Synergistic Effect of Layered Double Hydroxides Nanodosage Form to Induce Apoptosis and Ferroptosis in Breast Cancer.

Background: Breast cancer is the most common cancer in women and one of the leading causes of cancer death worldwide. Ferroptosis, a promising mechanism of killing cancer cells, has become a research hotspot in cancer therapy. Simvastatin (SIM), as a potential new anti-breast cancer drug, has been shown to cause ferroptosis of cancer cells and inhibit breast cancer metastasis and recurrence. The purpose of this study is to develop a novel strategy boosting ferroptotic cascade for synergistic cancer therapy. Methods: In this paper, iron base form of layered double hydroxide supported simvastatin (LDHs-SIM) was synthesized by hydrothermal co-precipitation method. The characterization of LDHs-SIM were assessed by various analytical techniques, including ultraviolet-visible (UV-vis) spectroscopy, X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, and transmission electron microscopy (TEM). Biological activity, ferroptosis mechanism and biocompatibility were analyzed through in vivo and in vitro analysis, so as to evaluate its therapeutic effect on breast cancer. Results: The constructed LDHs-SIM nanosystem can not only release SIM through mevalonate (MVA) pathway, inhibit the expression of glutathione peroxidase 4 (GPX4), inhibit the expression of SLC7A11 and reduce the synthesis efficiency of GSH, but also promote the accumulation of Fe2+ in cells through the release of Fe3+, and increase the intracellular ROS content. In addition, LDHs-SIM nanosystem can induce apoptosis of breast cancer cells to a certain extent, and achieve the synergistic effect of apoptosis and ferroptosis. Conclusion: In the present study, we demonstrated that nanoparticles of layered double hydroxides (LDHs) loaded with simvastatin were more effective than a free drug at inhibiting breast cancer cell growth, In addition, superior anticancer therapeutic effects were achieved with little systemic toxicity, indicating that LDHs-SIM could serve as a safe and high-performance platform for ferroptosis-apoptosis combined anticancer therapy.

Non-targeted metabolomics revealed novel links between serum metabolites and primary ovarian insufficiency: a Mendelian randomization study.

Background: Primary ovarian insufficiency (POI) is a common clinical endocrine disorder with a high heterogeneity in both endocrine hormones and etiological phenotypes. However, the etiology of POI remains unclear. Herein, we unraveled the causality of genetically determined metabolites (GDMs) on POI through Mendelian randomization (MR) study with the overarching goal of disclosing underlying mechanisms. Methods: Genetic links with 486 metabolites were retrieved from GWAS data of 7824 European participants as exposures, while GWAS data concerning POI were utilized as the outcome. Via MR analysis, we selected inverse-variance weighted (IVW) method for primary analysis and several additional MR methods (MR-Egger, weighted median, and MR-PRESSO) for sensitivity analyses. MR-Egger intercept and Cochran's Q statistical analysis were conducted to assess potential heterogeneity and pleiotropy. In addition, genetic variations in the key target metabolite were scrutinized further. We conducted replication, meta-analysis, and linkage disequilibrium score regression (LDSC) to reinforce our findings. The MR Steiger test and reverse MR analysis were utilized to assess the robustness of genetic directionality. Furthermore, to deeply explore causality, we performed colocalization analysis and metabolic pathway analysis. Results: Via IVW methods, our study identified 33 metabolites that might exert a causal effect on POI development. X-11437 showed a robustly significant relationship with POI in four MR analysis methods (P IVW=0.0119; P weighted-median =0.0145; PMR-Egger =0.0499; PMR-PRESSO =0.0248). Among the identified metabolites, N-acetylalanine emerged as the most significant in the primary MR analysis using IVW method, reinforcing its pivotal status as a serum biomarker indicative of an elevated POI risk with the most notable P-value (P IVW=0.0007; PMR-PRESSO =0.0022). Multiple analyses were implemented to further demonstrate the reliability and stability of our deduction of causality. Reverse MR analysis did not provide evidence for the causal effects of POI on 33 metabolites. Colocalization analysis revealed that some causal associations between metabolites and POI might be driven by shared genetic variants. Conclusion: By incorporating genomics with metabolomics, this study sought to offer a comprehensive analysis in causal impact of serum metabolome phenotypes on risks of POI with implications for underlying mechanisms, disease screening and prevention.

High AMH levels are associated with gestational hypertension in patients with PCOS who underwent IVF/ICSI-ET.

BACKGROUND: Patients with polycystic ovary syndrome (PCOS) have a higher risk of obstetric complications. The association between anti-Müllerian hormone (AMH) and gestational hypertension in these patients is poorly understood. OBJECTIVE: To determine the association between serum AMH levels and gestational hypertension in patients with PCOS undergoing fresh embryo transfer. METHODS: This retrospective study included 649 patients with PCOS who had singleton live births after undergoing fresh embryo transfers. The association of AMH with gestational hypertension in these patients was estimated before and after propensity score matching (PSM). RESULTS: Patients with gestational hypertension had higher AMH levels than those without gestational hypertension. In single-factor logistic regression, the odds of gestational hypertension increased by 11.7% and 18.6% for every 1 ng/mL increase in AMH before and after adjusting for confounding factors [OR 1.117, 95% CI(1.025, 1.217), P = 0.012; adjusted OR 1.186, 95% CI(1.061, 1.327), adjusted P = 0.003], respectively. The odds of gestational hypertension increased more than 100% [adjusted OR 2.635, 95% CI(1.132, 6.137), adjusted P = 0.025] in the 75th percentile group (>9.30 ng/ml) and more than thrice [adjusted OR 4.75, 95% CI(1.672, 13.495), adjusted P = 0.003] in the 90th percentile group (>12.31 ng/ml) as compared to the without-gestational hypertension group. AMH level was still associated with gestational hypertension after PSM. The area under the curve of AMH predicting gestational hypertension was 0.654 [95% CI (0.532, 0.776), P = 0.011] with an optimal cutoff value of 11.975 ng/mL. CONCLUSIONS: High serum AMH level pre-pregnancy (especially at levels > 9.30 ng/mL) indicates a high odds of gestational hypertension in patients with PCOS undergoing fresh embryo transfer.

Adrenal pheochromocytoma impacts three main pathways: cysteine-methionine, pyrimidine, and tyrosine metabolism. / è‚¾ä¸Šè ºå—œé“¬ç»†èƒžç˜¤å½±å“ä¸‰æ¡ä¸»è¦ä»£è°¢é€šè·¯ï¼šåŠèƒ±æ°¨é ¸-è›‹æ°¨é ¸ä»£è°¢ã€å˜§å•¶ä»£è°¢å’Œé ªæ°¨é ¸ä»£è°¢é€šè·¯.

Pheochromocytomas and paragangliomas (PPGLs) cause symptoms by altering the circulation levels of catecholamines and peptide hormones. Currently, the diagnosis of PPGLs relies on diagnostic imaging and the detection of catecholamines. In this study, we used ultra-performance liquid chromatography (UPLC)/quadrupole time-of-flight mass spectrometry (Q-TOF MS) analysis to identify and measure the perioperative differential metabolites in the plasma of adrenal pheochromocytoma patients. We identified differentially expressed genes by comparing the transcriptomic data of pheochromocytoma with the normal adrenal medulla. Through conducting two steps of metabolomics analysis, we identified 111 differential metabolites between the healthy group and the patient group, among which 53 metabolites were validated. By integrating the information of differential metabolites and differentially expressed genes, we inferred that the cysteine-methionine, pyrimidine, and tyrosine metabolism pathways were the three main metabolic pathways altered by the neoplasm. The analysis of transcription levels revealed that the tyrosine and cysteine-methionine metabolism pathways were downregulated in pheochromocytoma, whereas the pyrimidine pathway showed no significant difference. Finally, we developed an optimized diagnostic model of two metabolites, L-dihydroorotic acid and vanylglycol. Our results for these metabolites suggest that they may serve as potential clinical biomarkers and can be used to supplement and improve the diagnosis of pheochromocytoma.

Elevated sperm DNA fragmentation is correlated with an increased chromosomal aneuploidy rate of miscarried conceptus in women of advanced age undergoing fresh embryo transfer cycle.

Background: Male sperm DNA fragmentation (SDF) may be associated with assisted reproductive technology (ART) outcomes, but the impact of SDF on the occurrence of aneuploid-related miscarriage remains controversial. Methods: Genome-wide single-nucleotide polymorphism-based chromosomal microarray analysis was performed on 495 miscarried chorionic villus samples undergone IVF/ICSI treatment from the Reproductive Medicine Center of the First Affiliated Hospital of Zhengzhou University. SDF was assessed using sperm chromatin structure assay. Patients were divided into four groups according to embryo transfer cycle type and maternal age, and the correlation between SDF and chromosome aberration was analyzed. A receiver operating characteristic (ROC) curve was utilized to find the optimal threshold. Results: Total chromosomal aneuploidy rate was 54.95%, and trisomy was the most common abnormality (71.32%). The chromosomally abnormal group had higher SDF than the normal group (11.42% [6.82%, 16.54%] vs. 12.95% [9.61%, 20.58%], P = 0.032). After grouping, elevated SDF was significantly correlated with an increasing chromosome aneuploidy rate only in women of advanced age who underwent fresh embryo transfer (adjusted odds ratio:1.14 [1.00-1.29], adjusted-P = 0.045). The receiver operating characteristic curve showed that SDF can predict the occurrence of chromosomal abnormality of miscarried conceptus in this group ((area under the curve = 0.76 [0.60-0.91], P = 0.005), and 8.5% was the optimum threshold. When SDF was ≥ 8.5%, the risk of such patients increased by 5.76 times (adjusted odds ratio: 6.76 [1.20-37.99], adjusted-P = 0.030). Conclusion: For women of advanced maternal age undergoing fresh embryo transfer, older oocytes fertilized using sperm with high SDF in IVF/ICSI treatment might increase the risk of chromosomal abnormality in miscarried conceptus.

Adrenal pheochromocytoma impacts three main pathways: cysteine-methionine, pyrimidine, and tyrosine metabolism. / è‚¾ä¸Šè ºå—œé“¬ç»†èƒžç˜¤å½±å“ä¸‰æ¡ä¸»è¦ä»£è°¢é€šè·¯ï¼šåŠèƒ±æ°¨é ¸-è›‹æ°¨é ¸ä»£è°¢ã€å˜§å•¶ä»£è°¢å’Œé ªæ°¨é ¸ä»£è°¢é€šè·¯.

Pheochromocytomas and paragangliomas (PPGLs) cause symptoms by altering the circulation levels of catecholamines and peptide hormones. Currently, the diagnosis of PPGLs relies on diagnostic imaging and the detection of catecholamines. In this study, we used ultra-performance liquid chromatography (UPLC)/quadrupole time-of-flight mass spectrometry (Q-TOF MS) analysis to identify and measure the perioperative differential metabolites in the plasma of adrenal pheochromocytoma patients. We identified differentially expressed genes by comparing the transcriptomic data of pheochromocytoma with the normal adrenal medulla. Through conducting two steps of metabolomics analysis, we identified 111 differential metabolites between the healthy group and the patient group, among which 53 metabolites were validated. By integrating the information of differential metabolites and differentially expressed genes, we inferred that the cysteine-methionine, pyrimidine, and tyrosine metabolism pathways were the three main metabolic pathways altered by the neoplasm. The analysis of transcription levels revealed that the tyrosine and cysteine-methionine metabolism pathways were downregulated in pheochromocytoma, whereas the pyrimidine pathway showed no significant difference. Finally, we developed an optimized diagnostic model of two metabolites, L-dihydroorotic acid and vanylglycol. Our results for these metabolites suggest that they may serve as potential clinical biomarkers and can be used to supplement and improve the diagnosis of pheochromocytoma.

Sestrin2 and Sestrin3 protect spermatogenesis against heat-induced meiotic defects.

Heat stress (HS) induces testicular oxidative stress, impairs spermatogenesis, and increases the risk of male infertility. Recent studies have highlighted the antioxidative properties of the Sestrins family in reducing cellular oxidative damage. However, the role of Sestrins (Sestrin1, 2, and 3) in the testicular response to heat stress remains unclear. Here, we found that Sestrin 2 and 3 were highly expressed in the testis relative to Sestrin1. Then, the Sestrin2-/- and Sestrin3-/- mice were generated by CRISPR/Cas9 to investigate the role of them on spermatogenesis after HS. Our data showed that Sestrin2-/- and Sestrin3-/- mice testes exhibited more severe damage manifested by exacerbated loss of germ cells and higher levels of oxidative stress as compared to wild-type counterparts after HS. Notably, Sestrin2-/- and Sestrin3-/- mice underwent a remarkable increase in heat-induced spermatocyte apoptosis than that of controls. Mechanistically, the transcriptome landscape of spermatocytes and chromosome spreading showed that loss of Sestrin2 and Sestrin3 exacerbated meiotic failure by compromising DNA double-strand breaks (DSBs) repair after heat stress. Taken together, our work demonstrated a critical protective function of Sestrin2 and Sestrin3 in mitigating the impairments of spermatogenesis against heat stress.

Circular RNAs in Breast Cancer: An Update.

Breast cancer (BC), characterized by high heterogeneity, is the most commonly reported malignancy among females across the globe. Every year, many BC patients die owing to delayed diagnosis and treatment. Increasing researches have indicated that aberrantly expressed circular RNAs (circRNAs) are implicated in the tumorigenesis and progression of various tumors, including BC. Hence, this article provides a summary of the biogenesis and functions of circRNAs, as well as an examination of how circRNAs regulate the progression of BC. Moreover, circRNAs have aroused incremental attention as potential diagnostic and prognostic biomarkers for BC. Exosomes enriched with circRNAs can be secreted into the tumor microenvironment to mediate intercellular communication, affecting the progression of BC. Detecting the expression levels of exosomal circRNAs may provide reference for BC diagnosis and prognosis prediction. Illuminating insights into the earlier diagnosis and better treatment regimens of BC will be potentially available following elucidation of deeper regulatory mechanisms of circRNAs in this malignancy.

Sestrin1, 2, and 3 are dispensable for female fertility in mice.

BACKGROUND: Sestrins have been implicated in regulating aging in various organs through multiple pathways. However, their roles in ovarian aging remain unrevealed. METHODS: Female Sestrin1-/-, Sestrin2-/-, and Sestrin3-/- mice were generated using the CRISPR-Cas9 system. Body weights, little sizes, ovarian weights, estrous cyclicity, and follicle number in female mice were observed. ELISA was utilized to measure serum anti-Müllerian hormone (AMH) levels. Real time PCR, western blot, immunofluorescence, and Masson trichrome staining were employed for assessment of aging-related change. RESULTS: The deletion of Sestrin 1, 2, or 3 had no discernible impact on body weights,or serum AMH levels in female mice at the age of 12 months. And there were no discernible differences in litter sizes or estrous cyclicity which were assessed at the age of 8 months. At the age of 12 months, no significant differences were observed in ovarian weights or follicle numbers among the knockout mice. Consistently, the extent of fibrosis within the ovaries remained comparable across all experimental groups at this age. Additionally, autophagy, apoptosis, DNA damage, and inflammation within the ovaries were also found to be comparable to those in wild-type mice of the same age. CONCLUSIONS: The loss of Sestrin 1, 2, or 3 does not exert a noticeable influence on ovarian function during the aging process. Sestrin1, 2, and 3 are not essential for female fertility in mice.

NADase CD38 is a key determinant of ovarian aging.

The ovary ages earlier than most other tissues, yet the underlying mechanisms remain elusive. Here a comprehensive analysis of transcriptomic landscapes in different organs in young and middle-aged mice revealed that the ovaries showed earlier expression of age-associated genes, identifying increased NADase CD38 expression and decreased NAD+ levels in the ovary of middle-aged mice. Bulk and single-cell RNA sequencing revealed that CD38 deletion mitigated ovarian aging, preserving fertility and follicle reserve in aged mice by countering age-related gene expression changes and intercellular communication alterations. Mechanistically, the earlier onset of inflammation induced higher expression levels of CD38 and decreased NAD+ levels in the ovary, thereby accelerating ovarian aging. Consistently, pharmacological inhibition of CD38 enhanced fertility in middle-aged mice. Our findings revealed the mechanisms underlying the earlier aging of the ovary relative to other organs, providing a potential therapeutic target for ameliorating age-related female infertility.

Comparing the clinical and singleton neonatal outcomes in male infertility patients with Oligoasthenospermia, OA, or NOA following fresh ICSI-ET using different sources of sperm.

Objective: To investigate clinical and singleton newborn outcomes in fresh cycles of embryo transfer after intracytoplasmic sperm injection (ICSI-ET) with diverse sperm sources (ejaculate, epididymis, and testis) in patients with Oligoasthenospermia, obstructive azoospermia (OA) or non-obstructive azoospermia (NOA). Methods: Patients who received fresh ICSI-ET for the first time at the First Affiliated Hospital of Zhengzhou University Reproductive Medicine Center between June 2011 and June 2021 were selected for this 10-year retrospective cohort analysis. After propensity score matching, only 1630 cycles were included in the investigation of ICS-ET clinical and singleton newborn outcomes in patients with Oligoasthenospermia, OA, and NOA using sperm from diverse sperm sources. Results: After propensity score matching, our data revealed a negligible difference in baseline and cycle parameters among groups. In patients with Oligoasthenospermia and OA, different sperm sources do not appear to influence clinical pregnancy rates and live birth rates, nor do they influence newborn outcomes, such as newborn weight, premature birth rate, and neonatal sex ratio in singleton births, except for OA patients who use epididymal sperm having higher low birth weight (LBW) rates in singleton pregnancies than those who use testicular sperm. In addition, clinical pregnancy rates, live birth rates, singleton gestation birth weights, premature birth rates, and neonatal sex ratios were similar between patients with Oligoasthenospermia, OA, and NOA using testicular sperm. Conclusions: Regardless of the type of male infertility (Oligoasthenospermia, OA, NOA) or sperm sources (ejaculate, epididymis, testis), a successful ICSI-ET procedure can result in similar clinical and neonatal outcomes, such as clinical pregnancy rate, live birth rate, abortion rate, neonatal birth weight and sex ratio of singleton pregnancies.

A Machine Learning-Based Preclinical Osteoporosis Screening Tool (POST): Model Development and Validation Study.

Background: Identifying persons with a high risk of developing osteoporosis and preventing the occurrence of the first fracture is a health care priority. Most existing osteoporosis screening tools have high sensitivity but relatively low specificity. Objective: We aimed to develop an easily accessible and high-performance preclinical risk screening tool for osteoporosis using a machine learning-based method among the Hong Kong Chinese population. Methods: Participants aged 45 years or older were enrolled from 6 clinics in the 3 major districts of Hong Kong. The potential risk factors for osteoporosis were collected through a validated, self-administered questionnaire and then filtered using a machine learning-based method. Bone mineral density was measured with dual-energy x-ray absorptiometry at the clinics; osteoporosis was defined as a t score of -2.5 or lower. We constructed machine learning models, including gradient boosting machines, support vector machines, and naive Bayes, as well as the commonly used logistic regression models, for the prediction of osteoporosis. The best-performing model was chosen as the final tool, named the Preclinical Osteoporosis Screening Tool (POST). Model performance was evaluated by the area under the receiver operating characteristic curve (AUC) and other metrics. Results: Among the 800 participants enrolled in this study, the prevalence of osteoporosis was 10.6% (n=85). The machine learning-based Boruta algorithm identified 15 significantly important predictors from the 113 potential risk factors. Seven variables were further selected based on their accessibility and convenience for daily self-assessment and health care practice, including age, gender, education level, decreased body height, BMI, number of teeth lost, and the intake of vitamin D supplements, to construct the POST. The AUC of the POST was 0.86 and the sensitivity, specificity, and accuracy were all 0.83. The positive predictive value, negative predictive value, and F1-score were 0.41, 0.98, and 0.56, respectively. Conclusions: The machine learning-based POST was conveniently accessible and exhibited accurate discriminative capabilities for the prediction of osteoporosis; it might be useful to guide population-based preclinical screening of osteoporosis and clinical decision-making.

Melatonin Potentiates Sensitivity to 5-Fluorouracil in Gastric Cancer Cells by Upregulating Autophagy and Downregulating Myosin Light-Chain Kinase.

5-Fluorouracil is an effective chemotherapeutic drug for gastric cancer. However, the acquisition of chemotherapeutic resistance remains a challenge in treatment. Melatonin can enhance the therapeutic effect of 5-fluorouracil; however, the underlying mechanisms are not well understood. We investigated the effects of combinations of melatonin and 5-fluorouracil on the proliferation, migration and invasion of gastric cancer cells. Melatonin significantly potentiated the 5-fluorouracil-mediated inhibition of proliferation, migration and invasion in gastric cancer cells, which potentiates sensitivity to 5-FU by promoting the activation of Beclin-1-dependent autophagy and targeting the myosin light-chain kinase (MLCK) signaling pathway. Previous studies have shown that autophagy might be associated with the MLCK signaling pathway. The autophagy inhibitor, 3-methyladenine, effectively rescued the migratory and invasive capabilities of gastric cancer cells, while also reducing expression level of MLCK and the phosphorylation level of MLC. This indicates that autophagy is involved in tumor metastasis, which may be related to inhibition of the MLCK signaling pathway. Our findings indicate that melatonin can improve the effectiveness of 5-fluorouracil in gastric cancer and could be used as a supplemental agent in the treatment of gastric cancer with 5-fluorouracil.

The impact of female BMI on sperm DNA damage repair ability of oocytes and early embryonic development potential in intracytoplasmic sperm injection cycles.

Background: Obesity adversely influences the quality of oocytes and embryos and can affect DNA repair in embryos, leading to reproductive issues. However, the effects of body mass index (BMI) on DNA repair ability in oocytes during intracytoplasmic sperm injection (ICSI) cycles have not yet been investigated. Therefore, this retrospective study aimed to analyze the influence of sperm DNA damage on embryo development and reproductive outcomes in overweight/obese and normal-weight women in ICSI cycles. Methods: A total of 1,141 patients who received the first fresh ICSI cycle treatments were recruited from July 2017 to July 2021. Based on the BMI of the women, all patients were divided into normal weight (18.5≤BMI<25 kg/m2; n=824; 72.22%) and overweight/obese (BMI≥25 kg/m2; n=317; 27.78%) groups. Furthermore, according to the sperm DNA fragmentation index (DFI), these two groups were subdivided into two subgroups: DFI<30% and DFI≥30%. Results: In the normal-weight women group, the embryonic development and reproductive outcomes of ICSI cycles were not statistically different between the two subgroups (DFI<30% and DFI≥30%). However, in the overweight/obese women group, couples with a sperm DFI≥30% had a significantly lower fertilization rate (76% vs. 72.7%; p=0.027), cleavage rate (98.7% vs. 97.2%; p=0.006), and high-quality embryo rate (67.8% vs. 62.6%; p=0.006) than couples with a sperm DFI<30%. Conclusion: When injected sperm with high DFI into the oocytes of overweight/obese women, resulting in lower fertilization, cleavage, and high-quality embryo rates in ICSI cycles, and the decreased early developmental potential of embryos from overweight/obese patients may be caused by the diminished capacity of oocytes to repair sperm DNA damage.

High sperm DNA fragmentation increased embryo aneuploidy rate in patients undergoing preimplantation genetic testing.

RESEARCH QUESTION: Is high sperm DNA fragmentation (SDF) associated with a high embryonic aneuploidy rate in patients undergoing intracytoplasmic sperm injection (ICSI)-preimplantation genetic testing (PGT)? DESIGN: This was a retrospective study of 426 couples with normal karyotypes undergoing ICSI-PGT at the authors' centre from March 2017 to March 2021. SDF was assessed using the sperm chromatin structure assay. The population was divided into low and high SDF groups according to cut-off values found by the receiver operating characteristic (ROC) curve. A 1:1 ratio propensity score matching (PSM) method was used to control for potential confounding factors, and a generalized linear mixed model was established to evaluate the relationship between SDF and the embryonic aneuploidy rate. RESULTS: The ROC curve indicated a threshold of 30%. In total, 132 couples were included after PSM, and the high SDF group (>30%) had significantly higher SDF (40.74% ± 9.78% versus 15.54% ± 7.86%, P < 0.001) and a higher embryo aneuploidy rate (69.36% versus 53.96%, P < 0.001) compared with the low SDF group (≤30%). The two pronuclear fertilization rate, cleavage rate, rate of high-quality embryos at day 3 rate, blastocyst rate, biochemical pregnancy rate, clinical pregnancy rate, miscarriage rate, live birth rate, caesarean section rate, preterm birth rate, singleton rate and low birthweight rate were similar in both groups (P > 0.05). After PSM, SDF > 30% was significantly correlated with an increased embryo aneuploidy rate after adjusting for all confounding variables (adjusted odds ratio 1.70, 95% CI 1.00-2.88, P = 0.049). CONCLUSIONS: SDF > 30% was associated with an increased embryo aneuploidy rate in couples with normal karyotypes undergoing PGT, but did not affect embryonic and clinical outcomes after transfer of euploid embryos.

Sequence tolerance of immunoglobulin variable domain framework regions to noncanonical intradomain disulfide linkages.

Most immunoglobulin (Ig) domains bear only a single highly conserved canonical intradomain, inter-ß-sheet disulfide linkage formed between Cys23-Cys104, and incorporation of rare noncanonical disulfide linkages at other locations can enhance Ig domain stability. Here, we exhaustively surveyed the sequence tolerance of Ig variable (V) domain framework regions (FRs) to noncanonical disulfide linkages. Starting from a destabilized VH domain lacking a Cys23-Cys104 disulfide linkage, we generated and screened phage-displayed libraries of engineered VHs, bearing all possible pairwise combinations of Cys residues in neighboring ß-strands of the Ig fold FRs. This approach identified seven novel Cys pairs in VH FRs (Cys4-Cys25, Cys4-Cys118, Cys5-Cys120, Cys6-Cys119, Cys22-Cys88, Cys24-Cys86, and Cys45-Cys100; the international ImMunoGeneTics information system numbering), whose presence rescued domain folding and stability. Introduction of a subset of these noncanonical disulfide linkages (three intra-ß-sheet: Cys4-Cys25, Cys22-Cys88, and Cys24-Cys86, and one inter-ß-sheet: Cys6-Cys119) into a diverse panel of VH, VL, and VHH domains enhanced their thermostability and protease resistance without significantly impacting expression, solubility, or binding to cognate antigens. None of the noncanonical disulfide linkages identified were present in the natural human VH repertoire. These data reveal an unexpected permissiveness of Ig V domains to noncanonical disulfide linkages at diverse locations in FRs, absent in the human repertoire, whose presence is compatible with antigen recognition and improves domain stability. Our work represents the most complete assessment to date of the role of engineered noncanonical disulfide bonding within FRs in Ig V domain structure and function.

Overexpression of SlPRE3 alters the plant morphologies in Solanum lycopersicum.

KEY MESSAGE: Overexpression of SlPRE3 is detrimental to the photosynthesis and alters plant morphology and root development. SlPRE3 interacts with SlAIF1/SlAIF2/SlPAR1/SlIBH1 to regulate cell expansion. Basic helix-loop-helix (bHLH) transcription factors play crucial roles as regulators in plant growth and development. In this study, we isolated and characterized SlPRE3, an atypical bHLH transcription factor gene. SlPRE3 exhibited predominant expression in the root and moderate expression in the senescent leaves. Comparative analysis with the wild type revealed significant differences in plant morphology in the 35S:SlPRE3 lines. These differences included increased internode length, rolling leaves with reduced chlorophyll accumulation, and elongated yet fewer adventitious roots. Additionally, 35S:SlPRE3 lines displayed elevated levels of GA3 (gibberellin A3) and reduced starch accumulation. Furthermore, utilizing the Y2H (Yeast two-hybrid) and the BiFC (Bimolecular Fluorescent Complimentary) techniques, we identified physical interactions between SlPRE3 and SlAIF1 (ATBS1-interacting factor 1)/SlAIF2 (ATBS1-interacting factor 2)/SlPAR1 (PHYTOCHROME RAPIDLY REGULATED 1)/SlIBH1 (ILI1-binding bHLH 1). RNA-seq analysis of root tissues revealed significant alterations in transcript levels of genes involved in gibberellin metabolism and signal transduction, cell expansion, and root development. In summary, our study sheds light on the crucial regulatory role of SlPRE3 in determining plant morphology and root development.

Exosome-derived miR-372-5p promotes stemness and metastatic ability of CRC cells by inducing macrophage polarization.

Colorectal cancer (CRC) is the most common malignancy in the digestive system, and tumor metastasis is the main cause of death in clinical patients with CRC. It has been shown that exosomes promote phenotypic changes in macrophages and tumor metastasis in the CRC tumor microenvironment. In this study, we used miRNA-seq technology to screen out the highly expressed miR-372-5p among the miRNAs differentially expressed in plasma exosomes of clinical CRC patients. It was found that miR-372-5p highly expressed in HCT116 exosomes could be phagocytosed by macrophages and promote their polarization into M2 macrophages by regulating the PTEN/AKT pathway. Meanwhile, co-culture of CRC cells with conditioned medium (CM) of macrophages enhanced the EMT, stemness and metastasis of CRC cells. Mechanistically, CRC cells exosome-derived miR-372-5p induced polarized M2 macrophages to secrete chemokine C-X-C-Motif Ligand 12 (CXCL12), which activated the WNT/ß-catenin pathway to promote the EMT, stemness and metastatic ability of CRC cells. In summary, this study elucidated the molecular mechanism of exosomal miR-372-5p promoting metastasis and stemness in CRC, which may provide new therapeutic targets for CRC metastasis and prognosis assessment.

Colorectal cancer tumor cell-derived exosomal miR-203a-3p promotes CRC metastasis by targeting PTEN-induced macrophage polarization.

(1) Background: Tumor-associated macrophages (TAMs) are important immunocytes associated with liver metastasis of colorectal cancer (CRLM). However, the molecular processes underpinning the interaction between the TME and the tumour-derived exosomal miRNAs in CRLM are not being fully understood; (2) Methods: Transmission electron microscopy was utilized to confirm the existence of exosomes after differential ultracentrifugation. To determine the roles of exosomal miR-203a-3p, an in vivo and in vitro investigation was conducted. The mechanism by which exosomal miR-203a-3p governs the interaction between CRC cells and M2 macrophages was investigated using a dual-luciferase reporter assay, western blot, and other techniques; (3) Results: Overexpression of miR-203a-3p was associated with poor prognosis and liver metastasis in CRC patients. Exosomal miR-203a-3p was upregulated in the plasma of CRC patients and highly metastatic CRC cells HCT116, and it could be transported to macrophages via exosomes. Exosomal miR-203a-3p induced M2 polarization of macrophages by controlling PTEN and activating the PI3K/Akt signaling pathway. M2-polarized macrophages secreted the CXCL12, which increased cancer metastasis and resulted in pre-metastatic niches in CRLM by CXCL12/CXCR4/NF-κB signaling pathway. Co-culture of macrophages with miR-203a-3p-transfected or exosome-treated cells increased the ability of HCT116 cells to metastasize both in vitro and in vivo; (4) Conclusions: Exosomes produced by highly metastatic CRC cells and rich in miR-203a-3p may target PTEN and alter the TME, promoting liver metastasis in CRC patients. These findings offer fresh understanding of the liver metastatic process in CRC.

The Multimorbidity and Lifestyle Correlates in Chinese Population Residing in Macau: Findings from a Community-Based Needs Assessment Study.

Multimorbidity has become one of the most pressing public health concerns worldwide. The objectives of this study were to understand the prevalence of multimorbidity and its relationship with lifestyle factors among Chinese adults in Macau, and to investigate the combined contribution of common lifestyle factors in predicting multimorbidity. Data were collected through face-to-face interviews using a self-reported questionnaire on common chronic diseases, lifestyle factors and sociodemographics. BMI, physical activity, drinking status, smoking status and sleep quality were assessed, and a composite lifestyle score (0 to 9 points) was calculated, and the higher the score, the healthier the lifestyle. A total of 1443 participants were included in the analysis, of whom 55.2% were female, 51.8% were middle aged or elderly and 30.5% completed tertiary education or higher. The prevalence of multimorbidity was 10.3%. The combination of hypertension and hyperlipidaemia was the most common (22.2%) multimorbidity among the participants with multimorbidity. After the adjustment of the covariates, it was found that the participants who were overweight (OR: 1.95, 95% CI: 1.18-3.20, p = 0.009) or obese (OR: 3.76, 95% CI: 2.38-5.96, p < 0.001), former drinkers (OR: 2.43, 95% CI: 1.26-4.69, p = 0.008), and those who reported poor sleep quality (OR: 2.25, 95% CI: 1.49-3.40, p < 0.001) had a high risk of developing multimorbidity. A one-unit increase in the lifestyle score was associated with a 0.33-times reduction in the risk of developing multimorbidity (OR: 0.67; 95% CI: 0.59-0.77, p < 0.001). A combination of lifestyle factors can influence a variety of multimorbidity among the Chinese adults in Macau. Thus, comprehensively assessing the combined contribution of several lifestyle factors in predicting multimorbidity is important.