[Clinical, genetic, and pathological analysis in 165 children with disorders of sex development]. / 165ä¾‹å„¿ç«¥æ€§å‘è‚²å¼‚å¸¸ç–¾ç— çš„ä¸´åºŠã€é—ä¼ å­¦ä¸Žç— ç†å­¦åˆ†æž.

OBJECTIVES: To investigate the clinical phenotypes, genetic characteristics, and pathological features of children with disorders of sex development (DSD). METHODS: A retrospective analysis was conducted on epidemiological, clinical phenotype, chromosomal karyotype, gonadal pathology, and genotype data of 165 hospitalized children with DSD at Children's Hospital of Hebei Province and Tangshan Maternal and Child Health Hospital from August 2008 to December 2022. RESULTS: Among the 165 children with DSD, common presenting symptoms were short stature (62/165, 37.6%), clitoromegaly (33/165, 20.0%), cryptorchidism (28/165, 17.0%), hypospadias (24/165, 14.5%), and skin pigmentation abnormalities/exteriorized pigmented labia majora (19/165, 11.5%). Chromosomal karyotype analysis was performed on 127 cases, revealing 36 cases (28.3%) of 46,XX DSD, 34 cases (26.8%) of 46,XY DSD, and 57 cases (44.9%) of sex chromosome abnormalities. Among the sex chromosome abnormal karyotypes, the 45,X karyotype (11/57, 19%) and 45,X/other karyotype mosaicism (36/57, 63%) were more common. Sixteen children underwent histopathological biopsy of gonadal tissues, resulting in retrieval of 25 gonadal tissues. The gonadal tissue biopsies revealed 3 cases of testes, 3 cases of dysplastic testes, 6 cases of ovaries, 11 cases of ovotestes, and 1 case each of streak gonad and agenesis of gonads. Genetic testing identified pathogenic/likely pathogenic variants in 23 cases (23/36, 64%), including 12 cases of 21-hydroxylase deficiency congenital adrenal hyperplasia caused by CYP21A2 pathogenic variants. CONCLUSIONS: Short stature, clitoromegaly, cryptorchidism, hypospadias, and skin pigmentation abnormalities are common phenotypes in children with DSD. 45,X/other karyotype mosaicism and CYP21A2 compound heterozygous variants are major etiological factors in children with DSD. The most commonly observed gonadal histopathology in children with DSD includes ovotestes, ovaries, and testes/dysgenetic testes.

GADD45α alleviates the CDDP resistance of SK-OV3/cddp cells via redox-mediated DNA damage.

Epithelial ovarian cancer has the highest mortality rate of all malignant ovarian cancer types. Great progress has been made in the treatment of ovarian cancer in recent years. However, drug resistance has led to a low level of 5-year survival rate of epithelial ovarian cancer, and the molecular mechanism of which remains unknown. The aim of the present study was to identify the role of redox status in the cisplatin (CDDP) resistance of ovarian cancer. CDDP-resistant SK-OV3 (SK-OV3/cddp) cells were prepared and their reactive oxygen species and glutathione levels were investigated. The effects of hydrogen peroxide on the CDDP sensitivity of the SK-OV3/cddp cells and their expression levels of the redox-associated protein growth arrest and DNA damage 45a (GADD45α) were also investigated. In addition, the impact of GADD45α overexpression on cell viability was evaluated in vitro and in vivo, and the levels of Ser-139 phosphorylated H2A histone family member X (γ-H2AX), which is associated with DNA damage, were detected. The results suggested that redox status affected the drug resistance of the ovarian cancer cells by increasing the expression of GADD45α. The overexpression of GADD45α reversed the CDDP resistance of the SK-OV3/cddp cells and increased the level of γ-H2AX. In conclusion, GADD45α alleviated the CDDP resistance of SK-OV3/cddp cells via the induction of redox-mediated DNA damage.

Dexmedetomidine Reduces the Lidocaine-Induced Neurotoxicity by Inhibiting Inflammasome Activation and Reducing Pyroptosis in Rats.

Local anesthetic toxicity is closely related to neuronal death and activation of the inflammatory response. Dexmedetomidine (Dex) is an adrenergic α2 receptor agonist that can reduce the neurotoxicity induced by lidocaine. It also has anti-inflammatory effects. However, the mechanism underlying the neuroprotective effects of Dex against lidocaine-induced toxicity remains to be defined. We hypothesized that Dex exerts its neural protective effect through inhibiting inflammasome activation and through anti-pyroptosis effects against local anesthetic-induced nerve injury. In a rat model of lidocaine-induced spinal cord injury, we studied the protective effect of Dex on lidocaine-induced changes in spinal cord function, inflammasome formation and pyroptosis, pro-inflammatory cytokine expression, and protein kinase C (PKC)-δ phosphorylation. Dex reduced lidocaine-induced neurotoxicity and inhibited PKC-δ phosphorylation in the spinal cord of rats. Furthermore, Dex inhibited pyroptosis and inflammasome formation (caspase-1, NLRP3, and apoptosis-associated speck-like protein (ASC)). Finally, Dex attenuated interleukin (IL)-1ß and IL-18 expression, as well as microglia response. In conclusion, Dex can reduce the severity of lidocaine-induced spinal cord injury in rats by inhibiting priming and inflammasome activation and reducing pyroptosis via PKC-δ phosphorylation.

[The NMD escape mechanism and its application in disease therapy].

Nonsense-mediated mRNA decay (NMD) refers to the degradation of mRNA due to the presence of premature stop codon (PTC) on mRNA under pathological or physiological conditions. NMD is widely considered an mRNA-specific quality control process. Recently it was discovered that some PTCs do not trigger NMD in a variety of diseases - a process known as NMD escape; however, its exact mechanism remains unclear. At present, there are two widely accepted mechanistic hypotheses during NMD escape. The first is PTC read-through, in which protein translation undergoes PTC until the normal stop codon is encountered, producing a full-length protein. The second is translation reinitiation, in which protein translation recommences at the potential start codon downstream of PTC and terminates at the stop codon, producing an N-terminal truncated protein. Currently, an increasing number of drugs or small molecules that use PTC read-through have been successfully applied to treat nonsense variation-associated diseases. In this review, we summarize the NMD mechanism and discuss the application and progress in our understanding of NMD escape in disease therapy. This review should provide a useful framework to advance current understanding of the research and application of NMD escape.

Diagnosis of Spinal Muscular Atrophy: A Simple Method for Quantifying the Relative Amount of Survival Motor Neuron Gene 1/2 Using Sanger DNA Sequencing.

BACKGROUND: Spinal muscular atrophy (SMA) is caused by homozygous deletion or compound heterozygous mutation of survival motor neuron gene 1 (SMN1), which is the key to diagnose SMA. The study was to establish and evaluate a new diagnostic method for SMA. METHODS: A total of 1494 children suspected with SMA were enrolled in this study. Traditional strategy, including multiplexed ligation-dependent probe amplification (MLPA) and TA cloning, was used in 1364 suspected SMA children from 2003 to 2014, and the 130 suspected SMA children were tested by a new strategy from 2015 to 2016, who were also verified by MLPA combined with TA cloning. The SMN1 and SMN2 were simultaneously amplified by polymerase chain reaction using the same primers. Mutation Surveyor software was used to detect and quantify the SMN1 variants by calculating allelic proportions in Sanger sequencing. Finally, turnaround time and cost of these two strategies were compared. RESULTS: Among 1364 suspected SMA children, 576 children had SMN1 homozygous deletion and 27 children had SMN1 compound heterozygous mutation. Among the 130 cases, 59 had SMN1 homozygous deletion and 8 had heterozygous deletion: the SMN1-specific peak proportion on exon 7 was 34.6 ± 1.0% and 25.5 ± 0.5%, representing SMN1:SMN2 to be 1:2 and 1:3, respectively. Moreover, five variations, including p.Ser8Lysfs *23 (in two cases), p.Leu228*, p.Pro218Hisfs *26, p.Ser143Phefs*5, and p.Tyr276His, were detected in 6/8 cases with heterozygous deletion, the mutant allele proportion was 31.9%, 23.9%, 37.6%, 32.8%, 24.5%, and 23.6%, which was similar to that of the SMN1-specific site on exon 7, suggesting that those subtle mutations were located in SMN1. All these results were consistent with MLPA and TA cloning. The turnaround times of two strategies were 7.5 h and 266.5 h, respectively. Cost of a new strategy was only 28.5% of the traditional strategy. CONCLUSION: Sanger sequencing combined with Mutation Surveyor analysis has potential application in SMA diagnosis.

Flavonoid biosynthesis controls fiber color in naturally colored cotton.

The existence of only natural brown and green cotton fibers (BCF and GCF, respectively), as well as poor fiber quality, limits the use of naturally colored cotton (Gossypium hirsutum L.). A better understanding of fiber pigment regulation is needed to surmount these obstacles. In this work, transcriptome analysis and quantitative reverse transcription PCR revealed that 13 and 9 phenylpropanoid (metabolic) pathway genes were enriched during pigment synthesis, while the differential expression of phenylpropanoid (metabolic) and flavonoid metabolic pathway genes occurred among BCF, GCF, and white cotton fibers (WCF). Silencing the chalcone flavanone isomerase gene in a BCF line resulted in three fiber phenotypes among offspring of the RNAi lines: BCF, almost WCF, and GCF. The lines with almost WCF suppressed chalcone flavanone isomerase, while the lines with GCF highly expressed the glucosyl transferase (3GT) gene. Overexpression of the Gh3GT or Arabidopsis thaliana 3GT gene in BCF lines resulted in GCF. Additionally, the phenylpropanoid and flavonoid metabolites of BCF and GCF were significantly higher than those of WCF as assessed by a metabolomics analysis. Thus, the flavonoid biosynthetic pathway controls both brown and green pigmentation processes. Like natural colored fibers, the transgenic colored fibers were weaker and shorter than WCF. This study shows the potential of flavonoid pathway modifications to alter cotton fibers' color and quality.

p.Val19Glyfs*21 and p.Leu228* variants in the survival of motor neuron 1 trigger nonsense-mediated mRNA decay causing the SMN1 PTC+ transcripts degradation.

Spinal Muscular Atrophy (SMA) results from loss-of-function mutations in the survival of motor neuron 1 (SMN1) gene. Our previous research showed that 40% of variants were nonsense or frameshift variants and SMN1 mRNA levels in the patients carrying these variants were significantly decreased. Here we selected one rare variant (p.Val19Glyfs*21) and one common variant (p.Leu228*) to explore the degradation mechanism of mutant transcripts. The levels of full-length (FL)-SMN1 transcripts and SMN protein in the cell lines from the patients with these variants were both significantly reduced (p<0.01). Treatment with two translation inhibitors (puromycin and Cycloheximide (CHX)) markedly increased the levels of FL-SMN1 transcripts with premature translation termination codons (PTCs) (p<0.01) and showed time-dependent (10h>5.5h) but not dose-dependent effects. Moreover, the knockdown of UPF1, a key factor in nonsense-mediated mRNA decay (NMD) by lentivirus, led to a 3.1-fold increase (p<0.01) in FL-SMN1 transcript levels in patient fibroblasts. Our research provides evidence that these two PTC-generating variants (p.Val19Glyfs*21 and p.Leu228*) can trigger NMD, causing rapid degradation of SMN1 transcripts thereby resulting in SMN protein deficiency. These two variants are highly pathogenic and are associated with more severe SMA phenotypes. Varying NMD efficiency after treatment with puromycin and CHX in different cell types was also observed.

Tuning of nanofocused vector vortex beam of metallic granary-shaped nanotip with spin-dependent dielectric helical cone.

We present the combined configuration of dielectric helical cone and metallic granary-shaped nanotip to produce three -dimensional vector vortex nanofocused optical field. The intensity and phase of the electric fields, and Povnting vector of the optical field generated by the combined configuration with linearly polarized illumination are studied with three-dimensional finite difference time-domain method. The localized vector electric field near the apex of the metallic granary-shaped nanotip is strongly depended on the chirality of the dielectric helical cone and the bottom radius of the metallic granary-shaped nanotip. The localized vector electric field is wavelength selective with the maximum intensity enhancement up to 104 times and minimum size of about 900 nm2, and the maximum radial electric field rotates 67.0° along z axis. This indicates the vector vortex beam generated by the combined configuration can be applied in nanofabrication, nano-sensing and nano-manipulation.

Role of Diffusion-weighted and Contrast-enhanced Magnetic Resonance Imaging in Differentiating Activity of Ankylosing Spondylitis.

BACKGROUND: Previous studies showed that combining apparent diffusion coefficient (ADC) value with the Spondyloarthritis Research Consortium of Canada (SPARCC) index value might provide a reliable evaluation of the activity of ankylosing spondylitis (AS), and that contrast-enhanced (CE) magnetic resonance imaging (MRI) is unnecessary. However, the results were based on confirming only a small random sample. This study aimed to assess the role of CE-MRI in differentiating the disease activity of AS by comparing ADC value with a large sample. METHODS: A total of 115 patients with AS were enrolled in accordance with Bath AS Disease Activity Index and laboratory indices, and 115 patients were divided into two groups, including active group (n = 69) and inactive group (n = 46). SPARCC, ΔSI, and ADC values were obtained from the short tau inversion recovery (STIR), diffusion-weighted imaging (DWI), and CE-MRI, respectively. One-way analysis of variance and receiver operating characteristic analysis were performed for all parameters. RESULTS: The optimal cutoff values (with sensitivity, specificity, respective area under the curve, positive likelihood ratio, and negative likelihood ratio) for the differentiation between active and inactive groups are as follows: SPARCC = 6 (72.06%, 82.61%, 0.836, 4.14, 0.34); ΔSI (%) = 153 (80.6%, 84.78%, 0.819, 5.3, 0.23); ADC value = 1.15 × 10-3 mm2/s (72.73%, 81.82%, 0.786, 4, 0.33). No statistical differences were found among the predictive values of SPARCC, ΔSI, and ADC. Multivariate analysis showed no significant difference between the combination of SPARCC and ADC values with and without ΔSI. CONCLUSIONS: Using large sample, we concluded that the combination of STIR and DWI would play significant roles in assessing the disease activity, and CE-MRI sequence is not routinely used in imaging of AS to avoid renal fibrosis and aggravation of kidney disease.