TAF5L and TAF6L Maintain Self-Renewal of Embryonic Stem Cells via the MYC Regulatory Network.

Self-renewal and pluripotency of the embryonic stem cell (ESC) state are established and maintained by multiple regulatory networks that comprise transcription factors and epigenetic regulators. While much has been learned regarding transcription factors, the function of epigenetic regulators in these networks is less well defined. We conducted a CRISPR-Cas9-mediated loss-of-function genetic screen that identified two epigenetic regulators, TAF5L and TAF6L, components or co-activators of the GNAT-HAT complexes for the mouse ESC (mESC) state. Detailed molecular studies demonstrate that TAF5L/TAF6L transcriptionally activate c-Myc and Oct4 and their corresponding MYC and CORE regulatory networks. Besides, TAF5L/TAF6L predominantly regulate their target genes through H3K9ac deposition and c-MYC recruitment that eventually activate the MYC regulatory network for self-renewal of mESCs. Thus, our findings uncover a role of TAF5L/TAF6L in directing the MYC regulatory network that orchestrates gene expression programs to control self-renewal for the maintenance of mESC state.

Discovery of Natural Products Alleviating Renal Fibrosis with a Viscosity-Responsive Molecular Probe.

Renal fibrosis poses a significant threat to individuals suffering from chronic progressive kidney disease. Given the absence of effective medications for treating renal fibrosis, it becomes crucial to assess the extent of fibrosis in real time and explore the development of novel drugs with substantial therapeutic benefits. Due to the accumulation of renal tissue damage and the uncontrolled deposition of fibrotic matrix during the course of the disease, there is an increase in viscosity both intracellularly and extracellularly. Therefore, a viscosity-sensitive near-infrared fluorescence (NIRF) and photoacoustic (PA) imaging probe, BDP-KY, was developed to detect aberrant changes in viscosity during fibrosis. Furthermore, BDP-KY has been applied to screen the effective components of herbal medicine, rhubarb, resulting in the identification of potential antirenal fibrotic compounds such as emodin-8-glucoside and chrysophanol 8-O-glucoside. Ultrasound, PA, and NIRF imaging of a unilateral uretera obstruction mice model show that different concentrations of emodin-8-glucoside and chrysophanol 8-O-glucoside effectively reduce viscosity levels during the renal fibrosis process. The histological results showed a significant decrease in fibrosis factors α-smooth muscle actin and collagen deposition. Combining these findings with their pharmacokinetic characteristics, these compounds have the potential to fill the current market gap for effective antirenal fibrosis drugs. This study demonstrates the potential of BDP-KY in the evaluation of renal fibrosis, and the two identified active components from rhubarb hold great promise for the treatment of renal fibrosis.

A novel method for evaluating pseudoallergy based on ß-hexosaminidase and its application for traditional Chinese medicine injections.

Pseudoallergy is a typical and common adverse drug reaction to injections, especially in traditional Chinese medicine injections (TCMIs). At present, the evaluation methods for pseudoallergy include cell methods in vitro and animal methods in vivo. The mast cell evaluation method based on the ß-hexosaminidase (ß-Hex)-catalyzed substrate, 4-nitrophenyl-ß-N-acetyl-D-glucosaminide (4-NPG), is an important method for the evaluation of drug-induced pseudoallergy, but it is prone to false positive results and has insufficient sensitivity. In this study, a novel ß-Hex evaluation system with rat basophilic leukemia-2H3 cells based on high-performance liquid chromatography-fluorescence detection (HPLC-FLD) was established, which effectively increased the sensitivity and avoided false positive results. Cell viabilities were measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay. In addition, a method for the determination of histamine, which is another indicator in the development of pseudoallergy, was established to validate the above method. The results of this novel method indicated that two TCMIs (Shuxuening injection and Shenqi Fuzheng injection), which were considered to be pseudoallergenic using 4-NPG, were not pseudoallergenic. Overall, the novel ß-Hex/HPLC-FLD evaluation system using Rat basophilic leukemia-2H3 cells established was effective and precise. It could be used for the evaluation of pseudoallergic reactions caused by TCMIs and other injections.

[Value of SOX1 and PAX1 Gene Methylation Detection in Secondary Triage of High-Grade Cervical Lesions].

Objective To evaluate the value of SOX1 and PAX1 gene methylation detection in the secondary triage of high-grade cervical lesions.Methods Exfoliated cervical cells were collected from 122 patients tested positive for human papilloma virus (HPV) and subjected to thin-prep cytologic test (TCT) and SOX1/PAX1 gene methylation tests.Results The HPV test combined with TCT showed the sensitivity of 95.24% and the specificity of 23.75% for detecting cervical intraepithelial neoplasia (CIN) grade 2 and above (CIN2+).After the addition of the SOX1/PAX1 gene methylation detection in secondary triage,the sensitivity for detecting CIN2+ was 83.33%,which had no statistically significant difference from the sensitivity of TCT combined with HPV test (P=0.078).However,the specificity reached 77.50%,which was significantly higher than that of HPV test combined with TCT (P<0.001).The SOX1/PAX1 gene methylation level in the CIN2+ group was higher than those in the normal cervical tissue and the CIN1 group(P<0.001).The cut-off values of SOX1 and PAX1 gene methylation for CIN2+ detection were -11.81 and -11.98,respectively.Conclusion Adding the detection of SOX1/PAX1 gene methylation in secondary triage significantly improves the efficiency and accuracy of CIN2+ detection.

Small-Molecule Photoacoustic Imaging Probe with Aggregation-Enhanced Amplitude for Real-Time Visualization of Acute Kidney Injury.

Acute kidney injury (AKI) has become a growing issue for patients with the extensive use of all kinds of drugs in clinic. Photoacoustic (PA) imaging provides a noninvasive and real-time imaging method for studying kidney injury, but it has inherent shortages in terms of high background signal and low detection sensitivity for exogenous imaging agents. Intriguingly, J-aggregation offers to tune the optical properties of the dyes, thus providing a platform for developing new PA probes with desired performance. In this study, a small-molecule PA probe (BDP-3) was designed and synthesized. We serendipitously discovered that BDP-3 can transform into renal clearable nanoaggregates under physiological conditions. The hydrodynamic diameter of the BDP-3 increased from 0.64 ± 0.11 to 3.74 ± 0.39 nm when the content of H2O increased from 40 to 90%. In addition, it was surprising that such a transforming process can significantly enhance its PA amplitude (2.06-fold). On this basis, PA imaging with BDP-3 was applied as a new method for the noninvasive detection of AKI induced by anticancer drugs, traditional Chinese medicine, and clinical contrast agents in animal models and exhibited higher sensitivity than the conventional serum index test, demonstrating great potential for further clinical diagnostic applications.

Smad4 promotes diabetic nephropathy by modulating glycolysis and OXPHOS.

Diabetic nephropathy (DN) is the leading cause of end-stage kidney disease. TGF-ß1/Smad3 signalling plays a major pathological role in DN; however, the contribution of Smad4 has not been examined. Smad4 depletion in the kidney using anti-Smad4 locked nucleic acid halted progressive podocyte damage and glomerulosclerosis in mouse type 2 DN, suggesting a pathogenic role of Smad4 in podocytes. Smad4 is upregulated in human and mouse podocytes during DN. Conditional Smad4 deletion in podocytes protects mice from type 2 DN, independent of obesity. Mechanistically, hyperglycaemia induces Smad4 localization to mitochondria in podocytes, resulting in reduced glycolysis and oxidative phosphorylation and increased production of reactive oxygen species. This operates, in part, via direct binding of Smad4 to the glycolytic enzyme PKM2 and reducing the active tetrameric form of PKM2. In addition, Smad4 interacts with ATPIF1, causing a reduction in ATPIF1 degradation. In conclusion, we have discovered a mitochondrial mechanism by which Smad4 causes diabetic podocyte injury.

Aminoacylase-1 plays a key role in myocardial fibrosis and the therapeutic effects of 20(S)-ginsenoside Rg3 in mouse heart failure.

We previously found that the levels of metabolite N-acetylglutamine were significantly increased in urine samples of patients with heart failure (HF) and in coronary artery ligation (CAL)-induced HF mice, whereas the expression of its specific metabolic-degrading enzyme aminoacylase-1 (ACY1) was markedly decreased. In the current study, we investigated the role of ACY1 in the pathogenesis of HF and the therapeutic effects of 20(S)-ginsenoside Rg3 in HF experimental models in vivo and in vitro. HF was induced in mice by CAL. The mice were administered Rg3 (7.5, 15, 30 mg · kg-1· d-1, i.g.), or positive drug metoprolol (Met, 5.14 mg · kg-1· d-1, i.g.), or ACY1 inhibitor mono-tert-butyl malonate (MTBM, 5 mg · kg-1 · d-1, i.p.) for 14 days. We showed that administration of MTBM significantly exacerbated CAL-induced myocardial injury, aggravated cardiac dysfunction, and pathological damages, and promoted myocardial fibrosis in CAL mice. In Ang II-induced mouse cardiac fibroblasts (MCFs) model, overexpression of ACY1 suppressed the expression of COL3A1 and COL1A via inhibiting TGF-ß1/Smad3 pathway, whereas ACY1-siRNA promoted the cardiac fibrosis responses. We showed that a high dose of Rg3 (30 mg · kg-1· d-1) significantly decreased the content of N-acetylglutamine, increased the expression of ACY1, and inhibited TGF-ß1/Smad3 pathway in CAL mice; Rg3 (25 µM) exerted similar effects in Ang II-treated MCFs. Meanwhile, Rg3 treatment ameliorated cardiac function and pathological features, and it also attenuated myocardial fibrosis in vivo and in vitro. In Ang II-treated MCFs, the effects of Rg3 on collagen deposition and TGF-ß1/Smad3 pathway were slightly enhanced by overexpression of ACY1, whereas ACY1 siRNA partially weakened the beneficial effects of Rg3, suggesting that Rg3 might suppress myocardial fibrosis through ACY1. Our study demonstrates that N-acetylglutamine may be a potential biomarker of HF and its specific metabolic-degrading enzyme ACY1 could be a potential therapeutic target for the prevention and treatment of myocardial fibrosis during the development of HF. Rg3 attenuates myocardial fibrosis to ameliorate HF through increasing ACY1 expression and inhibiting TGF-ß1/Smad3 pathway, which provides some references for further development of anti-fibrotic drugs for HF.

Ruscogenin attenuates particulate matter-induced acute lung injury in mice via protecting pulmonary endothelial barrier and inhibiting TLR4 signaling pathway.

The inhalation of particulate matter (PM) is closely related to respiratory damage, including acute lung injury (ALI), characterized by inflammatory fluid edema and disturbed alveolar-capillary permeability. Ruscogenin (RUS), the main active ingredient in the traditional Chinese medicine Ophiopogonis japonicus, has been found to exhibit anti-inflammatory activity and rescue LPS-induced ALI. In this study, we investigated whether and how RUS exerted therapeutic effects on PM-induced ALI. RUS (0.1, 0.3, 1 mg·kg-1·d-1) was orally administered to mice prior to or after intratracheal instillation of PM suspension (50 mg/kg). We showed that RUS administration either prior to or after PM challenge significantly attenuated PM-induced pathological injury, lung edema, vascular leakage and VE-cadherin expression in lung tissue. RUS administration significantly decreased the levels of cytokines IL-6 and IL-1ß, as well as the levels of NO and MPO in both bronchoalveolar lavage fluid (BALF) and serum. RUS administration dose-dependently suppressed the phosphorylation of NF-κB p65 and the expression of TLR4 and MyD88 in lung tissue. Furthermore, TLR4 knockout partly diminished PM-induced lung injury, and abolished the protective effects of RUS in PM-instilled mice. In conclusion, RUS effectively alleviates PM-induced ALI probably by inhibition of vascular leakage and TLR4/MyD88 signaling. TLR4 might be crucial for PM to initiate pulmonary lesion and for RUS to exert efficacy against PM-induced lung injury.

HPLC-QTOF/MS-based metabolomics to explore the molecular mechanisms of Yiqi Fumai Lyophilized Injection in heart failure mice.

Chronic heart failure is a common and fatal disease triggered by loss of normal cardiac function. Yiqi Fumai Lyophilized Injection is widely used in the treatment of cardiovascular diseases, especially chronic heart failure. In this study, a model of chronic heart failure in mice was established with permanent coronary artery ligation followed by Yiqi Fumai Lyophilized Injection intervention for 14 days. Then, the endogenous metabolites of mice plasma and urine samples were screened through nontargeted metabolomics techniques. The results indicated that Yiqi Fumai Lyophilized Injection treatment changed the metabolic pattern of chronic heart failure and regulated valine, leucine, and isoleucine biosynthesis, taurine and hypotaurine metabolism, histidine metabolism and arginine biosynthesis, etc. Finally, the cardioprotective mechanism of Yiqi Fumai Lyophilized Injection was further verified in the mouse model of chronic heart failure and angiotensin II-induced cardiac fibroblasts based on metabolomics. The results showed that Yiqi Fumai Lyophilized Injection could inhibit myocardial fibrosis to improve chronic heart failure. This study firstly elucidated the metabolic network and pathways regulated by Yiqi Fumai Lyophilized Injection, which might facilitate the realization of the clinically accurate application of Yiqi Fumai Lyophilized Injection in the treatment of chronic heart failure.

Quality assessment and traceability study of Angelicae Sinensis Radix via binary chromatography, triple quadrupole tandem mass spectrometry, and multivariate statistical analysis.

Angelicae Sinensis Radix is a world-renowned herbal medicine originating in China. Owing to many environmental and geographical factors, Angelicae Sinensis Radix from various origins may have a difference in the content of ingredients, which made the confusion in the clinical practice and market. Herein, a binary chromatographic fingerprinting analysis method is developed via hydrophilic interaction chromatography and reversed-phase liquid chromatography to obtain more chemical information. Following that, an ultra-performance liquid chromatography with a triple quadrupole mass spectrometry method is furnished to simultaneously detect 17 ingredients of Angelicae Sinensis Radix gathered from six geographic zones in China. Eventually, the principal component analysis is successfully carried out to classify and differentiate the Angelicae Sinensis Radix from different origins, meanwhile the quantitative volcano plots was used to observe the changes of ingredient trends vividly. Accordingly, the proposed binary chromatography and triple quadrupole tandem mass spectrometry coupled with multivariate statistical analysis can be utilized as a facile and reliable method for origin tracing and quality control of Angelicae Sinensis Radix.

[Analysis on ingredients and metabolites of Gegen Decoction absorbed into blood based on UHPLC-Q-TOF-MS].

This study analyzed the plasma components of Gegen Decoction(GGD) by ultra-high-performance liquid chromatography-quadrupole-time of flight mass spectrometry(UHPLC-Q-TOF-MS), which is expected to serve as a reference for exploring the pharmacodynamic substances of GGD. Female Wistar rats were given(ig) GGD and then plasma samples were collected and analyzed by UHPLC-Q-TOF-MS. The results showed that 42 chemical components were identified: 25 prototypes(14 from Puerariae Lobatae Radix, 6 from Glycyrrhizae Radix et Rhizoma, 3 from Paeoniae Radix Alba, and 2 from Ephedrae Herba) and 17 metabolites(from isoflavonoids in Puerariae Lobatae Radix and Glycyrrhizae Radix et Rhizoma). UHPLC-Q-TOF-MS was employed to achieve rapid analysis of plasma components of GGD, laying a basis for elucidating the therapeutic material basis and mechanism of GGD.

[Role of Puerariae Lobatae Radix in Gegen Decoction for treatment of primary dysmenorrhea].

This study aimed to explore the characteristic role of Puerariae Lobatae Radix(PLR) in Gegen Decoction for the treatment of primary dysmenorrhea(PD). Estrogen(E_2) was combined with oxytocin to establish a mouse model of PD. The mice were randomly divided into a normal group, a model group, a Gegen Decoction group, a PLR-free Gegen Decoction group, a PLR group, and a positive drug group(ibuprofen). Writhing response times and writhing incubation of mice in each group were tested by behavio-ral assessment, and the serum levels of prostaglandin F_(2α)(PGF_(2α)), prostaglandin E_2(PGE_2), E_2, and progesterone(PROG) were detected by ELISA kits. Western blot method was adopted to detect cyclooxygenase-2(COX-2) and estrogen receptor alpha(ER_α) expression levels in uterine tissues. Doppler ultrasound was employed to detect changes in uterine artery blood flow in mice, including peak systolic blood flow velocity(maximum velocity), end-diastolic velocity(minimum velocity), peak systolic blood flow velocity/end-diastolic velocity(S/D), pulsatility index(PI), and resistive index(RI). Histopathological changes in the uterus were detected by HE staining. Based on the oxytocin-induced isolated uterine contraction model, the effects of Gegen Decoction, PLR-free Gegen Decoction, and PLR on the amplitude, frequency, and activity of isolated uterine contraction were compared to investigate the role of PLR in Gegen Decoction for the treatment of PD. The results showed that compared with the Gegen Decoction group, the PLR-free Gegen Decoction improved the indicators of PD except for E_2 content, ER_α expression, and uterine artery blood flow. PLR could significantly down-regulate the serum content of E_2 and the protein expression of uterine ER_α, and improve the uterine artery blood flow. The data suggested that PLR, as the sovereign drug of Gegen Decoction, might function in Gegen Decoction for the treatment of PD by mediating E_(2 )and improving the uterine artery blood flow.

Rapid and sensitive detection of NGAL for the prediction of acute kidney injury via a polydopamine nanosphere/aptamer nanocomplex coupled with DNase I-assisted recycling amplification.

Early detection of acute kidney injury (AKI) is important, as early intervention and treatment can prevent further kidney injury and improve kidney health. Neutrophil gelatinase-associated lipocalin (NGAL) has emerged as the earliest and promising non-invasive biomarker of AKI in urine, and has been used as a new predictive biomarker of AKI in the bench-to-bedside journey. In this work, a nanocomplex composed of a polydopamine nanosphere (PDANS) and a fluorophore-labelled aptamer has been constructed for the detection of NGAL using a DNase I-assisted recycling amplification strategy. After the addition of NGAL, the fluorescence intensity increases linearly over the NGAL concentration range from 12.5 to 400 pg mL-1. The limit of detection of this strategy is found to be 6.25 pg mL-1, which is almost 5 times lower than that of the method that does not involve DNase I. The process can be completed within 1 h, indicating a fast fluorescence response. Furthermore, the method using the nanocomplex coupled with DNase I has been successfully utilized for the detection of NGAL in the urine from cisplatin-induced AKI and five-sixths nephrectomized mice, demonstrating its promising ability for the early prediction of AKI. This method also demonstrates the protective effect of the Huangkui capsule on AKI, and provides an effective way to screen potentially protective drugs for renal disease.

Exploring the protective effects of schizandrol A in acute myocardial ischemia mice by comprehensive metabolomics profiling integrated with molecular mechanism studies.

Schizandrol A (SA) is an bioactive component isolated from the Schisandra chinensis (Turcz.) Baill., which has been used as a remedy to prevent oxidative injury. However, whether the cardioprotective effect of SA is associated with regulating endogenous metabolites remains unclear, thus we performed comprehensive metabolomics profiling in acute myocardial ischemia (AMI) mice following SA treatment. AMI was induced in ICR mice by coronary artery ligation, then SA (6 mg·kg-1·d-1, ip) was administered. SA treatment significantly decreased the infarct size, preserved the cardiac function, and improved the biochemical indicators and cardiac pathological alterations. Moreover, SA (10, 100 M) significantly decreased the apoptotic index in OGD-treated H8c2 cardiomycytes in vitro. By using HPLC-Q-TOF/MS, we conducted metabonomics analysis to screen the significantly changed endogenous metabolites and construct the network in both serum and urine. The results revealed that SA regulated the pathways of glycine, serine and threonine metabolism, lysine biosynthesis, pyrimidine metabolism, arginine and proline metabolism, cysteine and methionine metabolism, valine, leucine and isoleucine biosynthesis under the pathological conditions of AMI. Furthermore, we selected the regulatory enzymes related to heart disease, including ecto-5'-nucleotidase (NT5E), guanidinoacetate N-methyltransferase (GAMT), platelet-derived endothelial cell growth factor (PD-ECGF) and methionine synthase (MTR), for validation. In addition, SA was found to facilitate PI3K/Akt activation and inhibit the expression of NOX2 in AMI mice and OGD-treated H9c2 cells. In conclusion, we have elucidated SA-regulated endogenous metabolic pathways and constructed a regulatory metabolic network map. Furthermore, we have validated the new potential therapeutic targets and underlying molecular mechanisms of SA against AMI, which might provide a reference for its future application in cardiovascular diseases.

[Analysis on new research and development ideas and technical points of traditional Chinese medicine for prevention and treatment of chronic heart failure].

Chronic heart failure(CHF), a serious and end stage of various heart diseases, is a common chronic cardiovascular disease in the 21 st century. Literature data show that the 5-year mortality rate of hospitalized patients with heart failure is as high as 50%. Nowadays, the development of drugs treating heart failure has become a hot spot, meanwhile, traditional Chinese medicine(TCM) has shown the advantages in the treatment of chronic heart failure. In this article, four stages to develop traditional Chinese medicine for chronic heart failure were proposed. Firstly, discuss and screen ideas and methods with regard to the development of TCM and its prescriptions based on clinical needs. Secondly, study the preparation process and quality control method by referring to the existing clinical background of TCM prescriptions and analyzing the chemical compositions and pharmacological action characteristics of each herb in the prescription. Then, design non-clinical evaluation programs and carry out researches on pharmacodynamics and toxicology by combining the experience of clinical use of TCM prescriptions and future clinical positioning, and gradually adjust and improve the programs during implementation. Finally, conduct clinical trial application(IND) by submitting registration application data which are base on the clinical drug experience, preclinical research pharmacy, main pharmacodynamics, safety test results of the prescription, clinical positioning, and reasonable clinical trial plan designed by the theory of TCM. After passing the IND technical review, the clinical trial study shall be officially launched to achieve the desired results and obtain effective Chinese patent medicines for heart failure treatment.

Synergistic combination of DT-13 and Topotecan inhibits aerobic glycolysis in human gastric carcinoma BGC-823 cells via NM IIA/EGFR/HK II axis.

DT-13 combined with topotecan (TPT) showed stronger antitumour effects in mice subcutaneous xenograft model compared with their individual effects in our previous research. Here, we further observed the synergistically effect in mice orthotopic xenograft model. Metabolomics analysis showed DT-13 combined with TPT alleviated metabolic disorders induced by tumour and synergistically inhibited the activity of the aerobic glycolysis-related enzymes in vivo and in vitro. Mechanistic studies revealed that the combination treatment promoted epidermal growth factor receptor (EGFR) degradation through non-muscle myosin IIA (NM IIA)-induced endocytosis of EGFR, further inhibited the activity of hexokinase II (HK II), and eventually promoted the aerobic glycolysis inhibition activity more efficiently compared with TPT or DT-13 monotherapy. The combination therapy also inhibited the specific binding of HK II to mitochondria. When using the NM II inhibitor (-)002Dblebbistatin or MYH-9 shRNA, the synergistic inhibition effect of DT-13 and TPT on aerobic glycolysis was eliminated in BGC-823 cells. Immunohistochemical analysis revealed selective up-regulation of NM IIA while specific down-regulation of p-CREB, EGFR, and HK II by the combination therapy. Collectively, these findings suggested that this regimen has significant clinical implications, warranted further investigation.

Targeted Myocardial Hypoxia Imaging Using a Nitroreductase-Activatable Near-Infrared Fluorescent Nanoprobe.

Development of a highly selective and sensitive imaging probe for accurate detection of myocardial hypoxia will be helpful to estimate the degree of ischemia and subsequently guide personalized treatment. However, an efficient optical approach for hypoxia monitoring in myocardial ischemia is still lacking. In this work, a cardiomyocyte-specific and nitroreductase-activatable near-infrared nanoprobe has been developed for selective and sensitive imaging of myocardial hypoxia. The nanoprobe is a liposome-based nanoarchitecture which is functionalized with a peptide (GGGGDRVYIHPF) for targeting heart cells and encapsulating a nitrobenzene-substituted BODIPY for nitroreductase imaging. The nanoprobe can specifically recognize and bind to angiotensin II type 1 receptor that is overexpressed on the ischemic heart cells by the peptide and is subsequently uptaken into heart cells, in which the probe is released and activated by hypoxia-related nitroreductase to produce fluorescence emission at 713 nm. The in vitro response of the nanoprobe toward nitroreductase resulted in 55-fold fluorescence enhancement with the limit of detection as low as 7.08 ng/mL. Confocal fluorescence imaging confirmed the successful uptake of nanoprobe by hypoxic heart cells and intracellular detection of nitroreductase. More significantly, in vivo imaging of hypoxia in a murine model of myocardial ischemia was achieved by the nanoprobe with high sensitivity and good biocompatibility. Therefore, this work presents a new tool for targeted detection of myocardial hypoxia and will promote the investigation of the hypoxia-related physiological and pathological process of ischemic heart disease.

Plasminogen activator, tissue type regulates germinal vesicle breakdown and cumulus expansion of bovine cumulus-oocyte complex in vitro†.

Plasminogen activator, tissue type (PLAT) and its inhibitor serpin family E member 1 (SERPINE1) cooperatively regulate PLAT activity in various reproductive processes. However, it is unknown whether this includes bovine oocyte maturation. We addressed this question in the present study by evaluating PLAT and SERPINE1 protein localization in immature cumulus-oocyte complexes (COCs), as well as PLAT mRNA and protein expression in cultured COCs after 0, 8, 16, and 24 h of in vitro maturation (IVM). We also examined the effects of PLAT and SERPINE1 on germinal vesicle breakdown (GVBD) and oocyte cyclic 3' 5' adenosine monophosphate (cAMP) levels, cumulus expansion index, and expansion-related gene expression in oocytes derived from bovine COCs cultured for 4, 8, and 12 h and in COCs cultured for 16 h. Both PLAT and SERPINE1 localized in cumulus cells but only the latter was detected in oocytes. PLAT and SERPINE1 transcript levels increased during IVM; however, from 8 to 16 h, the levels of PLAT remained stable whereas those of SERPINE1 increased, resulting in a decline in PLAT concentration. Additionally, PLAT delayed GVBD, increased oocyte cAMP levels, and blocked cumulus expansion and associated gene expression, which was reversed by SERPINE1 supplemented. Thus, PLAT delays bovine oocyte GVBD by enhancing oocyte cAMP levels during the first 8 h of IVM; suppression of PLAT activity via accumulation of SERPINE1 in COCs results in cumulus expansion from 8 to 16 h of IVM. These findings provide novel insights into the molecular mechanisms underlying in vitro bovine oocyte maturation.

A hairpin DNA-fueled nanoflare for simultaneous illumination of two microRNAs in drug-induced nephrotoxic cells with target catalytic recycling amplification.

The detection of specific extracellular microRNAs (miRNAs) is beneficial for the prediction of drug-induced kidney injury. Here, a novel hairpin DNA-fueled nanoflare was developed for the simultaneous detection of drug-induced nephrotoxicity-related miRNA-21 and miRNA-200c with target catalytic recycling amplification. The nanoflare utilized gold nanoparticles (AuNPs) as the highly efficient quencher to ensure a low background signal. With the help of the fueled hairpin DNA, the miRNA targets could serve as the catalysts for the assembly of DNA duplexes. Therefore, the nanoflare could respond to the miRNAs to yield signal outputs of 1 : n (target : signal) rather than an equivalent reaction ratio of 1 : 1, achieving the signal amplified detection of low-abundant miRNAs. The targets can be concurrently detected with the detection limit of 18.1 and 21.1 pM for miRNA-21 and miRNA-200c, respectively, which are approximately 2 orders of magnitude lower than that of the non-catalytic probes. In addition, this nanoflare offered a high selectivity for determination between perfectly matched targets and single-base mismatched targets. It should be noted that the nanoflare was successfully employed to predict the drug-induced nephrotoxicity by the detection of miRNAs in culture media excreted from the drug-treated renal cells using a fluorescent microplate reader. Our hairpin DNA-fueled nanoflare could also accurately detect the divergence of miRNA-21 and miRNA-200c between drug-treated nephrotoxic cells and tumor cells, demonstrating a promising potential for exploring the pathogenesis of drugs and auxiliary diagnosis of drug-induced nephrotoxicity.

Prediction of Drug-Induced Nephrotoxicity with a Hydroxyl Radical and Caspase Light-Up Dual-Signal Nanoprobe.

The development of well-designed nanoprobes for specific imaging of multiple biomarkers in renal cells will afford beneficial information related to the transmutation process of drug-induced kidney injury (DIKI). However, the most reported nanoprobes for DIKI detection were dependent on single-signal output and lack of kidney targeting. In this work, we reported a renal cell targeting and dual-signal nanoprobe by encapsulating Brite 670 and Dabcyl-KFFFDEVDK-FAM into a low molecular weight chitosan nanoparticle. Confocal fluorescence imaging results demonstrated that the nanoprobe could visualize the upregulation of hydroxyl radical in early stage and activation of caspase-3 in late stage of DIKI at both the renal cell and tissue level. In a mouse DIKI model, the positive time of 8 h using nanoprobe imaging was superior to that of 72 h for serum creatinine or blood urea nitrogen, 16 h for cystatin-C, and 24 h for kidney injury molecule-1 with conventional methods. These results demonstrated that the nanoprobe may be a promising tool for effective early prediction and discriminative imaging of DIKI.