Unraveling the impact of hyperleptinemia on female reproduction: insights from transgenic pig model.

BACKGROUND: Infertility is a growing global health concern affecting millions of couples worldwide. Among several factors, an extreme body weight adversely affects reproductive functions. Leptin is a well-known adipokine that serves as an endocrine signal between adiposity and fertility. However, the exact mechanisms underlying the effects of high leptin level on female reproduction remain unclear. METHODS: Transgenic pigs overexpressing leptin (♀) were produced by backcrossing and screened for leptin overexpression. The growth curve, fat deposition, reproductive performance, apoptosis, serum hormones and cholesterol production, RNA sequencing, and single-nucleus RNA sequencing (snRNA-seq) of the leptin-overexpressing pigs and wild-type group were evaluated. RESULTS: Transgenic pigs overexpressing leptin (♀) were obtained, which exhibited significantly reduced body weight, body size, and back fat thickness. These pigs manifested a late onset of puberty (330 ± 54.3 vs. 155 ± 14.7 days), irregular estrous behavior characterized by increased inter-estrous interval (29.2 ± 0 vs. 21.3 ± 0.7 days), and more number of matings until pregnancy (at least 3 times). This reproductive impairment in leptin pigs was related to hormonal imbalances characterized by increased levels of FSH, LH, prolactin, E2, P4, and TSH, altered steroidogenesis such as increased levels of serum cholesterol esters along with steroidogenic markers (StAR, CYP19A), and ovarian dysfunctions manifested by neutrophilic infiltration and low expression of caspase-3 positive cells in the ovaries. Moreover, bulk RNA sequencing of the ovaries also revealed neutrophilic infiltration followed by upregulation of inflammation-related genes. Furthermore, snRNA-seq reflected that leptin overexpression triggered immune response, suppressed follicle development and luteinization, resulting in metabolic dysfunction and hormone imbalance in the ovary. CONCLUSIONS: Low body weight in leptin overexpressing pigs adversely affects the reproductive performance, causing delayed puberty, irregular estrous cycles, and reduced breeding efficiency. This is linked to metabolic imbalances, an increased immune response, and altered ovarian functions. This study provides a theoretical basis for the complex mechanisms underlying leptin, and infertility by employing leptin-overexpressing female pigs.

Integrating mTOR Inhibition and Photodynamic Therapy Based on Carrier-Free Nanodrugs for Breast Cancer Immunotherapy.

Conventional photodynamic therapy (PDT) in cancer treatment needs to utilize oxygen to produce reactive oxygen species to eliminate malignant tissues. However, oxygen consumption in tumor microenvironment exacerbates cancer cell hypoxia and may promote vasculature angiogenesis. Since the mammalian target of rapamycin (mTOR) signaling pathway plays a vital role in endothelial cell proliferation and fibrosis, mTOR inhibitor drugs hold the potential to reverse hypoxia-evoked angiogenesis for improved PDT effect. In this study, a carrier-free nanodrug formulation composed of Torin 1 as mTORC1/C2 dual inhibitor and Verteporfin as a photosensitizer and Yes-associated protein inhibitor is developed. These two drug molecules can self-assemble into stable nanoparticles through π-π stacking and hydrophobic interactions with good long-term stability. The nanodrugs can prompt synergistic apoptosis, combinational anti-angiogenesis, and strong immunogenic cell death effects upon near-infrared light irradiation in vitro. Furthermore, the nanosystem also exhibits improved antitumor effect, anti-cancer immune response, and distant tumor inhibition through tumor microenvironment remodeling in vivo. In this way, the nanodrugs can reverse PDT-elicited angiogenesis and promote cancer immunotherapy to eliminate tumor tissues and prevent metastasis. This nanosystem provides insights into integrating mTOR inhibitors and photosensitizers for safe and effective breast cancer treatment in clinical settings.

Progress on SOX9 and its enhancers in mammalian sex determination.

The sex determination in mammals refers to the development of an initial bipotential organ, termed the bipotential gonad/genital ridge, into either a testis or an ovary at the early stages of embryonic development, under the precise regulation of transcription factors. SOX9 (SRY-box transcription factor 9) is a multifunctional transcription factor in mammalian development and plays a critical role in sex determination and subsequent male reproductive organs development. Recent studies have shown that several enhancers upstream of SOX9 also play an important role in the process of sex determination. In this review, we summarize the progress on the role of SOX9 and its gonadal enhancers in sex determination. This review will facilitate to understand the regulatory mechanism of sex determination of SOX9 and provides a theoretical basis for the further development of animal sex manipulation technologies.

Generation of RAG2 Knockout Immune-Deficient Miniature Pigs.

Recombination-activating genes (RAGs) play a crucial role in the V(D)J recombination process and the development of immune cells. The development of the immune system and its mechanisms in pigs exhibit greater similarity to those of humans compared to other animals, thus rendering pigs a valuable tool for biomedical research. In this study, we utilized CRISPR/Cas9 gene editing and somatic cell nuclear transfer technology to generate RAG2 knockout (KO) pigs. Furthermore, we evaluated the impact of RAG2 KO on the immune organs and immune cell development through morphological observations, blood analysis and flow cytometry technology. RAG2 KO cell lines were used as donors for cloning. The reconstructed embryos were transplanted into 4 surrogate sows, and after 116 days of gestation, 2 sows gave birth to 12 live piglets, all of which were confirmed to be RAG2 KO. The thymus and spleen sizes of RAG2 KO pigs were significantly smaller than those of wild-type (WT) pigs. Hematoxylin-eosin staining results revealed that the thymus and spleen tissue structures of RAG2 KO pigs were disorganized and lacked the characteristic structures, indicating that RAG2 KO leads to dysplasia of the thymus and spleen. Hematological analysis demonstrated that the total number of white blood cells and lymphocytes in the circulation of RAG2 KO pigs was significantly lower, while the number of eosinophils was higher. Flow cytometry results indicated that the proportions of mature T and B lymphocytes were significantly reduced compared to WT pigs. These findings successfully verified the immunodeficiency phenotype of RAG2 KO pigs. This study may provide experimental animals for the development of tumor models and humanized animals.

Effects of cross-sectional shapes and arrangement patterns on windbreak effects of shelterbelts by large eddy simulation.

Investigating the wind speed flow field and aerodynamic characteristics of shelterbelts with different structural features is of significant importance for the rational arrangement of shelterbelts and the mitigation of wind-blown sand disasters. Considering five cross-sectional shapes of shelterbelts (rectangle, windward right-angle triangle, leeward right-angle triangle, isosceles triangle, and parabolic) and four layout forms (single shelterbelt, L-shaped network, U-shaped network, and rectangular network), we conducted computational fluid dynamics (CFD) simulations using the large eddy simulation (LES) turbulence model to understand mean wind speed flow field and turbulence structure of shelterbelts with different structural features, and investigated the effects of shelterbelt cross-sectional shapes and layout forms on windbreak indicators, such as protection distance and area. We considered tree canopies as porous media and conducted simulation with the 'Tsujimatsu' shelterbelt in Japan with a total height (H) of 7 m, canopy height of 5.8 m, and a canopy base width of 2 m. The results showed that the average relative errors of mean wind speed and turbulent kinetic energy at different heights obtained by numerical simulations and field measurement were small, being 5.5% and 12%, respectively, indicating that the porous medium canopy model successfully reproduced the mean wind speed and turbulent kinetic energy in the leeward area of the shelterbelt. The rectangular cross-section shelterbelt, with the largest canopy volume, significantly obstructed airflow. The mean wind speed and turbulent kinetic energy showed a notable reduction in the leeward area near the shelterbelt, especially in the upper region (z≥0.5H, where z denoted the height), showing the largest protection range. The parabolic cross-section shelterbelt ranked second in terms of protection range, followed by shelterbelts with windward right-angle, leeward right-angle, and isosceles triangular cross-sections. In the downstream area where horizontal distance x≥10H, the mean wind speed and turbulent kinetic energy of shelterbelts with different cross-sectional shapes tended to be the same. Comparing the flow field structures of single shelterbelts and L-shaped, U-shaped, and rectangular networks, it revealed that the more shelterbelts oriented perpendicular to the incoming wind speed, the more pronounced the wind speed attenuation behind the canopy, a longer distance would be required for airflow to recover to the incoming wind speed. In contrast, the wind protection effect of shelterbelts paralleled to the wind direction was extremely limited, making the U-shaped and rectangular networks more effective in wind protection than single shelterbelts and L-shaped networks. The findings would provide references for the structural configuration and optimal layout of shelterbelt systems.

New CRISPR/Cas9-based Fgfr2C361Y/+ mouse model of Crouzon syndrome exhibits skull and behavioral abnormalities.

Crouzon syndrome (CS), a syndromic craniosynostosis, is a craniofacial developmental deformity caused by mutations in fibroblast growth factor receptor 2 (FGFR2). Previous CS mouse models constructed using traditional gene editing techniques faced issues such as low targeting efficiency, extended lineage cycles, and inconsistent and unstable phenotypes. In this study, a CRISPR/Cas9-mediated strategy was employed to induce a functional augmentation of the Fgfr2 point mutation in mice. Various techniques, including bone staining, micro-CT, histological methods, and behavioral experiments, were employed to systematically examine and corroborate phenotypic disparities between mutant mice (Fgfr2C361Y/+) and their wild-type littermates. Confirmed via PCR-Sanger sequencing, we successfully induced the p.Cys361Tyr missense mutation in the Fgfr2 IIIc isoform of the extracellular domain (corresponding to the p.Cys342Tyr mutation in humans) based on Fgfr2-215 transcript (ENSMUST00000122054.8). Fgfr2C361Y/+ mice exhibited characteristics consistent with the phenotypic features associated with CS, including skull-vault craniosynostosis, skull deformity, shallow orbits accompanied by exophthalmos, midface hypoplasia with malocclusion, and shortened skull base, notably without any apparent limb defects. Furthermore, mutant mice displayed behavioral abnormalities encompassing deficits in learning and memory, social interaction, and motor dysfunction, without anxiety-related disorders. Histopathological examination of the hippocampal region revealed structural abnormalities, suggesting possible brain development impairment secondary to craniosynostosis. In conclusion, we constructed a novel gene-edited Fgfr2C361Y/+ mice strain based on CRISPR/Cas9, which displayed skull and behavioral abnormalities, serving as a new model for studying genetic molecular mechanisms and exploring treatments for CS. KEY MESSAGES: CRISPR/Cas9 crafted a Crouzon model by enhancing Fgfr2-C361Y in mice. Fgfr2C361Y/+ mice replicate CS phenotypes-craniosynostosis and midface anomalies. Mutant mice show diverse behavioral abnormalities, impacting learning and memory. Fgfr2C361Y/+ mice offer a novel model for cranial suture studies and therapeutic exploration.

Surgical outcomes and intraoperative parameters of 3D visualization system versus conventional microscopes for macular surgery in highly myopic eyes, a prospective randomized clinical trial.

PURPOSE: To investigate the surgical outcomes and intraoperative parameters of 3D visualization system for macular diseases in highly myopic eyes. METHODS: In this single-center, prospective, randomized, comparative interventional study, 40 highly myopic eyes (axial length > 26mm) were randomly assigned to either a 3D visualization system or a conventional microscope (CM) group. Surgical outcomes and intraoperative parameters, including the number of indocyanine green (ICG) injections, surgical time, and epiretinal membrane (ERM)/ internal limiting membrane (ILM) peeling time, were compared. RESULTS: The 3D group required significantly fewer ICG injections (1.3 ± 0.5 vs. 2.3 ± 0.7, p < 0.001), had shorter ERM/ILM peeling times (522.8 ± 258.0 vs. 751.8 ± 320.2 sec, p < 0.05), and experienced fewer intraoperative retinal hemorrhages (0 vs. 7 cases, p < 0.05) compared to the CM group. Anatomical and functional outcomes were comparable between the two groups. CONCLUSION: The 3D system exhibited a lower number of ICG injections, shorter ERM/ILM peeling times and a reduced incidence of intraoperative retinal hemorrhages, suggesting the 3D visualization system may offer advantages for macular surgery in highly myopic eyes.

Production of Four-Gene (GTKO/hCD55/hTBM/hCD39)-Edited Donor Pigs and Kidney Xenotransplantation.

BACKGROUND: The number of multigene-modified donor pigs for xenotransplantation is increasing with the advent of gene-editing technologies. However, it remains unclear which gene combination is suitable for specific organ transplantation. METHODS: In this study, we utilized CRISPR/Cas9 gene editing technology, piggyBac transposon system, and somatic cell cloning to construct GTKO/hCD55/hTBM/hCD39 four-gene-edited cloned (GEC) pigs and performed kidney transplantation from pig to rhesus monkey to evaluate the effectiveness of these GEC pigs. RESULTS: First, 107 cell colonies were obtained through drug selection, of which seven were 4-GE colonies. Two colonies were selected for somatic cell nuclear transfer (SCNT), resulting in seven fetuses, of which four were GGTA1 biallelic knockout. Out of these four, two fetuses had higher expression of hCD55, hTBM, and hCD39. Therefore, these two fetuses were selected for two consecutive rounds of cloning, resulting in 97 live piglets. After phenotype identification, the GGTA1 gene of these pigs was inactivated, and hCD55, hTBM, and hCD39 were expressed in cells and multiple tissues. Furthermore, the numbers of monkey IgM and IgG binding to the peripheral blood mononuclear cells (PBMCs) of the 4-GEC pigs were markedly reduced. Moreover, 4-GEC porcine PBMCs had greater survival rates than those from wild-type pigs through complement-mediated cytolysis assays. In pig-to-monkey kidney xenotransplantation, the kidney xenograft successfully survived for 11 days. All physiological and biochemical indicators were normal, and no hyperacute rejection or coagulation abnormalities were found after transplantation. CONCLUSION: These results indicate that the GTKO/hCD55/hTBM/hCD39 four-gene modification effectively alleviates immune rejection, and the pig kidney can functionally support the recipient monkey's life.

Relationship between genotype, phenotype, and refractive status in patients of inherited retinal degeneration.

BACKGROUND: To elucidate the relationship between inherited retinal disease, visual acuity and refractive error development in Asian patients. SUBJECTS: Five hundred phakic eyes with refractive data were analysed in this retrospective cohort. Diseases were categorized by clinical phenotypes, and the prevalent genotypes identified in the Taiwan Inherited Retinal Degeneration Project were analysed. Consecutive surveys in Taiwan have provided the rates of myopia in the general population. RESULTS: No differences were observed among the disease phenotypes with respect to myopia (P = 0.098) and high myopia rates (P = 0.037). The comparison of refractive error between retinitis pigmentosa and diseases mainly affecting the central retina showed no difference, and the refraction analyses in diseases of different onset ages yielded no significance. Moreover, there was no difference in the myopia rate between the diseases and general population. Among the genotypes, a higher spherical equivalent was seen in RPGR and PROM1-related patients and emmetropic trends were observed in patients with CRB1 and PRPF31 mutations. Furthermore, significantly poorer visual acuity was found in ABCA4, CRB1 and PROM1-related patients, and more preserved visual acuity was seen in patients with EYS, USH2A, and RDH12 mutations. CONCLUSIONS: No significant differences were observed in visual acuity, refractive state and myopia rate between patients with inherited retinal disease and the general population, and different subtypes of inherited retinal disease shared similar refractive state, except for higher cylindrical dioptres found in patients with Leber's congenital amaurosis. The heterogeneity of disease-causing genes in Asian patients may lead to variable refractive state.

Autocrine IL-8 Contributes to Propionibacterium Acnes-induced Proliferation and Differentiation of HaCaT Cells via AKT/FOXO1/ Autophagy.

OBJECTIVE: Proprionibacterium acnes (P. acnes)-induced inflammatory responses, proliferation and differentiation of keratinocytes contribute to the progression of acne vulgaris (AV). P. acnes was found to enhance the production of interleukin-8 (IL-8) by keratinocytes. This study aimed to investigate the role of IL-8 in P. acnes-induced proliferation and differentiation of keratinocytes and the underlying mechanism. METHODS: The P. acnes-stimulated HaCaT cell (a human keratinocyte cell line) model was established. Western blotting and immunofluorescence were performed to detect the expression of the IL-8 receptors C-X-C motif chemokine receptor 1 (CXCR1) and C-X-C motif chemokine receptor 2 (CXCR2) on HaCaT cells. Cell counting kit-8 (CCK-8) assay, 5-ethynyl-20-deoxyuridine (EdU) assay and Western blotting were performed to examine the effects of IL-8/CXCR2 axis on the proliferation and differentiation of HaCaT cells treated with P. acnes, the IL-8 neutralizing antibody, the CXCR2 antagonist (SB225002), or the CXCR1/CXCR2 antagonist (G31P). Western blotting, nuclear and cytoplasmic separation, CCK-8 assay, and EdU assay were employed to determine the downstream pathway of CXCR2 after P. acnes-stimulated HaCaT cells were treated with the CXCR2 antagonist, the protein kinase B (AKT) antagonist (AZD5363), or the constitutively active forkhead box O1 (FOXO1) mutant. Finally, autophagy markers were measured in HaCaT cells following the transfection of the FOXO1 mutant or treatment with the autophagy inhibitor 3-methyladenine (3-MA). RESULTS: The expression levels of CXCR1 and CXCR2 were significantly increased on the membrane of HaCaT cells following P. acnes stimulation. The IL-8/CXCR2 axis predominantly promoted the proliferation and differentiation of P. acnes-induced HaCaT cells by activating AKT/FOXO1/autophagy signaling. In brief, IL-8 bound to its receptor CXCR2 on the membrane of keratinocytes to activate the AKT/FOXO1 axis. Subsequently, phosphorylated FOXO1 facilitated autophagy to promote the proliferation and differentiation of P. acnes-induced keratinocytes. CONCLUSION: This study demonstrated the novel autocrine effect of IL-8 on the proliferation and differentiation of P. acnes-induced keratinocytes, suggesting a potential therapeutic target for AV.

Identification of organic constituents on atmospheric particulate matter in the East Asian background air of free troposphere by GC×GC-TOFMS.

The presence of organic compounds on the particulate matter (PM) or aerosols can arise from the condensation of gaseous organic compounds on the existing aerosols, or from organic precursors to form secondary organic aerosols (SOA) through photochemistry. The objective of this study is to characterize organic constituents on aerosols relevant to their emission sources and the key compounds revealing the evolution of aerosols with the use of a novel analytical technique. A time-of-flight mass spectrometry (TOFMS) coupled with comprehensive two-dimensional gas chromatography (GC×GC) was developed using a flow type of modulator instead of a thermal type as a prelude to field applications without the need for cryogen. The methodology of GC×GC-TOFMS is discussed in this study in detail. Since the coarse PM (PM10-2.5) may exhibit with a relatively high OC content compared to PM2.5, the GC×GC results have been obtained by analyzing PM10 samples collected in parallel with OC/EC analysis of PM2.5 samples at the Lulin Atmospheric Background Station (LABS, 23.47°N, 120.87°E, 2862 m ASL) as the high-mountain background site in East Asia. We found that the organic analytes were in a majority in the range of 12-30 carbon numbers falling in the category of semi-volatile organic compounds (SVOCs) with 43 compounds of alcohol, aldehyde, ketone, and ester varieties if excluding alkanes. Intriguingly, trace amounts of plasticizers and phosphorus flame retardants such as phthalates (PAEs) and triphenyl phosphate (TPP) were also found, likely originating from regions involved in open burning of household solid waste in Southeast Asia or e-waste recycling in southern China and along the long-range transport route. Compounds such as these are unique to the specific sources, demonstrating the wide spread of these hazardous compounds in the environment.

Synthesis of Etrasimod (APD334): Al2O3-Promoted Decarboxylative Rearrangements of Cyclopentenones with Stereochemical Inversion.

This study presents an efficient synthesis pathway for etrasimod, starting from (+)-cis-4-acetoxy-2-cyclopenten-1-ol, yielding 5.6% overall with 98% enantiomeric excess. The crucial intermediate, (4R)-anilinocyclopent-2-enone, was derived from the (S)-alcohol/isocyanate adduct through a concerted, Al2O3-promoted decarboxylative rearrangement, which inverted the configuration. A tetracyclic fused lactam was formed via a one-pot acylation-Michael addition, followed by keto α-arylation. Subsequent removal of the oxo group facilitated the synthesis of cyclopenta[b]indol-3-ylacetic acid through a series of reactions, including methanolysis, indoline oxidation, and hydrolysis.

Constructing an interaction in plant polyphenol-modified carbon fiber with amylopectin-based waterborne polyurethane sizing agent via hydrogen bonding to improve the interfacial performance of carbon fiber/nylon 6 composites.

The adhesive strength between the sizing agent and carbon fiber (CF) plays a crucial role in improving the interfacial properties of composites, while such a vital aspect has been consistently disregarded. In this study, a hyperbranched waterborne polyurethane (HWPU) sizing agent was synthesized from biogenetically raw materials including gallic acid, l-Lysine diisocyanate and amylopectin. Concurrently, hydrogen-bonded cross-linked network structures were established utilizing a botanical polyphenol tannin as coupling agent to effectively connect CF with HWPU. This meticulous process yielded CF/nylon 6 composites with improved properties and their mechanical characteristics were systematically investigated. The findings showcased a noteworthy boost in flexural strength and interlaminar shear strength (ILSS), showing enhancements of 54.6 % and 61.4 %, respectively, surpassing those of untreated CF. Furthermore, the interfacial shear strength (IFSS) test indicated a remarkable 70.3 % improvement. This approach presents a highly promising concept aimed at developing sustainable green waterborne polyurethane sizing agent and improving the interfacial performance of CF composite materials.

Shikonin ameliorates depressive- and anxiogenic-like behaviors in rats via the suppression of inflammation in the hippocampus.

Shikonin is an active naphthoquinone with antioxidative, anti-inflammatory, and anticancer properties. In this study, we investigated the effects of shikonin on depressive- and anxiety-like behaviors in lipopolysaccharide- (LPS-) induced depression and chronic unpredictable mild stress (CUMS) rat models and explored the potential mechanism. First, a 14-day intraperitoneal administration of shikonin (10 mg/kg) significantly decreased immobility time in forced swimming test (FST) and increased open arm entries in elevated plus maze (EPM) test, without affecting line crossings in open field test (OFT), indicating that shikonin has anti-depressant- and anxiolytic-like effects. Second, chronic shikonin administration (10 mg/kg) reversed depressive- and anxiety-like behaviors in LPS-induced and CUMS depression models, as shown in the sucrose preference test (SPT), FST, EPM, and novel object recognition test (NORT). Finally, shikonin significantly reduced the levels of interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor-α (TNF-α) in hippocampus, indicating that the anti-depressant- and anxiolytic-like effects of shikonin are related to the reduction of neuroinflammation in hippocampus. These findings suggest that shikonin exerts anti-depressant- and anxiolytic-like effects via an anti-inflammatory mechanism of shikonin in the hippocampus.

Physical activity promotes brain development through serotonin during early childhood.

Early childhood serves as a critical period for neural development and skill acquisition when children are extremely susceptible to the external environment and experience. As a crucial experiential stimulus, physical activity is believed to produce a series of positive effects on brain development, such as cognitive function, social-emotional abilities, and psychological well-being. The World Health Organization recommends that children engage in sufficient daily physical activity, which has already been strongly advocated in the practice of preschool education. However, the mechanisms by which physical activity promotes brain development are still unclear. The role of neurotransmitters, especially serotonin, in promoting brain development through physical activity has received increasing attention. Physical activity has been shown to stimulate the secretion of serotonin by increasing the bioavailability of free tryptophan and enriching the diversity of gut microbiota. Due to its important role in modulating neuronal proliferation, differentiation, synaptic morphogenesis, and synaptic transmission, serotonin can regulate children's explicit cognitive and social interaction behavior in the early stages of life. Therefore, we hypothesized that serotonin emerges as a pivotal transmitter that mediates the relationship between physical activity and brain development during early childhood. Further systematic reviews and meta-analyses are needed to specifically explore whether the type, intensity, dosage, duration, and degree of voluntariness of PA may affect the role of serotonin in the relationship between physical activity and brain function. This review not only helps us understand the impact of exercise on development but also provides a solid theoretical basis for increasing physical activity during early childhood.

Impact of COVID-19 on emotional and behavioral problems among preschool children: a meta-analysis.

The impact of the Coronavirus Disease 2019 (COVID-19) on society is continuous, resulting in negative psychological consequences. Given the vulnerability and sensitivity to the environment among preschool children, their emotional and behavioral problems deserve more attention. The current study aimed to explore the impact of the epidemic on preschool children's mental health by determining the pooled prevalence of emotional and behavioral problems amidst the Coronavirus Disease 2019 pandemic and to reveal potential reasons for variations between studies. Published studies were searched in Embase, PubMed, ProQuest, PsycINFO, Web of Science, CNKI, and Wanfang. Based on the inclusion criteria outlined in this study, a total of 10 studies encompassing 38,059 participants were incorporated. Employing a random-effect model for estimating the prevalence of emotional and behavioral problems, the results revealed a pooled prevalence rate of 24.3% (95% CI, 0.15-0.38; I²=99.9%) among preschool children. This rate surpasses the pre-outbreak prevalence observed in different countries, signifying a detrimental influence of the epidemic on the mental well-being of preschoolers. Therefore, mental health care and recovery are essential for the vulnerable group during and after the public health crisis. Specific emotional and behavioral problems among preschool children are expected to be researched in the future to provide more targeted guidance for intervention.

Cross-kingdom regulation of ginseng miRNA156 on immunity and metabolism.

AIM OF THE STUDY: To study the cross-border regulation of immunity and energy metabolism by ginseng miRNA156, and to provide a new perspective for further exploring the possibility of ginseng miRNA156 as a pharmacodynamic substance. MATERIALS AND METHODS: Combined with the previous research results of our research group, miRNA156 with high expression in blood sequencing of intragastrically administered with ginseng decoction was selected. Bioinformatics analysis was performed on the selected differential miRNA156. The target genes of differential miRNA156 were mainly enriched in metabolic, immune and other signaling pathways. According to the analysis results, the experimental part will use qi deficiency fatigue model and RAW264.7 cells. The contents of lactic acid (LA), creatine kinase (CK), blood urea nitrogen (BUN), lactate dehydrogenase (LD), liver glycogen (LG), muscle glycogen (MG), interleukin 4 (IL-4), matrix metallo-proteinase 9 (MMP-9), superoxide dismutase (SOD), malondialdehyde, phosphor-enolpyruvate carboxykinase (PEPCK), glucose-6-phosphatase (G6pase), nitric oxide (NO) and tumor necrosis factor-α (TNF-α) were measured after administration of miRNA156. RESULTS: Ginseng miRNA156 can accelerate the removal of metabolic waste during exercise. Increase the glycogen reserve in, provide energy for the body, regulate the activity of key gluconeogenesis enzyme phosphorus, improve the energy metabolism system of, and enhance the endurance of fatigue mice. The contents of matrix metalloproteinase 9, superoxide dismutase and malondialdehyde were affected, and the content of TNF-α in the supernatant of RAW264.7 cells was significantly increased, which had certain antioxidant capacity and potential immunomodulatory effects. CONCLUSION: Ginseng miRNA156 has a certain regulatory effect on the energy metabolism and immune function of mice, which makes it possible to regulate the cross-species regulation of ginseng miRNA in theory, provides ideas for ginseng miRNA to become a new pharmacodynamic substance.

The HD-ZIP II gene PaHAT14 increases cuticle deposition by down-regulating ERF gene PaERF105 in Phalaenopsis.

To analyze the gene involved in orchid floral development, a HD-Zip II gene PaHAT14, which specifically and highly expressed in perianth during early flower development was identified from Phalaenopsis. Transgenic Arabidopsis plants expressing 35S::PaHAT14 and 35S::PaHAT14+SRDX (fused with the repressor motif SRDX) exhibited similar altered phenotypes, including small leaves, early flowering, and bending petals with increased cuticle production. This suggests that PaHAT14 acts as a repressor. In contrast, transgenic Arabidopsis plants expressing 35S::PaHAT14+VP16 (fused with the activation domain VP16) exhibited curled leaves, late flowering, and folded petals with decreased cuticle production within hardly opened flowers. Additionally, the expression of the ERF gene DEWAX2, which negatively regulates cuticular wax biosynthesis, was down-regulated in 35S::PaHAT14 and 35S::PaHAT14+SRDX transgenic Arabidopsis, while it was up-regulated in 35S::PaHAT14+VP16 transgenic Arabidopsis. Furthermore, transient overexpression of PaHAT14 in Phalaenopsis petal/sepal increased cuticle deposition due to the down-regulation of PaERF105, a Phalaenopsis DEWAX2 orthologue. On the other hand, transient overexpression of PaERF105 decreased cuticle deposition, whereas cuticle deposition increased and the rate of epidermal water loss was reduced in PaERF105 VIGS Phalaenopsis flowers. Moreover, ectopic expression of PaERF105 not only produced phenotypes similar to those in 35S::PaHAT14+VP16 Arabidopsis but also compensated for the altered phenotypes observed in 35S::PaHAT14 and 35S::PaHAT14+SRDX Arabidopsis. These results suggest that PaHAT14 promotes cuticle deposition by negatively regulating downstream gene PaERF105 in orchid flowers.

[Discussion on reference sample research of traditional Chinese medicine compound preparations developed from catalogued ancient classical prescriptions].

The concept of reference sample was put forward in the Guidance on CMC of Traditional Chinese Medicine Compound Preparations Developed from Catalogued Ancient Classical Prescriptions(Interim). The research on reference sample is a key link in the research and development of traditional Chinese medicine(TCM) compound prescriptions from catalogued ancient classical prescriptions(known as Category 3.1 TCM). This paper discusses the content of research on reference sample by analyzing the characteristics of Category 3.1 TCM and the purpose of research on reference sample. Furthermore, suggestions on the research of reference sample are proposed according to the development and evaluation practice of Category 3.1 TCM and research achievements of TCM regulatory science, aiming to provide reference for colleagues in this industry.