20-HETE downregulates Na/K-ATPase α1 expression via the ubiquitination pathway.

In this article, we found that 20-Hydroxyeicosatetraenoic acid (20-HETE) reduced Na/K-ATPase α1 expression via the ubiquitin-proteasome pathway. The ubiquitination level of Na/K-ATPase α1 protein was increased in 20-HETE-treated mouse cortical collecting duct cells and the kidney tissues of CYP4F2 transgenic mice. We also demonstrated that 20-HETE-induced high level phosphorylation of Na/K-ATPase α1 was necessary for its ubiquitination.The protein kinase C inhibitor sotrastaurin significantly reduced the phosphorylation of Na/K-ATPase α1 and increased the expression of Na/K-ATPase α1 although 20-HETE stimulus being applied at the same time. Moreover, high level of 20-HETE increased the expression and neddylation of Cullin3,which is an important ubiquitin E3 ligase in kidney. MLN4924, an inhibitor of NEDD8-activating enzyme, inhibited neddylation of Cullin3 and reversed the reduction of Na/K-ATPase α1 expression caused by 20-HETE. Thus, 20-HETE downregulates Na/K-ATPase α1 via the ubiquitination pathway, and phosphorylation of Na/K-ATPase α1 is a prerequisite to ubiquitination. Additionally, 20-HETE regulates Cullin3 expression via neddylation.

Projected patterns of land uses in Africa under a warming climate.

Land-use change is a direct driver of biodiversity loss, projection and future land use change often consider a topical issue in response to climate change. Yet few studies have projected land-use changes over Africa, owing to large uncertainties. We project changes in land-use and land-use transfer under future climate for three specified time periods: 2021-2040, 2041-2060, and 2081-2100, and compares the performance of various scenarios using observational land-use data for the year 2020 and projected land-use under seven Shared Socioeconomic Pathways Scenarios (SSP): SSP1-1.9, SSP1-2.6, SSP2-4.5, SSP3-7.0, SSP4-3.4, SSP4-6.0 and SSP5-8.5 from 2015 to 2100 in Africa. The observational land-use types for the year 2020 depict a change and show linear relationship between observational and simulated land-use with a strong correlation of 0.89 (P < 0.01) over Africa. Relative to the reference period (1995-2014), for (2021-2040), (2041-2060), (2081-2100), barren land and forest land are projected to decrease by an average of (6%, 11%, 16%), (9%, 19%, 38%) respectively, while, crop land, grassland and urban land area are projected to increase by (36%, 58%, and 105%), (4%, 7% and 11%), and (139%, 275% and 450%) respectively. Results show a substantial variations of land use transfer between scenarios with major from barren land to crop land, for the whole future period (2015-2100). Although SSP4-3.4 project the least transfer. Population and GDP show a relationship with cropland and barren land. The greatest conversion of barren land to crop land could endanger biodiversity and have negative effects on how well the African continent's ecosystem's function.

Prenatal whole-exome sequencing for fetal structural anomalies: a retrospective analysis of 145 Chinese cases.

BACKGROUND: Whole-exome sequencing (WES) significantly improves the diagnosis of the etiology of fetal structural anomalies. This study aims to evaluate the diagnostic value of prenatal WES and to investigate the pathogenic variants in structurally abnormal fetuses. METHODS: We recruited 144 fetuses with structural anomalies between 14 and 2020 and 15 December 2021 in the study. Genetic screening was performed by WES combined with karyotyping and chromosomal microarray analysis. The molecular diagnostic yield of prenatal WES for each type of fetal structural anomaly and the identified pathogenic genes and mutations were reported. RESULTS: In this study, we retrospectively analyzed the clinical and genetic data of 145 structurally anomalous fetuses. These cases were classified into 9 phenotypic classes based on antenatal ultrasound findings. Thirty-eight pathogenic variants in 24 genes were identified in 35 of the 145 cases, including 14 novel variants in 13 genes (EP300, MYH3, TSC2, MMP9, CPLANE1, INVS, COL1A1, EYA1, TTC21B, MKS1, COL11A2, PDHA1 and L1CAM). Five additional pathogenic variants were classified as incidental findings. Our study showed that the overall diagnosis rate of WES was 28.1% (27/96) in the parent-fetus trio cases and 16.3% (8/49) in the proband-only cases. Fetuses with musculoskeletal anomalies had the highest diagnostic yield (51.4%, 19/37). In addition, FGFR3 and COL1A1 were the most common pathogenic genes. CONCLUSIONS: Our work expands the mutation spectrum of the genes associated with fetal structural anomalies and provides valuable information for future parental genetic counselling and pregnancy management of the structurally anomalous fetuses.

Identification of a novel 10.3 kb deletion causing α0-thalassemia by third-generation sequencing: Pedigree analysis and genetic diagnosis.

BACKGROUND: α-thalassemia is an inherited blood disorder caused by variants in the α-globin gene cluster. Identification of the pathogenic α-globin gene variants is important for the diagnosis and management of thalassemia. METHODS: Two suspected families from Xiantao, Hubei Province were recruited in this study. The family members underwent hemoglobin testing. Polymerase Chain Reaction based reverse dot blot (PCR-RDB) was employed to identify the known variants. Next-generation sequencing (NGS) and third-generation sequencing (TGS) were performed to screen the potential disease-causing variants, which were validated by Sanger sequencing and multiplex ligation-dependent probe amplification (MLPA). RESULTS: Hematological analysis suggested that proband A had α-thalassemia traits, and proband B had HbH disease traits. However, only a -α3.7 mutation had been detected by PCR-RDB and NGS in the proband of family B. Subsequent TGS identified a novel 10.3 kb deletion (NC_000016.10:g.172342-182690del) covering the HBA1, HBQ1 and HBA2 genes in the α-globin gene cluster in both family A and B, which was confirmed by Sanger sequencing and MLPA. These results indicated that the novel deletion is likely responsible for α-thalassemia. CONCLUSION: A novel α-thalassemia deletion was identified for the two families by TGS. Our work broadened the molecular spectrum of α-thalassemia, and was beneficial for the diagnosis, genetic counseling and management of α-thalassemia.

The prevalence and outcomes of α- and ß-thalassemia among pregnant women in Hubei Province, Central China: An observational study.

ABSTRACT: There is no information concerning the prevalence of thalassemia among pregnant women in Hubei Province currently. This study is aimed to explore the prevalence of α- and ß-thalassemia genotypes among pregnant women in Hubei Province, and to explore the clinically applicable screening approach, as well as to investigate the pregnancy outcomes of α- and ß-thalassemia carriers.Pregnant participants were recruited from 4 hospitals for the screening of α- and ß-thalassemia mutations in Hubei Province. Polymerase Chain Reaction and flow cytometry methods were used to examine α- and ß-thalassemia mutations. The hematological parameters and pregnancy outcomes of α- and ß-thalassemia carriers were obtained from the hospital information system. The chi-square tests were used to evaluate the difference in hematological parameters between pregnant thalassemia carriers and the control group.Among 11,875 participants, 414 (3.49%) were confirmed with α-thalassemia carriers, 228 (1.92%) were confirmed with ß-thalassemia carriers, and 3 (0.03%) were confirmed with both α- and ß-thalassemia carriers. The frequency of -α3.7 accounted for 2.05% and it was the most frequent genotype of α-thalassemia; the proportion of IVS-II-654 was 0.85% and it was the most frequent genotype of ß-thalassemia in Hubei Province. Furthermore, the proportion of patients with low mean corpuscular volume (MCV) or mean cell hemoglobin (MCH) values was accounted for 36.64% and 93.97% among α-thalassemia and ß-thalassemia carriers, respectively. And participants with normal MCV and MCH values were accounted for 95.07% among non-thalassemia participants. High prevalence of pregnancy-induced diabetes (16.97%), preterm birth (9.96%), pregnancy-induced hypertension (8.12%), and low birth weight (5.90%) were observed among pregnant thalassemia carriers.MCV and MCH values were suggested to apply on the preliminary screening of pregnant ß-thalassemia; however, it's unpractical on that of α-thalassemia. Furthermore, thalassemia carriers might have a high risk of negative pregnancy outcomes. These findings could be useful for the preliminary screening of thalassemia and perinatal care for the pregnant thalassemia carriers.

[Genetic Mutation Characteristics of Glucose-6-Phosphate Dehydrogenase Deficiency Patients in Wuhan].

OBJECTIVE: To explore the genotype mutation characteristics of patients with glucose-6-phosphate dehydrogenase(G6PD) deficiency in Wuhan. METHODS: A total of 1 321 neonates with positive screening and outpatients were received G6PD mutation detection, 12 kinds of common G6PD mutation in Chinese people was detected by using multicolor melting curve analysis (MMCA) method, for those with negative results, the enzyme activity and clinical information were analyzed, sequencing was recommended after informed consent when it is necessary. RESULTS: Among 1321 patients, a total of 768 mutations were detected out, with a detection rate of 58.1%. A total of 18 types of G6PD genotypes were identified, including c.1388G>A, c.1376G>T, c.95G>A, c.1024C>T, c.871G>A, c.392G>T, c.487G>A, c.1360C>T, c.1004C>A, c.517T>C, c.592C>T, c.94C>G, c.152C>T, c.320A>G, c.1028A>G, c.1316G>A, c.1327G>C and c.1376G>C, including 683 male hemizygotes, 3 female homozygotes, 80 female heterozygotes and 2 female compound heterozygous. CONCLUSION: A total of 18 types of G6PD mutations are identified in the reaserch, and c.94C>G, c.1028A>G and c.1327G>C are first reported in Chinese population. The most common G6PD mutation types in Wuhan are c.1388G>A, c.1376G>T, c.95G>A.

A feasibility study of noninvasive prenatal diagnosis in facioscapulohumeral muscular dystrophy type 1 in a Chinese family.

Objective: To demonstrate the feasibility of haplotype-based noninvasive prenatal diagnosis of Facioscapulohumeral Muscular Dystrophy type 1 (FSHD1). Methods: Bionano optical mapping was used to identify the D4Z4 structural variation of the genomic DNA sample from the proband affected with FSHD1. In addition, based on the technique of next generation sequencing, the pathogenic haplotype was determined by using trio strategy through genotyping his parents, and also fetal inheritance of paternal haplotypes was then deduced using the Hidden Markov Model. Results: Bionano optical mapping analysis revealed that the proband has only three D4Z4 repeats left in the 4q35 chromosomal region and a disease-permitting 4qA haplotype. The other normal allele of the proband contains 29 D4Z4 repeats and also a 4qA haplotype. The noninvasive cell-free fetal DNA (cffDNA)-based haplotype analysis suggested that the fetus inherited the pathogenic allele from his father and thus was predicted to be affected by FSHD1. In addition, Bionano optical mapping also demonstrated the presence of the pathogenic allele in the fetus by interrogating the genomic DNA from the amniotic fluid cells. Conclusion: Our study showed the cffDNA-based haplotyping was feasible for the noninvasive prenatal diagnosis of FSHD1, which is able to provide earlier testing results with a lower risk of miscarriage and infection than invasive techniques.

20-Hydroxyeicosatetraenoic acid regulates the expression of Nedd4­2 in kidney and liver through a neddylation modification pathway.

The present study aimed to test whether 20-hydroxyeicosatetraenoic acid (20­HETE) affected neddylation modification of E3­ligase Nedd4­2 (neural precursor cell expressed, developmentally down­regulated 4­like, E3 ubiquitin protein ligase). A cytochrome P450 family 4 subfamily F member 2 (CYP4F2) transgenic mouse model that overproduces 20­HETE in the kidney and the liver was used in the present study. Transgenic mice with high salt intake exhibited increased activation of Nedd4­2­mediated ubiquitin­proteasome pathway. Nedd4­2 expression is increased in the kidney and decreased in the liver of transgenic mice compared with wild­type mice. Subsequently, co­immunoprecipitation analysis indicated that Nedd4­2 was modified by Nedd8, and the level of neddylation on Nedd4­2 was reduced in the kidney and increased in the liver of transgenic mice compared with controls. In addition, sentrin­specific protease 8 (Senp8), a deneddylation enzyme, is expressed higher in the kidney and lower in the liver of transgenic mice compared with wild­type controls. The function of 20­HETE on modulation of Nedd4­2 were also confirmed in mouse M1 kidney and mouse NCTC1469 liver cell lines, and the function was restored by neddylation inhibitor MLN4924 Data from the present study demonstrated that 20­HETE upregulated the expression of Nedd4­2 in the kidney and downregulated expression in the liver through the neddylation modification pathway, at least partly, depending on the effects on Senp8 deneddylation.

20-HETE attenuates the response of glucose-stimulated insulin secretion through the AKT/GSK-3ß/Glut2 pathway.

We previously generated cytochrome P450 4F2 (CYP4F2) transgenic mice that have high levels of 20-hydroxyeicosatetraenoic acid (20-HETE) production; these mice exhibit both hypertension and hyperglycemia without insulin resistance. Currently, it is unclear whether and how 20-HETE affects insulin secretion, thus resulting in hyperglycemia. In this study, we found that 20-HETE attenuated glucose-stimulated insulin secretion (GSIS) in CYP4F2 transgenic mice as well as in rat insulinoma INS-1E cells treated with 0.5 µM 20-HETE. HET0016, a selective inhibitor of 20-HETE synthesis, reversed the reduction in GSIS leading to a decrease in blood glucose in the transgenic mice. Furthermore, the expression of glucose transporter 2 (Glut2), Ser473 phosphorylation of protein kinase B (AKT), and Ser9 phosphorylation of glycogen synthase kinase-3ß (GSK-3ß) were decreased in CYP4F2 transgenic mice compared with wild-type mice. In vitro experiments in INS-1E cells revealed that 20-HETE activated the AKT/GSK-3ß pathway and thereby decreased Glut2 expression by inhibiting activator protein 1 (AP-1). TWS119, a GSK-3ß selective inhibitor, blocked the 20-HETE-mediated reduction in Glut2 expression. Therefore, we concluded that 20-HETE inhibition of Glut2 contributes to the reduction in GSIS, at least in part, through the AKT/GSK-3ß/AP-1/Glut2 pathway.