Gender Control of Mouse Embryos by Activation of TLR7/8 on X Sperm via Ligands dsRNA-40 and dsRNA-DR.

Gender control technologies are promising for enhancing the production efficiency of the farm animal industry, and preventing sex-linked hereditary diseases in humans. It has been shown that the X sperm of mammalian animals specifically expresses X-chromosome-derived toll-like receptor 7/8 (TLR7/8), and the activation of TLR7/8 on the X sperm by their agonist, R848, can separate X and Y sperm via the specific inhibition of X sperm motility. The use of R848-preselected sperm for fertilization resulted in sex-ratio-skewed embryos or offspring. In this study, we aimed to investigate whether two other TLR7/8 ligands, double-stranded RNA-40 (dsRNA-40) and double-stranded RNA-DR (dsRNA-DR), are also effective in the separation of mouse X and Y sperm and the subsequent generation of gender-ratio-skewed in vitro fertilization (IVF) embryos. Our results indicated that cholesterol modification significantly enhances the transfection of dsRNA-40 and dsRNA-DR into sperm cells. dsRNA-40 and dsRNA-DR incubation with mouse sperm could separate X and Y sperm by the specific suppression of X sperm motility by decreasing its ATP level and mitochondrial activity. The use of a dsRNA-40- or dsRNA-DR-preselected upper layer of sperm, which predominantly contains high-motility Y sperm, for IVF caused a male-biased sex ratio shift in resulting embryos (with 65.90-74.93% of embryos being male). This study develops a simple new method for the efficient separation of mammalian X and Y sperm, enabling the selective production of male or female progenies.

Luteolin regulates the distribution and function of organelles by controlling SIRT1 activity during postovulatory oocyte aging.

The quality of oocytes determines their development competence, which will be rapidly lost if the oocytes are not fertilized at the proper time after ovulation. SIRT1, one of the sirtuin family members, has been proven to protect the quality of oocytes during postovulatory oocyte aging. However, evidence of the effect of SIRT1 on the activity of organelles including the mitochondria, the endoplasmic reticulum (ER), the Golgi apparatus, and the lysosomes in postovulatory aging oocyte is lacking. In this study, we investigated the distribution and function of organelles in postovulatory aged oocytes and discovered abnormalities. Luteolin, which is a natural flavonoid contained in vegetables and fruits, is an activator of SIRT1. When the oocytes were treated with luteolin, the abnormal distribution of mitochondria, ER, and Golgi complex were restored during postovulatory oocyte aging. The ER stress protein GRP78 and the lysosome protein LAMP1 increased, while the mitochondrial membrane potential and the Golgi complex protein GOLPH3 decreased in aged oocytes, and these were restored by luteolin treatment. EX-527, an inhibitor of SIRT1, disrupted the luteolin-mediated normal distribution and function of mitochondria, ER, Golgi apparatus, and lysosomes. In conclusion, we demonstrate that luteolin regulates the distribution and function of mitochondria, ER, Golgi apparatus, and lysosomes during postovulatory oocyte aging by activating SIRT1.

The Relationship between the Serum NLRP1 Level and Coronary Lesions in Patients with Coronary Artery Disease.

Background: The pathogenesis of coronary artery disease is complex, and inflammation is one of the regulatory factors. The nucleotide-binding oligomerization domain (NOD)-like receptor protein 1 (NLRP1) plays an important role in the cellular inflammatory response, cell apoptosis, cell death, and autoimmune diseases. Whether the level of NLRP1 is related to the severity of coronary artery stenosis in patients with coronary artery disease (CAD) has not been reported. Objective: To test the serum level of NLRP1 in unstable angina (UA) patients and investigate the effect of NLRP1 on coronary stenosis severity of the coronary artery disease (CAD). Methods: 307 patients hospitalized in the Department of Cardiology of the Affiliated Hospital of Xuzhou Medical University for coronary angiography from January 1, 2021, to December 31, 2022 were included. We detect the level of NLRP1 in the serum of the included patients. Patients were divided into UA group and control group according to coronary angiography results and other clinical data. We use logistic regression to screen the influencing factors of UA. Then, subgroups were divided according to the Gensini score and the number of coronary artery lesions, and the difference of serum NLRP1 level between the groups was compared. Spearman correlation analysis was used to explore the correlation between the serum NLRP1 level and Gensini score. We analyze the diagnostic value of NLRP1 for UA by drawing ROC curve. Results: The median level of serum NLRP1 in patients with UA (n = 257) was 49.71 pg/ml, IQR 30.15, 80.21, and that in patients without UA (n = 50) was 24.75 pg/ml, IQR 13.49, 41.95. Serum NLRP1 levels were significantly different among different subgroups. The patient's Gensini score was correlated with the patient's serum NLRP1 level. Conclusion: The serum NLRP1 level is increased in patients with UA, which is increased with the increasing severity of coronary lesions.

Knockout of Rlim Results in a Sex Ratio Shift toward Males but Superovulation Cannot Compensate for the Reduced Litter Size.

Technologies that can preselect offspring gender hold great promise for improving farm animal productivity and preventing human sex-related hereditary diseases. The maternal Rlim allele is required for imprinted X-chromosome inactivation, which is essential for the normal development of female mouse embryos. In this study, we inactivated the maternal Rlim allele in embryos by crossing a male transgenic mouse line carrying an X-linked CMV-Cre transgene with a female line carrying a loxP-flanked Rlim gene. Knockout of the maternal Rlim gene in embryos resulted in a male-biased sex ratio skew in the offspring. However, it also reduced litter size, and this effect was not compensated for by superovulation in the mother mice. In addition, we showed that siRNA-mediated knockdown of Rlim in mouse embryos leads to the birth of male-only progenies. This study provides a new promising method for male-biased sex selection, which may help to improve the productivity in livestock and prevent sex-associated hereditary diseases in humans.

Early monitoring of parasitism by Aphidiinae parasitoids on the grain aphid Sitobion miscanthi in wheat fields using DNA barcoding.

BACKGROUND: Sitobion miscanthi is a major wheat pest at the grain-filling stage found in China. Identifying parasitoid species and understanding parasitism rates are keys to controlling the aphids via natural enemies in the wheat field. RESULTS: In the present study, a method based on DNA barcoding for early determination of the community composition of Aphidiinae parasitoids and parasitism on the aphid was developed. The proposed method detected Aphidius gifuensis as the predominant parasite, with parasitism rates of 40.1 ± 2.8% in 2019 and 65.7 ± 3.7% in 2022, and found that the rate varied significantly among different wheat varieties. COI primers efficiently amplified the Aphidiinae parasitoids COI fragments and amplified the aphid COI fragments derived from parasitized (mummified) S. miscanthi. Thus, the COI barcode is not sufficiently specific to unambiguously detect immature parasitoids inside their S. miscanthi hosts. However, it can be used to detect the DNA extracted from mummified aphids. In contrast, the 16S and LWRh primers effectively amplified and identified the parasitoids in parasitized aphids. The 16S primer was reliable even in the early stages of parasitism (24 h) and for DNA samples stored at -20 °C for 5 days. The three barcodes from COI, 16S, and LWRh genes could not clearly distinguish a few certain Aphidiinae species owing to relatively low intraspecific and interspecific diversity. CONCLUSION: The morphological features remain indispensable when identifying Aphidiinae species. Nonetheless, the COI and 16S primers could be used in combination for monitoring the parasitism rates on S. miscanthi in wheat fields. © 2022 Society of Chemical Industry.

KLF9 Aggravates Streptozotocin-Induced Diabetic Cardiomyopathy by Inhibiting PPARγ/NRF2 Signalling.

AIMS: Krüppel-like Factor 9 (KLF9) is a transcription factor that regulates multiple disease processes. Studies have focused on the role of KLF9 in the redox system. In this study, we aimed to explore the effect of KLF9 on diabetic cardiomyopathy. METHODS AND RESULTS: Cardiac-specific overexpression or silencing of KLF9 in C57BL/6 J mice was induced with an adeno-associated virus 9 (AAV9) delivery system. Mice were also subjected to streptozotocin injection to establish a diabetic cardiomyopathy model. In addition, neonatal rat cardiomyocytes were used to assess the possible role of KLF9 in vitro by incubation with KLF9 adenovirus or small interfering RNA against KLF9. To clarify the involvement of peroxisome proliferator-activated receptors (PPARγ), mice were subjected to GW9662 injection to inhibit PPARγ. KLF9 was upregulated in the hearts of mice with diabetic cardiomyopathy and in cardiomyocytes. In addition, KLF9 overexpression in the heart deteriorated cardiac function and aggravated hypertrophic fibrosis, the inflammatory response and oxidative stress in mice with diabetic cardiomyopathy. Conversely, cardiac-specific silencing of KLF9 ameliorated cardiac dysfunction and alleviated hypertrophy, fibrosis, the cardiac inflammatory response and oxidative stress. In vitro, KLF9 silencing in cardiomyocytes enhanced inflammatory cytokine release and oxidative stress; KLF9 overexpression increased these detrimental responses. Moreover, KLF9 was found to regulate the transcription of PPARγ, which suppressed the expression and nuclear translocation of nuclear Factor E2-related Factor 2 (NRF2). In mice injected with a PPARγ inhibitor, the protective effects of KLF9 knockdown on diabetic cardiomyopathy were counteracted by GW9662 injection. CONCLUSIONS: KLF9 aggravates cardiac dysfunction, the inflammatory response and oxidative stress in mice with diabetic cardiomyopathy. KLF9 may become a therapeutic target for diabetic cardiomyopathy.

Effects of Crop Resistance on the Tritrophic Interactions between Wheat Lines, Schizaphis graminum (Hemitera: Aphididae), and Propylaea japonica (Coleoptera: Coccinellidae).

Crop resistance and biological control are both considered efficient and environmentally friendly methods of sustainable pest control. In this study, we aimed at investigating the direct influence of four wheat lines with varying resistance level on the life-history traits of the greenbug, Schizaphis graminum, and the mediational effect on the functional response of a predatory ladybird, Propylaea japonica, under laboratory conditions. Results showed that the aphid fitness was the lowest for aphids that had been feeding on wheat line '98-10-19' for one year. These aphids had the longest development time, and least adult mass, minimal mean relative growth rate, and lowest reproductive fitness. In contrast, the aphids that fed on wheat line '98-10-30' were the fittest, with the shortest development time and highest levels of reproductive fitness. The predatory activities of the ladybeetle, especially the adult male significantly decreased following the consumption of aphids belonging to the '98-10-19'-acclimated population. However, there were no significant differences in predatory efficiency (net attack frequency) among the four aphid acclimated populations. Our results showed that the wheat line '98-10-19' has a relative higher resistance to S. graminum than the other three wheat lines, which could further decrease the amount of prey available for consumption. However, the ecological effect of the resistance of '98-10-19' to S. graminum posed no negative influence on the biocontrol potential of P. japonica to these aphids, as their predatory efficiency increases at the fourth instar larvae phase.

Construction and analysis of lncRNA-associated ceRNA network identified potential prognostic biomarker in gastric cancer.

BACKGROUND: Long non-coding RNAs (lncRNAs) are defined as non-coding RNA (ncRNA) with transcripts longer than 200 nucleotides with tissue specificity. Recently it has been found participate in cancer tumorigenesis and progression via transcriptional regulation, post-transcriptional regulation and epigenetic gene regulation. Competitive endogenous RNA (ceRNA) hypothesis assume that lncRNAs compete the target RNA by sponging the common miRNA response elements (MREs) to complete the post-transcriptional regulation. To explore the function and mechanisms of lncRNAs as ceRNAs in gastric cancer (GC), this study performed a genome-wide analysis. METHODS: The lncRNAs, mRNAs and microRNAs (miRNAs) profiles of 375 GC samples and 32 normal samples were obtained from The Cancer Genome Atlas (TCGA) Stomach Adenocarcinoma (STAD) datasets. The data was standardized with a cross match in the miRBase (a database at http://www.mirbase.org/), which made 365 samples as the analysis objects. We identify differentially expressed RNAs (DERNAs), including differentially expressed mRNAs (DEmRNAs), differentially expressed miRNAs (DEmiRNAs) and differentially expressed lncRNAs (DElncRNAs) by applying edge R package with thresholds of |log2FC| >2 and false discovery rate (FDR) <0.01. The potential RNAs for the gastric ceRNA network were screened out from the DERNAs based on "ceRNA hypothesis". The further construction of the network and analysis of its topological properties were performed by Cytoscape. Gene oncology (GO) function enrichment was analyzed by BINGO plugin of Cytoscape. Survival analysis was estimated according to Kaplan-Meier curve analysis. RESULTS: The constructed gastric ceRNA network involved 61 mRNAs, 44 lncRNAs and 22 miRNAs. Five lncRNAs out of the DElncRNAs, namely MIR100HG, MAGI2-AS3, AC080038.1, AC010478.1 and MEF2C-AS1, were found mostly involved in the network. The lncRNA AL139147 were detected negatively correlated with overall survival (log-rank, P<0.05). CONCLUSIONS: In conclusion, our study identified promising lncRNAs, which might be potential diagnostic biomarker and therapeutic targets and contribute to further understanding of the ceRNA pathogenesis in GC and guide for further investigation.

Ionic liquid-based single-drop liquid-phase microextraction combined with high-performance liquid chromatography for the determination of sulfonamides in environmental water.

A simple, rapid and environment-friendly technique of single-drop liquid-phase microextraction has been developed for the determination of sulfonamides in environmental water. Several important parameters including stirring rate, extraction solvent, extraction pH, salinity and extraction time were optimized to maximize the extract efficiency. Extraction solvent 1-octyl-3-methylimidazolium hexafluorophosphate [C(8) MIM][PF(6) ] ionic liquid showed better extraction efficiency than 1-butyl-3-methylimidazolium hexafluorophosphate [C(4) MIM][PF(6) ] and 1-octanol. The optimum experimental conditions were: pH, 4.5; sodium chloride content, 36% w/v; extraction time, 20 min. This method provided low detection limits (0.5-1 ng/mL), good repeatability (the RSD ranging from 4.2 to 9.9%, n=5) and wide linear range (1-1500 ng/mL), with determination coefficients (r(2) ) higher than 0.9989 for all the target compounds. Real sample analysis showed relative recoveries between 63.5 and 115.8% for all the target compounds.

[Stromal tumor of the seminal vesicle: a case report and literature review].

OBJECTIVE: To investigate the clinical manifestations, pathological features, diagnosis and treatment of stromal tumor in the seminal vesicle. METHODS: We retrospectively analyzed 1 case of stromal tumor of the seminal vesicle, reviewed relevant domestic and international literature, and summarized the clinical manifestations, pathological features, diagnosis and treatment of the tumor. RESULTS: The patient was a 50 years old male, who underwent excision of the tumor together with the seminal vesicle. Pathology showed it to be stromal tumor of the seminal vesicle. Ultrasonography and CT found no recurrence 10 months after surgery. CONCLUSION: Stromal tumor of the seminal vesicle is rare and easy to be misdiagnosed. Digital rectal examination, and ultrasonography, CT and MRI of the urinary system are useful for its diagnosis. The currently accepted treatment is surgical removal of the tumor, and the prognosis is good.

[Minimally invasive surgery for colorectal cancer with liver metastasis].

OBJECTIVE: To investigate the minimal invasive surgery for simultaneously treating synchronous colorectal cancer with liver metastasis. METHODS: From the clinical data of our department between March 2006 to June 2008, retrospectively reviewed and analyzed 5 typical cases of synchronous colorectal cancer with liver metastasis. All were treated with 5 different strategies of minimal invasive surgery. RESULTS: All underwent laparoscopic colorectal surgery, i.e. open hepatectomy, laparoscopic-assisted hepatectomy, laparoscopic hepatectomy, laparoscopic hepatectomy plus radiofrequency ablation and multiple therapy respectively. The mean operative time was 177 minutes, the mean blood loss 126 ml, the time of gastrointestinal function recovery 48-72 hours and the mean hospital stay 7 days. In the follow-up of 10-27 months for all 5 patients, one suffered hepatic recurrence and one died. CONCLUSION: Simultaneous minimal invasive surgery is an optional and ideal method for treating synchronous colorectal cancer with liver metastasis.