Concurrent of compound heterozygous variant of a novel in-frame deletion and the common hypomorphic haplotype in TBX6 and inherited 17q12 microdeletion in a fetus.

BACKGROUND: TBX6, a member of the T-box gene family, encodes the transcription factor box 6 that is critical for somite segmentation in vertebrates. It is known that the compound heterozygosity of disruptive variants in trans with a common hypomorphic risk haplotype (T-C-A) in the TBX6 gene contribute to 10% of congenital scoliosis (CS) cases. The deletion of chromosome 17q12 is a rare cytogenetic abnormality, which often leads to renal cysts and diabetes mellitus. However, the affected individuals often exhibit clinical heterogeneity and incomplete penetrance. METHODS: We here present a Chinese fetus who was shown to have CS by ultrasound examination at 17 weeks of gestation. Trio whole-exome sequencing (WES) was performed to investigate the underlying genetic defects of the fetus. In vitro functional experiments, including western-blotting and luciferase transactivation assay, were performed to determine the pathogenicity of the novel variant of TBX6. RESULTS: WES revealed the fetus harbored a compound heterozygous variant of c.338_340del (p.Ile113del) and the common hypomorphic risk haplotype of the TBX6 gene. In vitro functional study showed the p.Ile113del variant had no impact on TBX6 expression, but almost led to complete loss of its transcriptional activity. In addition, we identified a 1.85 Mb deletion on 17q12 region in the fetus and the mother. Though there is currently no clinical phenotype associated with this copy number variation in the fetus, it can explain multiple renal cysts in the pregnant woman. CONCLUSIONS: This study is the first to report a Chinese fetus with a single amino acid deletion variant and a T-C-A haplotype of TBX6. The clinical heterogeneity of 17q12 microdeletion poses significant challenges for prenatal genetic counseling. Our results once again suggest the complexity of prenatal genetic diagnosis.

The functional identification and evaluation of endophytic bacteria sourced from the roots of tolerant Achyranthes bidentata to overcome monoculture problems of Rehmannia glutinosa.

The isolation and identification of plant growth-promoting endophytic bacteria (PGPEB) from Achyranthes bidentata roots have profound theoretical and practical implications in ecological agriculture, particularly as bio-inoculants to address challenges associated with continuous monoculture. Our research revealed a significant increase in the abundance of these beneficial bacteria in A. bidentata rhizosphere soil under prolonged monoculture conditions, as shown by bioinformatics analysis. Subsequently, we isolated 563 strains of endophytic bacteria from A. bidentata roots. Functional characterization highlighted diverse plant growth-promoting traits among these bacteria, including the secretion of indole-3-acetic acid (IAA) ranging from 68.01 to 73.25 mg/L, phosphorus and potassium solubilization capacities, and antagonistic activity against pathogenic fungi (21.54%-50.81%). Through 16S rDNA sequencing, we identified nine strains exhibiting biocontrol and growth-promoting potential. Introduction of a synthetic microbial consortium (SMC) in pot experiments significantly increased root biomass by 48.19% in A. bidentata and 27.01% in replanted Rehmannia glutinosa. These findings provide innovative insights and strategies for addressing continuous cropping challenges, highlighting the practical promise of PGPEB from A. bidentata in ecological agriculture to overcome replanting obstacles for non-host plants like R. glutinosa, thereby promoting robust growth in medicinal plants.

Thinning alters nitrogen transformation processes in subtropical forest soil: Key roles of physicochemical properties.

Thinning-a widely used forest management practice-can significantly influence soil nitrogen (N) cycling processes in subtropical forests. However, the effects of different thinning intensities on nitrification, denitrification, and their relationships with soil properties and microbial communities remain poorly understood. Here, we conducted a study in a subtropical forest in China and applied three thinning treatments, i.e., no thinning (0 %), intermediate thinning (10-15 %), and heavy thinning (20-25 %), and investigated the effects of thinning intensity on the potential nitrification rate (PNR), potential denitrification rate (PDR), and microbial communities. Moreover, we explored the relationships among soil physicochemical properties, microbial community structure, and nitrogen transformation rates under different thinning intensities. Our results showed that intermediate and heavy thinning significantly increased the PNR by 87 % and 61 % and decreased the PDR by 31 % and 50 % compared to that of the control, respectively. Although the bacterial community structure was markedly influenced by thinning, the fungal community structure remained stable. Importantly, changes in microbial community composition and diversity had minimal impacts on the nitrogen transformation processes, whereas soil physicochemical properties, such as pH, organic carbon content, and nitrogen forms, were identified as the primary drivers. These findings highlight the critical role of managing soil physicochemical properties to regulate nitrogen transformations in forest soils. Effective forest management should focus on precisely adjusting the thinning intensity to enhance the soil physicochemical conditions, thereby promoting more efficient nitrogen cycling and improving forest ecosystem health in subtropical regions.

Copy number variation sequencing for the products of conception: What is the optimal testing strategy.

BACKGROUND: Copy number variation sequencing (CNV-seq) is crucial in prenatal diagnosis, but its limitations in detecting polyploidy, maternal cell contamination (MCC), and uniparental disomy (UPD) restrict its application in the analysis of products of conception (POCs). This study aimed to investigate an optimal genetic testing strategy for POCs in the era of CNV-seq. METHODS: CNV-seq and quantitative fluorescent polymerase chain reaction (QF-PCR) were performed in all 4,211 spontaneous miscarriage cases. Different testing strategies were compared and the optimal testing strategies were proposed. RESULTS: Of the 4,211 cases, 2561 (60.82%) exhibited clinically significant chromosomal abnormalities. CNV-seq alone, without QF-PCR, might misdiagnose 311 (7.39%) cases, including 278 polyploidy, 13 UPD, and 20 MCC. In 20 MCC cases identified by QF-PCR, CNV-seq successfully pinpointed the cause of miscarriage in 13 cases. Furthermore, in cases where QF-PCR suggested polyploidy, CNV-seq improved the diagnostic accuracy in 54 (1.28%) hypo/hypertriploidy cases. After comparing four different strategies, the sequential approach (initiating with CNV-seq followed by QF-PCR if necessary) emerged as advantageous, reducing approximately 70% of the cost associated with QF-PCR while maintaining result accuracy. CONCLUSIONS: We propose an initial CNV-seq followed by QF-PCR if needed-an efficient and cost-effective strategy for the genetic analysis of POCs.

Chemical Components, Nutritional Value, Volatile Organic Compounds and Biological Activities In Vitro of Coconut (Cocos nucifera L.) Water with Different Maturities.

The differences in chemical components, nutritional value, volatile organic compounds, antioxidant activity and α-glucosidase inhibiting capacity in vitro in coconut waters with different maturities (8, 10, and 12 months after pollination and germination height below 10 cm were named CW-8, CW-10, CW-2, and MCW, respectively) from the tall coconut variety were compared and analyzed. Results showed that as the maturity increased, the ash and reducing sugar in coconut water gradually decreased, while the protein content and fatty acids continued to increase. Potassium, phosphorus, and sodium in four coconut waters showed a trend of first increasing and then decreasing, and CW-12 had the highest content of 2133.85 mg/kg, 239.74 mg/kg, and 310.75 mg/kg, respectively. The volatile organic compounds (VOCs) present in higher amounts are alcohols and esters in coconut waters, among which 2-methylbutyl acetate, ethyl acetate monomer, and 2-methyl-1-propanol dimer were the characteristic volatile substances that distinguish MCW from the other three coconut waters. MCW has the best DPPH-scavenging and ferrous-ion-chelating ability (87.39% and 7.65%), while CW-8 had the highest hydroxyl and ABTS radicals scavenging rate (97.31% and 83.48%) and α-glucosidase inhibitory rate (81.36%). These results can provide support for the differential and high-value utilization of coconut water with different maturities.

C-to-G editing generates double-strand breaks causing deletion, transversion and translocation.

Base editors (BEs) introduce base substitutions without double-strand DNA cleavage. Besides precise substitutions, BEs generate low-frequency 'stochastic' byproducts through unclear mechanisms. Here, we performed in-depth outcome profiling and genetic dissection, revealing that C-to-G BEs (CGBEs) generate substantial amounts of intermediate double-strand breaks (DSBs), which are at the centre of several byproducts. Imperfect DSB end-joining leads to small deletions via end-resection, templated insertions or aberrant transversions during end fill-in. Chromosomal translocations were detected between the editing target and off-targets of Cas9/deaminase origin. Genetic screenings of DNA repair factors disclosed a central role of abasic site processing in DSB formation. Shielding of abasic sites by the suicide enzyme HMCES reduced CGBE-initiated DSBs, providing an effective way to minimize DSB-triggered events without affecting substitutions. This work demonstrates that CGBEs can initiate deleterious intermediate DSBs and therefore require careful consideration for therapeutic applications, and that HMCES-aided CGBEs hold promise as safer tools.

Improvement effect and mechanism of paeoniflorin on glucose metabolism，inflammation and oxidative stress in rats with gestational diabetes mellitus / 中国药房

OBJECTIVE To explore the effect of paeoniflorin on glucose metabolism， inflammation and oxidative stress in rats with gestational diabetes mellitus （GDM） and its potential mechanism based on nuclear factor-erythroid 2-related factor 2 （Nrf2）/ heme oxygenase-1 （HO-1）/nicotinamide adenine dinucleotide phosphate：quinone oxidoreductase 1 （NQO1） signaling pathway. METHODS The female rats fed with high fat and high sugar diet and the male rats fed with an ordinary diet were caged， the successfully conceived rats were collected， and streptozotocin was injected intraperitoneally once to induce the GDM model. The successfully modeled rats were randomly divided into the model group， metformin hydrochloride group （200 mg/kg metformin by gavage）， paeoniflorin low-， high-dose groups （45， 90 mg/kg paeoniflorin by gavage， respectively）， paeoniflorin+ML385 group （90 mg/kg paeoniflorin by gavage and intraperitoneal injection of 30 mg/kg Nrf2 inhibitor ML385）， with 12 rats in each group； in addition， another 12 conceived rats fed with an ordinary diet were selected as the control group. The rats in each drug group were given the corresponding drug/normal saline， once a day， for 2 consecutive weeks. Glucose metabolism indexes [fasting blood glucose （FBG）， fasting insulin （FINS）， insulin resistance index （HOMA-IR）]， serum inflammatory factors [interleukin-6 （IL-6）， tumor necrosis factor- α （TNF- α）] and renal tissue oxidative stress indexes [superoxide dismutase （SOD）， malondialdehyde （MDA）， glutathione peroxidase （GSH-Px）] were detected； the pathological changes of renal tissue were observed， and the protein expressions of Nrf2， HO-1 and NQO1 in renal tissue were detected. RESULTS Compared with the control group， the renal tissue lesions of the model group were obvious， including glomerular atrophy， edema degeneration of renal tubular epithelial cells and a large number of inflammatory cell infiltration； the levels of FBG and FINS， HOMA-IR， the levels of IL-6 and TNF-α in serum， and the level of MDA in renal tissue were significantly increased （P＜0.05）， while the levels of SOD and GSH-Px and the protein expressions of Nrf2， HO-1 and NQO1 in renal tissue were significantly decreased （P＜0.05）. Compared with the model group， the renal tissue lesions of rats in paeoniflorin low-dose and high-dose groups were reduced， the above quantitative indexes were significantly improved， and the improvement effect was better in high-dose group （P＜0.05）， while ML385 could significantly reverse the improvement effect of paeoniflorin on the above indexes （P＜0.05）. CONCLUSIONS Paeoniflorin can improve the abnormal glucose metabolism， inflammation and oxidative stress damage of renal tissue in GDM rats， which may be related to the activation of Nrf2/HO-1/NOQ1 signaling pathway.

Advances in Biochemical Sensor Devices Based on Two-dimensional Material Field Effect Transistors / 分析化学

Field effect transistor(FET)biochemical sensors show great potential in the fields of environmental monitoring,food safety,disease diagnosis and clinical treatment due to their low noise,low power consumption,label-free,easy integration and miniaturization characteristics.Two-dimensional(2D)materials,as a new generation of channel materials for FET biochemical sensors,have atomic-level thickness,high carrier mobility,high specific surface area and tunable bandgap,which can further improve the performance of FET biochemical sensors,extend their application areas,and promote the rapid development of FET biochemical sensors.This review focused on the development and latest progress of 2D material-based FET biochemical sensors,along with the challenges and prospects of 2D material-based FET biochemical sensors,which aimed to provide new device design conceptions and promote the further development of biochemical sensing technology.

Bi-allelic variants in CEP295 cause Seckel-like syndrome presenting with primary microcephaly, developmental delay, intellectual disability, short stature, craniofacial and digital abnormalities.

BACKGROUND: Pathogenic variants in the centrosome protein (CEP) family have been implicated in primary microcephaly, Seckel syndrome, and classical ciliopathies. However, most CEP genes remain unlinked to specific Mendelian genetic diseases in humans. We sought to explore the roles of CEP295 in human pathology. METHODS: Whole-exome sequencing was performed to screen for pathogenic variants in patients with severe microcephaly. Patient-derived fibroblasts and CEP295-depleted U2OS and RPE1 cells were used to clarify the underlying pathomechanisms, including centriole/centrosome development, cell cycle and proliferation changes, and ciliogenesis. Complementary experiments using CEP295 mRNA were performed to determine the pathogenicity of the identified missense variant. FINDINGS: Here, we report bi-allelic variants of CEP295 in four children from two unrelated families, characterized by severe primary microcephaly, short stature, developmental delay, intellectual disability, facial deformities, and abnormalities of fingers and toes, suggesting a Seckel-like syndrome. Mechanistically, depletion of CEP295 resulted in a decrease in the numbers of centrioles and centrosomes and triggered p53-dependent G1 cell cycle arrest. Moreover, loss of CEP295 causes extensive primary ciliary defects in both patient-derived fibroblasts and RPE1 cells. The results from complementary experiments revealed that the wild-type CEP295, but not the mutant protein, can correct the developmental defects of the centrosome/centriole and cilia in the patient-derived skin fibroblasts. INTERPRETATION: This study reports CEP295 as a causative gene of the syndromic microcephaly phenotype in humans. Our study also demonstrates that defects in CEP295 result in primary ciliary defects. FUNDING: A full list of funding bodies that contributed to this study can be found under "Acknowledgments."

Autosomal dominant neurodevelopmental disorders associated with KIF1A gene variants in 6 pediatric patients. / å ­ä¾‹KIF1AåŸºå› å˜å¼‚ç›¸å ³å¸¸æŸ“è‰²ä½“æ˜¾æ€§é—ä¼ ç¥žç»å‘è‚²éšœç¢æ‚£å„¿ä¸´åºŠå’Œé—ä¼ å­¦åˆ†æž.

OBJECTIVES: To analyze the clinical and genetic characteristics of children with autosomal dominant neurodevelopmental disorders caused by kinesin family member 1A (KIF1A) gene variation. METHODS: Clinical and genetic testing data of 6 children with KIF1A gene de novo heterozygous variation diagnosed in Shanghai Children's Medical Center Affiliated to Shanghai Jiao Tong University School of Medicine from the year 2018 to 2020 were retrospectively analyzed. Pathogenic variants were identified based on whole exome sequencing, and verified by Sanger sequencing. Moreover, the effect of variants on three-dimensional structure and stability of protein was analyzed by bioinformatics. RESULTS: Among 6 patients there were 4 males and 2 females, and the age of consultation varied from 7 months to 18 years. All cases had varying degrees of motor developmental delay since childhood, and 4 of them had gait abnormalities or fell easily. In addition, 2 children were accompanied by delayed mental development, epilepsy and abnormal eye development. Genetic tests showed that all 6 cases had heterozygous de novo variations of KIF1A gene, including 4 missense mutations c.296C>T (p.T99M), c.761G>A (p.R254Q), c.326G>T (p.G109V), c.745C>G (p.L249V) and one splicing mutation c.798+1G>A, among which the last three variants have not been previously reported. Bioinformatics analysis showed that G109V and L249V may impair their interaction with the neighboring amino acid residues, thereby impacting protein function and reducing protein stability, and were assessed as "likely pathogenic". Meanwhile, c.798+1G>A may damage an alpha helix in the motor domain of the KIF1A protein, and was assessed as "likely pathogenic". CONCLUSIONS: KIF1A-associated neurological diseases are clinically heterogeneous, with motor developmental delay and abnormal gait often being the most common clinical features. The clinical symptoms in T99M carriers are more severe, while those in R254Q carriers are relatively mild.

Impact of dust deposition on the growth of marine autotrophic and heterotrophic microorganisms: Evidence from the South China Sea.

Aeolian dust can provide nutrients for the ocean and affect the growth of phytoplankton. However, the impacts of dust deposition on autotrophic and heterotrophic microorganisms have rarely been studied. In this study, we conducted two microcosm experiments in the low-nutrient and low-chlorophyll environment of the South China Sea and found that dust did not stimulate the abundance of autotrophic and heterotrophic microorganisms. Our results show that dust contains most of the unreacted iron-bearing minerals, and thus provides limited bioavailable iron and nitrogen for bacterioplankton and phytoplankton growth. These results elucidate the overlooked impacts of the properties of the iron-bearing minerals in aeolian dust on microbial communities, which may play an important role in marine ecosystems and climate change.

Molecular and phenotypic spectrum of cardio-facio-cutaneous syndrome in Chinese patients.

BACKGROUND: Cardio-facio-cutaneous (CFC) syndrome is a RASopathy subtype that presents with unique craniofacial dysmorphology, congenital heart disease, dermatologic abnormalities, growth retardation, and intellectual disability. This study describes the phenotypic spectrum of CFC in China and its association with CFC syndrome gene variants. RESULTS: Twenty Chinese CFC patients, aged 0.6-9.5 years old, were included in this study and their clinical phenotypic spectrum was compared with that of 186 patients with CFC from non-Chinese ethnicities. All 20 Chinese patients with CFC carried de novo heterozygous BRAF, MAP2K1, and MAP2K2 variants. Two novel variants were detected and consistently predicted to be deleterious using bioinformatic tools. The clinical features of CFC in the Chinese patients included hypertrophic cardiomyopathy (2/20, 10%), pulmonary valve stenosis (2/20, 10%), curly or sparse hair (7/20, 35%), epilepsy (1/20, 5%), and hypotonia (10/20, 50%); these features were less frequently observed in Chinese patients than non-Chinese patients (p < 0.05). In contrast, feeding difficulties (19/20, 95%) were more frequently observed in the Chinese patients. Absent eyebrows and severe short stature were more common in patients with BRAF variants than in those with MAP2K1/2 variants. Facial recognition software was used to recognize most CFC patients using artificial intelligence. CONCLUSION: This study identified novel and common variants in our cohort of 20 Chinese patients with CFC. We uncovered differences in clinical features between Chinese and non-Chinese patients and detected genotype-phenotype correlations among the BRAF and MAP2K1/2 variant subgroups. This is the largest cohort of Chinese CFC patients to our knowledge, providing new insights into a subtype of RASopathy.

SINE-VNTR-Alu retrotransposon insertion as a novel mutational event underlying Glanzmann thrombasthenia.

BACKGROUND: Glanzmann thrombasthenia (GT) is an autosomal recessive platelet aggregation disorder caused by mutations in ITGA2B or ITGB3. OBJECTIVES: We aimed to assess the phenotype and investigate the genetic etiology of a GT pedigree. METHODS: A patient with bleeding manifestations and mild mental retardation was enrolled. Complete blood count, coagulation, and platelet aggregation tests were performed. Causal mutations were identified via whole exome and genome sequencing and subsequently confirmed through polymerase chain reaction and Sanger sequencing. The transcription of ITGB3 was characterized using RNA sequencing and reverse transcription polymerase chain reaction. The αⅡb and ß3 biosynthesis was investigated via whole blood flow cytometry and in vitro studies. RESULTS: GT was diagnosed in a patient with defective platelet aggregation. Novel compound heterozygous ITGB3 variants were identified, with a maternal nonsense mutation (c.2222G>A, p.Trp741∗) and a paternal SINE-VNTR-Alu (SVA) retrotransposon insertion. The 5' truncated SVA element was inserted in a sense orientation in intron 11 of ITGB3, resulting in aberrant splicing of ITGB3 and significantly reducing ß3 protein content. Meanwhile, both the expression and transportation of ß3 were damaged by the ITGB3 c.2222G>A. Almost no αⅡb and ß3 expressions were detected on the patient's platelets surface. CONCLUSION: Novel compound heterozygous ITGB3 mutations were identified in the GT pedigree, resulting in defects of αⅡbß3 biosynthesis. This is the first report of SVA retrotransposon insertion in the genetic pathogenesis of GT. Our study highlights the importance of combining multiple high-throughput sequencing technologies for the molecular diagnosis of genetic disorders.

Effects of grazing prohibition on nirK- and nirS-type denitrifier communities in salt marshes.

Introduction: Grazing prohibition is an effective management practice to restore salt marsh functioning. However, the effects of grazing exclusion on denitrifying microbial communities and their controlling factors in salt marshes remain unclear. Methods: In this study, we surveyed soil physicochemical properties and above- and below-ground biomass and using quantitative polymerase chain reaction and Illumina MiSeq high-throughput sequencing technology to determine the relative abundance, composition, and diversity of nitrite reductase nirS- and nirK-type denitrifying bacterial communities associated with grazing prohibition treatments and elevations. Results: The abundance of nirS-type denitrifiers increased with grazing prohibition time, whereas the abundance of nirK-type denitrifiers remained unaltered. Moreover, nirS-type denitrifiers were more abundant and diverse than nirK-type denitrifiers in all treatments. Grazing prohibition significantly altered the operational taxonomic unit richness, abundance-based coverage estimator, and Chao1 indices of the nirS-type denitrifying bacterial communities, whereas it only minimally affected the structure of the nirK-type denitrifying bacterial community. Discussion: The results imply that the nirS community, rather than nirK, should be the first candidate for use as an indicator in the process of salt marsh restoration after grazing prohibition. Substances of concern, total nitrogen, and salinity were the key environmental factors affecting the abundance and community composition of nirS and nirK denitrifiers. The findings of this study provide novel insights into the influence of the length of grazing prohibition and elevation on nirS- and nirK-type denitrifying bacterial community composition in salt marshes.

Environmental heterogeneity shapes the C and S cycling-associated microbial community in Haima's cold seeps.

Environmental heterogeneity in cold seeps is usually reflected by different faunal aggregates. The sediment microbiome, especially the geochemical cycling-associated communities, sustains the ecosystem through chemosynthesis. To date, few studies have paid attention to the structuring and functioning of geochemical cycling-associated communities relating to environmental heterogeneity in different faunal aggregates of cold seeps. In this study, we profiled the microbial community of four faunal aggregates in the Haima cold seep, South China Sea. Through a combination of geochemical and meta-omics approaches, we have found that geochemical variables, such as sulfate and calcium, exhibited a significant variation between different aggregates, indicating changes in the methane flux. Anaerobic methanotrophic archaea (ANME), sulfate-reducing, and sulfide-oxidizing bacteria (SRB and SOB) dominated the microbial community but varied in composition among the four aggregates. The diversity of archaea and bacteria exhibited a strong correlation between sulfate, calcium, and silicate. Interspecies co-exclusion inferred by molecular ecological network analysis increased from non-seep to clam aggregates and peaked at the mussel aggregate. The networked geochemical cycling-associated species showed an obvious aggregate-specific distribution pattern. Notably, hydrocarbon oxidation and sulfate reduction by ANME and SRB produced carbonate and sulfide, driving the alkalization of the sediment environment, which may impact the microbial communities. Collectively, these results highlighted that geofluid and microbial metabolism together resulted in environmental heterogeneity, which shaped the C and S cycling-associated microbial community.

Elevated nutrients and surface chlorophyll-α associated with natural methane seeps in the Haima cold seep area of the Qiongdongnan Basin, northern South China Sea.

Cold seeps are a significant source of methane to the ocean. However, nutrients and Chl-α in the euphotic layer overlying cold seeps have been poorly studied. Variations in Chl-α, nutrients, environmental parameters, and CH4 concentrations in the Haima cold seeps were analyzed. Results show that the overlying water exhibits a typical low nutrient and low Chl-α marine environment. Phosphate and Chl-α were significantly elevated, and the average SCM in cold seeps was much higher than that in control stations. Spearman correlation analysis indicated Chl-α in cold seep was positively correlated with salinity and negatively with nutrient and CH4 concentrations. It implied that the CH4 concentrations may promote the increase of Chl-α, and may be linked to CH4 plumes, bringing cold, nutrient-rich waters to the thermocline. However, due to the CH4 plumes hardly to track, more sampling is needed to determine the effects on Chl-α and phytoplankton in the euphotic layer.

Effects of qufeng xuanfei formula in guinea pig model of airway hyperergy.

This work aimed to clarify the potential regulating effects of Qufeng Xuanfei formula (QFXF) on airway neurogenic inflammation and its underlying target signal pathway. Guinea pig model of airway hyperergy (AHR) was used. The relative susceptibility of major proteins to airway neurogenic inflammation was assessed using Western blot immunoassay followed by being separated by SDS-PAGE. Compared to the model group, QFXF of all concentrations effectively depressed the capsaicin enhanced cough in guinea pigs and the peak values of airway resistance significantly decreased. The results illustrated that QFXF alleviated cough symptom in guinea pigs and reduced airway neurogenic inflammation when compared to AHR model group. Airway inflammation and damage, as well as the levels of NGF, SP and c-Fos in QFXF decreased the most in the high-dose group. The mechanism of antitussive activity may be associated with reducing airway inflammation. QFXF displayed effect on chronic cough through reducing the levels of neuropeptides, attenuating airway inflammation and promoting recovery from disease to decrease the airway neuro sensitivity, suggesting that the potential mechanism may be related to Ras/ERK/c-Fos pathway.

Complex clinical manifestations and new insights in RNA sequencing of children with diabetes and WFS1 variants.

Background: WFS1-related disorders involve a wide range of clinical phenotypes, including diabetes mellitus and neurodegeneration. Inheritance patterns of pathogenic variants of this gene can be autosomal recessive or dominant, and differences in penetrance present challenges for accurate diagnosis and genetic counselling. Methods: Three probands and one elder brother from three families were systematically evaluated and the clinical data of other family members were collected from the medical history. Whole-exome sequencing was performed on the probands, and RNA sequencing was performed on four patients, their parents with WFS1 variants, and four gender- and age-matched children with type 1 diabetes mellitus. Results: There were six patients with diabetes. Dilated cardiomyopathy, a rare manifestation of WFS1-related disease, was identified in one patient, along with MRI findings of brain atrophy at age 7 years and 3 months, the earliest age of discovery we know of. Whole-exome sequencing revealed five pathogenic or likely pathogenic variants in the WFS1 gene, including c.1348dupC (p.His450Profs*93), c.1381A>C (p.Thr461pro), c.1329C>G (p.Ser443Arg), c.2081delA (p.Glu694Glyfs*16), c.1350-1356delinsGCA (p.His450Glnfs*26), of which 3 variants (c.1348dupC, c.2081delA, c.1350-1356delinsGCA) were novel that have not been previously reported. The differentially expressed genes were mainly associated with immune-related pathways according to the Gene Ontology enrichment analysis of the RNA sequencing data. The exon 1 region of HLA-DRB1 in two patients was not transcribed, while the transcription of the region in their parents was normal. Conclusion: This study emphasizes the clinical and genetic heterogeneity in patients, even in the same family with WFS1 variants. MRI evaluation of the brain should be considered when WFS1-related disorder is first diagnosed.

Defective Joint Development and Maintenance in GDF6-Related Multiple Synostoses Syndrome.

Multiple synostoses syndromes (SYNS) are a group of rare genetic bone disorders characterized by multiple joint fusions. We previously reported an SYNS4-causing GDF6 c.1330 T > A (p.Tyr444Asn) mutation, which reduced Noggin-induced GDF6 inhibition and enhanced SMAD1/5/8 signaling. However, the mechanisms by which GDF6 gain-of-function mutation alters joint formation and the comprehensive molecular portraits of SYNS4 remain unclear. Herein, we introduce the p.Tyr443Asn (orthologous to the human GDF6 p.Tyr444Asn) mutation into the mouse Gdf6 locus and report the results of extensive phenotype analysis, joint development investigation, and transcriptome profiling of Gdf6 p.Tyr443Asn limb buds. Gdf6 p.Tyr443Asn knock-in mice recapitulated the morphological features of human SYNS4, showing joint fusion in the wrists, ankles, phalanges, and auditory ossicles. Analysis of mouse embryonic forelimbs demonstrated joint interzone formation defects and excess chondrogenesis in Gdf6 p.Tyr443Asn knock-in mice. Further, RNA sequencing of forelimb buds revealed enhanced bone formation and upregulated bone morphogenetic protein (BMP) signaling in mice carrying the Gdf6 p.Tyr443Asn mutation. Because tightly regulated BMP signaling is critical for skeletal development and joint morphogenesis, our study shows that enhancing GDF6 activity has a significant impact on both prenatal joint development and postnatal joint maintenance. © 2023 American Society for Bone and Mineral Research (ASBMR).

Two patients with KDM3B variants and new presentations of Diets-Jongmans syndrome.

KDM3B is located on chromosome 5q31 and encodes KDM3B, which is involved in histone demethylation and epigenetic regulation. Pathogenic KDM3B variants cause a dominantly inherited disorder presenting with intellectual disability (ID), short stature, and facial dysmorphism, named Diets-Jongmans syndrome. We describe two patients with KDM3B variants presenting with Diets-Jongmans syndrome. Genetic testing was performed because of the clinical data and a lack of a clear diagnosis in both patients. Candidate variants were verified by Sanger sequencing. After KDM3B variants were detected, in silico tools were used to predict the pathogenicity of the missense variants. A minigene assay was performed to evaluate the splicing effects of the c.5070 + 1G > A variant on KDM3B. Patient 1 mainly presented with repetitive upper respiratory tract infection and patient 2 presented with palpitation, shortness of breath, and pitting edema; both had ID. Whole exome sequencing identified variants of KDM3B. Patient 1 had the de novo KDM3B c.5070 + 1G > A variant, whereas patient 2 had the c.2828G > A (p.R943Q) variant. Transcriptional experiments of the splicing variant c.5070 + 1G > A revealed aberrant transcripts leading to truncated protein products. We found two pathogenic variants in KDM3B, one of which is novel. Both patients had additional clinical presentations, and patient 1 had transient neutropenia. KDM3B c.5070 + 1G > A is the first KDM3B splice-site variant and was identified as a germline variant. Neutropenia and cardiomyopathy are newly found presentations of Diets-Jongmans syndrome. Our report enriches our knowledge of the genotypic spectrum of the KDM3B variants and phenotypic diversity of Diets-Jongmans syndrome.