 T > G(p.Trp2390Gly) in ACAN proved the structural alteration in protein level. CONCLUSION: Our data suggested ACAN is a common pathogenic gene of SE(M)D. This study enriched the genetic background of skeletal dysplasias, and expanded the mutation spectra of ACAN and PAPSS2.

Photoinduced Arylation of Acridinium Salts: Tunable Photoredox Catalysts for C-O Bond Cleavage.

A photoinduced arylation of N-substituted acridinium salts has been developed and has exhibited a high functional group tolerance (e.g., halogen, nitrile, ketone, ester, and nitro). A broad range of well-decorated C9-arylated acridinium-based catalysts with fine-tuned photophysical and photochemical properties, namely, excited-state lifetimes and redox potentials have been synthetized in a one-step procedure. These functionalized acridinium salts were later evaluated in the photoredox-catalyzed fragmentation of 1,2-diol derivatives (lignin models). Among them, 2-bromophenyl substituted N-methyl acridinium has outperformed all photoredox catalysts, including commercial Fukuzumi's catalyst, for the selective CßO-Ar bond cleavage of diol monoarylethers to afford 1,2-diols in good yields.

Genotypic and Phenotypic Analysis in Chinese Cohort With Autosomal Recessive Osteogenesis Imperfecta.

Osteogenesis imperfecta (OI) is a rare heritable skeletal disorder which is mainly caused by defected type I collagen. Autosomal recessive OI (AR-OI) is caused by mutations of genes that are responsible for type I collagen modification and folding, and is often associated with more severe phenotypes. Due to the limited number of recessive OI patients, it has been difficult to study the mutation spectrum as well as the correlation of genotype and phenotype. This study recruited a Chinese cohort of 74 AR-OI families, aiming to establish the mutation spectrum and to examine the genotypic and phenotypic correlation. We identified 82 variants including 25 novel variants and 57 HGMD reported variants in these AR-OI patients, using whole exome sequencing/panel sequencing combined with Sanger sequencing. Pathogenic mutations were found at WNT1 (n = 30, 40.54%), SERPINF1 (n = 22, 29.73%), FKBP10 (n = 10, 13.51%), CRTAP (n = 3, 4.05%), P3H1 (n = 3, 4.05%), SERPINH1 (n = 2, 2.70%), SEC24D (n = 3, 4.05%), and PLOD2 (n = 1, 1.35%) respectively. Thus, WNT1 represents the most frequent pathogenic gene of AR-OI in Chinese population. The most common clinical manifestations of AR-OI patients include walking problem (72.86%), scoliosis (65.28%) and frequent fractures (fractures ≥2/year) (54.05%). Interestingly, ptosis represents a unique phenotype of patients carrying WNT1 variants, and it was rare in patients harboring other pathogenic genes. Our study expanded the mutation spectrum of AR-OI and enriched the knowledge of genotypic and phenotypic correlation in Chinese cohort with AR-OI.

Catalytic Enantioselective Functionalizations of C-H Bonds by Chiral Iridium Complexes.

The development of catalytic enantioselective transformations, enabling the construction of complex molecular scaffolds from simple precursors, has been a long-standing challenge in organic synthesis. Recent achievements in transition-metal catalyzed enantioselective functionalizations of carbon-hydrogen (C-H) bonds represent a promising pathway toward this goal. Over the last two decades, iridium catalysis has evolved as a valuable tool enabling the stereocontrolled synthesis of chiral molecules via C-H activation. The development of iridium-based systems with various chiral ligand classes, as well as studies of their reaction mechanisms, has resulted in dynamic progress in this area. This review aims to present a comprehensive picture of the enantioselective functionalizations of C-H bonds by chiral iridium complexes with emphasis on the mechanisms of the C-H activation step.

A review on the recent development of cyclodextrin-based materials used in oilfield applications.

Cyclodextrin has been studied for more than 120 years. Due to the special structure and selectivity of the cyclodextrin cavity, native cyclodextrins, cyclodextrin derivatives and inclusion complexes have received extensive attention and are widely used in medicine, food, the environment, cosmetics, chemical analysis, separation technology, catalysts and other fields. Cyclodextrin-based materials have been applied to oilfields since the early 1980s and performed well in enhancing the performance of petroleum chemicals and improving the efficiency of oil exploration. This review introduces the characteristics of cyclodextrin-based materials, including cyclodextrin polymers, inclusion complexes and new cyclodextrin nanocomposites, and summarizes their applications in oilfields for the first time, such as retarded acid solution, enhanced oil recovery, clay stabilizers, corrosion and scale inhibitors, wastewater treatment, etc. Meanwhile, the action mechanisms, major challenges and development tendencies of cyclodextrin-based materials in oilfields are also comprehensively discussed.

Increase in the Number of Bone Marrow Osteoclast Precursors at Different Skeletal Sites, Particularly in Long Bone and Jaw Marrow in Mice Lacking IL-1RA.

Recently, it was shown that interleukin-1ß (IL-1ß) has diverse stimulatory effects on different murine long bone marrow osteoclast precursors (OCPs) in vitro. In this study, interleukin-1 receptor antagonist deficient (Il1rn-/-) and wild-type (WT) mice were compared to investigate the effects of enhanced IL-1 signaling on the composition of OCPs in long bone, calvaria, vertebra, and jaw. Bone marrow cells were isolated from these sites and the percentage of early blast (CD31hi Ly-6C-), myeloid blast (CD31+ Ly-6C+), and monocyte (CD31- Ly-6Chi) OCPs was assessed by flow cytometry. At the time-point of cell isolation, Il1rn-/- mice showed no inflammation or bone destruction yet as determined by histology and microcomputed tomography. However, Il1rn-/- mice had an approximately two-fold higher percentage of OCPs in long bone and jaw marrow compared to WT. Conversely, vertebrae and calvaria marrow contained a similar composition of OCPs in both strains. Bone marrow cells were cultured with macrophage colony stimulating factor (M-CSF) and receptor of NfκB ligand (RANKL) on bone slices to assess osteoclastogenesis and on calcium phosphate-coated plates to analyze mineral dissolution. Deletion of Il1rn increased osteoclastogenesis from long bone, calvaria, and jaw marrows, and all Il1rn-/- cultures showed increased mineral dissolution compared to WT. However, osteoclast markers increased exclusively in Il1rn-/- osteoclasts from long bone and jaw. Collectively, these findings indicate that a lack of IL-1RA increases the numbers of OCPs in vivo, particularly in long bone and jaw, where rheumatoid arthritis and periodontitis develop. Thus, increased bone loss at these sites may be triggered by a larger pool of OCPs due to the disruption of IL-1 inhibitors.

Novel mutations identified in Chinese families with autosomal dominant congenital cataracts by targeted next-generation sequencing.

BACKGROUND: Congenital cataract is a clinically and genetically heterogeneous visual impairment. The aim of this study was to identify causative mutations in five unrelated Chinese families diagnosed with congenital cataracts. METHODS: Detailed family history and clinical data were collected, and ophthalmological examinations were performed using slit-lamp photography. Genomic DNA was extracted from peripheral blood of all available members. Thirty-eight genes associated with cataract were captured and sequenced in 5 typical nonsyndromic congenital cataract probands by targeted next-generation sequencing (NGS), and the results were confirmed by Sanger sequencing. Bioinformatics analysis was performed to predict the functional effect of mutant genes. RESULTS: Results from the DNA sequencing revealed five potential causative mutations: c.154 T > C(p.F52 L) in GJA8 of Family 1, c.1152_1153insG(p.S385Efs*83) in GJA3 of Family 2, c.1804 G > C(p.G602R) in BFSP1 of Family 3, c.1532C > T(p.T511 M) in EPHA2 of Family 4 and c.356G > A(p.R119H) in HSF4 of Family 5. These mutations co-segregated with all affected individuals in the families and were not found in unaffected family members nor in 50 controls. Bioinformatics analysis from several prediction tools supported the possible pathogenicity of these mutations. CONCLUSIONS: In this study, we identified five novel mutations (c.154 T > C in GJA8, c.1152_1153insG in GJA3, c.1804G > C in BFSP1, c.1532C > T in EPHA2, c.356G > A in HSF4) in five Chinese families with hereditary cataracts, respectively. NGS can be used as an effective tool for molecular diagnosis of genetically heterogeneous disorders such as congenital cataract, and the results can provide more effective clinical diagnosis and genetic counseling for the five families.