Single-cell characterization of self-renewing primary trophoblast organoids as modeling of EVT differentiation and interactions with decidual natural killer cells.

BACKGROUND: Extravillous trophoblast cell (EVT) differentiation and its communication with maternal decidua especially the leading immune cell type natural killer (NK) cell are critical events for placentation. However, appropriate in vitro modelling system and regulatory programs of these two events are still lacking. Recent trophoblast organoid (TO) has advanced the molecular and mechanistic research in placentation. Here, we firstly generated the self-renewing TO from human placental villous and differentiated it into EVTs (EVT-TO) for investigating the differentiation events. We then co-cultured EVT-TO with freshly isolated decidual NKs for further study of cell communication. TO modelling of EVT differentiation as well as EVT interaction with dNK might cast new aspect for placentation research. RESULTS: Single-cell RNA sequencing (scRNA-seq) was applied for comprehensive characterization and molecular exploration of TOs modelling of EVT differentiation and interaction with dNKs. Multiple distinct trophoblast states and dNK subpopulations were identified, representing CTB, STB, EVT, dNK1/2/3 and dNKp. Lineage trajectory and Seurat mapping analysis identified the close resemblance of TO and EVT-TO with the human placenta characteristic. Transcription factors regulatory network analysis revealed the cell-type specific essential TFs for controlling EVT differentiation. CellphoneDB analysis predicted the ligand-receptor complexes in dNK-EVT-TO co-cultures, which relate to cytokines, immunomodulation and angiogenesis. EVT was known to affect the immune properties of dNK. Our study found out that on the other way around, dNKs could exert effects on EVT causing expression changes which are functionally important. CONCLUSION: Our study documented a single-cell atlas for TO and its applications on EVT differentiation and communications with dNKs, and thus provide methodology and novel research cues for future study of human placentation.

New technology for preparing energetic materials by nanofiltration membrane (NF): rapid and efficient preparation of high-purity ammonium dinitramide (ADN).

The development of environment-friendly and non-toxic green energetic materials and their safe, environmentally friendly, and economical production is very important to the national economy and national security. As an innovative, efficient, and environmentally friendly energetic material, the preferred preparation method of ammonium dinitramide (ADN) is the nitro-sulfur mixed acid method, which has the advantages of high yield, simple method, and easy access to raw materials. However, the large number of inorganic salt ions introduced by this method limits the large-scale production of ADN. Nanofiltration (NF) has been widely used in various industrial processes as a separation method with high separation efficiency and simple operation. In this study, NF was used for the desalination and purification of ADN synthesized by the mixed acid method. The effects of NF types, operation process (pressure, temperature, and feed solution concentration) on desalination efficiency, and membrane flux during purification were examined. The results showed that 600D NF could achieve the efficient desalination and purification of ADN. It was verified that the highest desalination and purification efficiency was achieved at 2 MPa pressure, 25 °C, and 1 time dilution of the feed solution, and the membrane flux of the desalination and purification process was stable. Under the optimized process conditions, the removal rate of inorganic salts and other impurities reached 99% (which can be recycled), the purity of ADN reached 99.8%, and the recovery rate reached 99%. This process has the potential for the large-scale production of ADN and provides a new process for the safe, efficient, and cheap preparation of energetic materials.

Application of a JEG-3 organoid model to study HLA-G function in the trophoblast.

The human placenta is a unique temporary organ with a mysterious immune tolerance. The formation of trophoblast organoids has advanced the study of placental development. HLA-G is uniquely expressed in the extravillous trophoblast (EVT) and has been linked to placental disorders. With older experimental methodologies, the role of HLA-G in trophoblast function beyond immunomodulation is still contested, as is its role during trophoblast differentiation. Organoid models incorporating CRISPR/Cas9 technology were used to examine the role of HLA-G in trophoblast function and differentiation. JEG-3 trophoblast organoids (JEG-3-ORGs) were established that highly expressed trophoblast representative markers and had the capacity to differentiate into EVT. CRISPR/Cas9 based on HLA-G knockout (KO) significantly altered the trophoblast immunomodulatory effect on the cytotoxicity of natural killer cells, as well as the trophoblast regulatory effect on HUVEC angiogenesis, but had no effect on the proliferation and invasion of JEG-3 cells and the formation of TB-ORGs. RNA-sequencing analysis further demonstrated that JEG-3 KO cells followed similar biological pathways as their wild-type counterparts during the formation of TB-ORGs. In addition, neither HLA-G KO nor the exogenous addition of HLA-G protein during EVT differentiation from JEG-3-ORGs altered the temporal expression of the known EVT marker genes. Based on the JEG-3 KO (disruption of exons 2 and 3) cell line and the TB-ORGs model, it was determined that HLA-G has a negligible effect on trophoblast invasion and differentiation. Despite this, JEG-3-ORG remains a valuable model for studying trophoblast differentiation.

Preparation and evaluation of attractive microspheres for control of Agrilus planipennis fairmaire.

Agrilus planipennis Fairmaire is an important wood boring pest of Fraxinus species in the family Oleaceae. Oxacyclotridecan-2-one is an attractant of A. planipennis. Traps with attractive lures can be used in mass trapping of insect pests, but the traps are a bit expensive and they must be set up and dismantled in the field. To develop an attract and kill method for A. planipennis, we enveloped oxacyclotridecan-2-one into sustained-released microspheres. The attractant microspheres were prepared using the solvent evaporation method. An orthogonal test L16(45) was used to optimize the five preparation factors: the quantities of polylactic acid (PLA), gelatin, Polyvinyl alcohol (PVA), attractant, and the rotational speed. The results showed that optimal conditions for preparation of microspheres were 2.5 g PLA, 0.5 g gelatin, 1.25 g PVA, 2 mL attractant and 600 r min-1 rotational speed. The encapsulation efficiency of the prepared microspheres was 95.22%, and the attractant loading rate was 15.61%. The release rate of attractant from prepared microspheres was about 26.74% on the first day, and then gradually entered a sustained-release stage for about 10 days that lasted for 17 days. Preliminary field control experiments showed that the prepared microspheres could attract and kill A. planipennis adults when sprayed together with insecticide.

Corrigendum: Identification of key endometrial microRNAs and their target genes associated with pathogenesis of recurrent implantation failure by integrated bioinformatics analysis.

[This corrects the article DOI: 10.3389/fgene.2022.919301.].

Identification of Key Endometrial MicroRNAs and Their Target Genes Associated With Pathogenesis of Recurrent Implantation Failure by Integrated Bioinformatics Analysis.

Purpose: Recurrent implantation failure (RIF) is an enormous challenge for in vitro fertilization (IVF) clinicians. An understanding of the molecular mechanisms of RIF helps to predict prognosis and develop new therapeutic strategies. The study is designed to identify diagnostic biomarkers for RIF as well as the potential mechanisms underlying RIF by utilizing public databases together with experimental validation. Methods: Two microarray datasets of RIF patients and the healthy control endometrium were downloaded from the Gene Expression Omnibus (GEO) database. First, differentially expressed microRNAs (miRNAs) (DEMs) were identified and their target genes were predicted. Then, we identified differentially expressed genes (DEGs) and selected hub genes through protein-protein interaction (PPI) analyses. Functional enrichment analyses of DEGs and DEMs were conducted. Furthermore, the key DEMs which targeted these hub genes were selected to obtain the key miRNA-target gene network. The key genes in the miRNA-target gene network were validated by a single-cell RNA-sequencing dataset of endometrium from GEO. Finally, we selected two miRNA-target gene pairs for further experimental validation using dual-luciferase assay and quantitative polymerase chain reaction (qPCR). Results: We identified 49 DEMs between RIF patients and the fertile group and found 136,678 target genes. Then, 325 DEGs were totally used to construct the PPI network, and 33 hub genes were selected. Also, 25 DEMs targeted 16 key DEGs were obtained to establish a key miRNA-target gene network, and 16 key DEGs were validated by a single-cell RNA-sequencing dataset. Finally, the target relationship of hsa-miR-199a-5p-PDPN and hsa-miR-4306-PAX2 was verified by dual-luciferase assay, and there were significant differences in the expression of those genes between the RIF and fertile group by PCR (p < 0.05). Conclusion: We constructed miRNA-target gene regulatory networks associated with RIF which provide new insights regarding the underlying pathogenesis of RIF; hsa-miR-199a-5p-PDPN and hsa-miR-4306-PAX2 could be further explored as potential biomarkers for RIF, and their detection in the endometrium could be applied in clinics to estimate the probability of successful embryo transfer.

Effect of precise nursing service mode on postoperative urinary incontinence prevention in patients with prostate disease.

BACKGROUND: Patients with benign prostatic disease often experience detrusor morphological changes and dysfunction. In severe cases, it leads to bladder detrusor dysfunction, resulting in dysuria, frequent urination, urgent urination, incomplete urination, and other symptoms including renal function injury. An operation to restore normal urination function and to control postoperative complications, as far as possible, is the most common method for benign prostatic disease. AIM: To observe the effect of precise nursing service mode on postoperative urinary incontinence prevention in patients with prostate disease. METHODS: In total, 130 patients diagnosed with benign prostatic disease, from January 2018 to June 2021, in our hospital, were selected and divided into observation and control groups according to their treatment options. Sixty-five cases in the control group were given routine nursing mode intervention and 65 cases in the observation group received precise nursing service mode intervention. The intervention with the observation group included psychological counseling about negative emotions, pelvic floor exercises, and post-hospital discharge care. The complications of the two groups were counted, and the general postoperative conditions of the two groups were recorded. The urinary flow dynamics indexes of the two groups were detected, and differences in clinical international prostate system score (IPSS) and urinary incontinence quality of life questionnaire (I-QOL) scores were evaluated. RESULTS: Postoperative exhaust time (18.65 ± 3.23 h and 24.63 ± 4.51 h), the time of indwelling catheter (4.85 ± 1.08 d and 5.63 ± 1.24 d), and hospitalization time (8.78 ± 2.03 d and 10.23 ± 2.28 d) in the observation group were lower than in the control group. The difference was statistically significant (P < 0.05). After the operation, the maximum urinary flow rate (Qmax) increased (P < 0.05), the residual urine volume (RUV) decreased (P < 0.05), and the maximum closed urethral pressure (MUCP) was not statistically significant (P > 0.05) compared to pre-operation. The Qmax of the observation group was higher than that of the control group, while the RUV was lower than that of the control group. There was no significant difference in MUCP between the observation and control groups (P > 0.05). The I-QOL score of the two groups improved (P < 0.05), and the IPSS decreased (P < 0.05). After the operation, the I-QOL score of the observation group was higher than that of the control group, and the IPSS was lower than that of the control group (P < 0.05). There were no significant differences in the incidence of urethral injury (1.54% and 3.08%), bladder spasm (0.00% and 1.54%), and secondary bleeding (1.54% and 4.62) between the observation and control groups (P > 0.05). CONCLUSION: The precise nursing service mode can reduce the incidence of postoperative urinary incontinence in patients with prostate disease, thus improving postoperative urodynamics and rehabilitation, and quality of life.

The Inhibitory Effect of Cyclodextrin on Oxygen Bioavailability Is a Key Factor for the Metabolic Flux Redistribution Toward Steroid Alcohols in Phytosterol Resting Cells Bioconversion.

In the present work, we tried to identify the mechanism why by which the steroid alcohols accumulated when hydroxypropyl-ß-cyclodextrin (HP-ß-CD) was present to enhance the sterol conversion rate. Compared with the bioconversion system without HP-ß-CD, the reaction rate was greatly improved in presence of HP-ß-CD, but the steroid alcohols largely accumulated concurrently. In a reaction system with an enhanced reaction rate, the higher intracellular NADH/NAD+ level was detected, and the production of steroid alcohols increased also. Mycobacterium neoaurum mutants with higher KshA activity (3-ketosteroid 9α-hydrolase, a monooxygenase hydroxylating the nucleus at C-9 at the expense of NAD(P)H consumption) reduced the steroid alcohol production, and in the meantime, the NADH/NAD+ level was decreased consequently. Further research found that oxygen availability was seriously inhibited by the cyclodextrin in a reaction system. These results indicated that NADH formed in the bioconversion was not properly regenerated via the respiratory chain because of the poor oxygen bioavailability. The inhibitory effect of cyclodextrin on oxygen bioavailability is a key factor for the metabolic flux redistribution toward steroid alcohols in phytosterol resting cells bioconversion.

Identification and in situ removal of an inhibitory intermediate to develop an efficient phytosterol bioconversion process using a cyclodextrin-resting cell system.

A classically versatile steroid intermediate, 9α-hydroxyandrost-4-ene-3,17-dione (9α-OH-AD), can be obtained by phytosterol (PS) bioconversion using Mycobacterium. In this study, a cyclodextrin-resting cell reaction system with a high concentration of PS (50 g L-1) was used to produce 9α-OH-AD. However, the inhibitory effect of metabolic intermediates is a key factor limiting production efficiency. After the separation and identification of a series of metabolic intermediates, it was found that 4-ene-3-keto steroids, which are the first metabolites of sterol side-chain degradation, accumulated at the beginning of the bioprocess and had a remarkable inhibitory effect on bioconversion. The bioconversion rate was greatly improved when 5 g L-1 of macroporous adsorbent resin D101 was added to the reaction system in the initial phase. A certain amount of resin acted as a reservoir to remove the inhibitory intermediate in situ and facilitated the bioconversion process, and the 9α-OH-AD space-time yield increased to 8.51 g L-1 d-1, which was 23.15% higher than that without resin addition (6.91 g L-1 d-1) after 72 h bioconversion. In summary, we identified an inhibitory intermediate that limits the bioconversion rate and provided a solution based on resin adsorption for improving 9α-OH-AD production efficiency in a commercial-scale process.

MicroRNAs: Key Regulators in the Central Nervous System and Their Implication in Neurological Diseases.

MicroRNAs (miRNAs) are a class of small, well-conserved noncoding RNAs that regulate gene expression post-transcriptionally. They have been demonstrated to regulate a lot of biological pathways and cellular functions. Many miRNAs are dynamically regulated during central nervous system (CNS) development and are spatially expressed in adult brain indicating their essential roles in neural development and function. In addition, accumulating evidence strongly suggests that dysfunction of miRNAs contributes to neurological diseases. These observations, together with their gene regulation property, implicated miRNAs to be the key regulators in the complex genetic network of the CNS. In this review, we first focus on the ways through which miRNAs exert the regulatory function and how miRNAs are regulated in the CNS. We then summarize recent findings that highlight the versatile roles of miRNAs in normal CNS physiology and their association with several types of neurological diseases. Subsequently we discuss the limitations of miRNAs research based on current studies as well as the potential therapeutic applications and challenges of miRNAs in neurological disorders. We endeavor to provide an updated description of the regulatory roles of miRNAs in normal CNS functions and pathogenesis of neurological diseases.

In situ fabrication of nanostructured titania coating on the surface of titanium wire: A new approach for preparation of solid-phase microextraction fiber.

Nanostructured titania-based solid-phase microextraction (SPME) fibers were fabricated through the in situ oxidation of titanium wires with H(2)O(2) (30%, w/w) at 80 degrees C for 24h. The obtained SPME fibers possess a approximately 1.2microm thick nanostructured coating consisting of approximately 100nm titania walls and 100-200nm pores. The use of these fibers for headspace SPME coupled with gas chromatography with electron capture detection (GC-ECD) resulted in improved analysis of dichlorodiphenyltrichloroethane (DDT) and its degradation products. The presented method to detect DDT and its degradation products has high sensitivity (0.20-0.98ngL(-1)), high precision (relative standard deviation R.S.D.=9.4-16%, n=5), a wide linear range (5-5000ngL(-1)), and good linearity (coefficient of estimation R(2)=0.991-0.998). As the nanostructured titania was in situ formed on the surface of a titanium wire, the coating was uniformly and strongly adhered on the titanium wire. Because of the inherent chemical stability of the titania coating and the mechanical durability of the titanium wire substrate, this new SPME fiber exhibited long life span (over 150 times).

Poly[penta-µ-aqua-µ(6)-methyl-ene-disulfonato-µ(5)-methyl-enedisulfonato-tetra-sodium(I)].

The title compound, [Na(4)(CH(2)O(6)S(2))(2)(H(2)O)(5)](n), was crystallized from an aqueous solution. The sodium ions are surrounded and bridged by O atoms from coordinated water mol-ecules and sulfonate ions in a three-dimensional neutral network. The crystal structure is also stabilized by an intricate system of hydrogen bonds.