Indole-3-carbinol ameliorates ovarian damage in female old mice through Nrf2/HO-1 pathway activation.

Ovarian aging leads to infertility and birth defects. We aimed to clarify the role of Indole-3-carbinol (I3C) in resistance to oxidative stress, apoptosis, and fibrosis in ovarian aging. I3C was administered via intraperitoneal injection for 3 weeks in young or old mice. Immunohistochemistry; Masson, Sirius red, and TUNEL staining; follicle counting; estrous cycle analysis; and Western blotting were used for validating the protective effect of I3C against ovarian senescence. Human granulosa-like tumor cell line and primary granulosa cells were used for in vitro assay. The results indicated that I3C inhibited ovarian fibrosis and apoptosis while increasing the number of primordial follicles. Mechanistic studies have shown that I3C promoted the nuclear translocation of nuclear factor-erythroid 2-related factor (Nrf2) and upregulated the expression of heme oxygenase 1 (HO-1). Additionally, I3C increased cell viability and decreased lactate dehydrogenase, malondialdehyde, reactive oxygen species and JC-1 levels. Furthermore, the antioxidant effect of I3C was found to be dependent on the activation of Nrf2 and HO-1, as demonstrated by the disappearance of the effect upon inhibition of Nrf2 expression. In conclusion, I3C can alleviate the ovarian damage caused by aging and may be a protective agent to delay ovarian aging.

The value of low-intensity pulsed ultrasound in reducing ovarian injury caused by chemotherapy in mice.

BACKGROUND: Ovarian damage and follicle loss are major side effects of chemotherapy in young female patients with cancer. However, effective strategies to prevent these injuries are still lacking. The purpose of this study was to verify low-intensity pulsed ultrasound (LIPUS) can reduce ovarian injury caused by chemotherapy and to explore its underlying mechanisms in mice model. METHODS: The mice were randomly divided into the Control group, Cisplatin group, and Cisplatin + LIPUS group. The Cisplatin group and Cisplatin + LIPUS group were intraperitoneally injected with cisplatin every other day for a total of 10 injections, and the Control group was injected with saline. On the second day of each injection, the Cisplatin + LIPUS group received irradiation, whereas the other two groups received sham irradiation. We used a variety of biotechnologies to detect the differences in follicle count, granulosa cell apoptosis, fibrosis, transcriptome level, oxidative damage, and inflammation in differently treated mice. RESULT: LIPUS was able to reduce primordial follicle pool depletion induced by cisplatin and inhibit the apoptosis of granulosa cells. Transcriptomic results confirmed that LIPUS can reduce ovarian tissue injury. We demonstrated that LIPUS can relieve ovarian fibrosis by inhibiting TGF-ß1/Smads pathway. Meanwhile, it can reduce the oxidative damage and reduced the mRNA levels of proinflammatory cytokines caused by chemotherapy. CONCLUSION: LIPUS can reduce the toxic effects of chemotherapy drugs on ovaries, inhibit ovarian fibrosis, reduce the inflammatory response, and redcue the oxidative damage, reduce follicle depletion and to maintain the number of follicle pools.

Recent advances in super-resolution optical imaging based on aggregation-induced emission.

Super-resolution imaging has rapidly emerged as an optical microscopy technique, offering advantages of high optical resolution over the past two decades; achieving improved imaging resolution requires significant efforts in developing super-resolution imaging agents characterized by high brightness, high contrast and high sensitivity to fluorescence switching. Apart from technical requirements in optical systems and algorithms, super-resolution imaging relies on fluorescent dyes with special photophysical or photochemical properties. The concept of aggregation-induced emission (AIE) was proposed in 2001, coinciding with unprecedented advancements and innovations in super-resolution imaging technology. AIE probes offer many advantages, including high brightness in the aggregated state, low background signal, a larger Stokes shift, ultra-high photostability, and excellent biocompatibility, making them highly promising for applications in super-resolution imaging. In this review, we summarize the progress in implementation methods and provide insights into the mechanism of AIE-based super-resolution imaging, including fluorescence switching resulting from photochemically-converted aggregation-induced emission, electrostatically controlled aggregation-induced emission and specific binding-regulated aggregation-induced emission. Particularly, the aggregation-induced emission principle has been proposed to achieve spontaneous fluorescence switching, expanding the selection and application scenarios of super-resolution imaging probes. By combining the aggregation-induced emission principle and specific molecular design, we offer some comprehensive insights to facilitate the applications of AIEgens (AIE-active molecules) in super-resolution imaging.

Inhibition of cisplatin-induced Acsl4-mediated ferroptosis alleviated ovarian injury.

Given that the severity of the chemotherapy-induced ovarian damage, effective fertility preservation is a necessary part of the treatment process. Ferroptosis is a regulated cell death triggered by excessive phospholipid peroxidation caused by iron and the role of ferroptosis in chemotherapy-induced ovarian damage remains unclear. In this study, we demonstrated that cisplatin treatment caused the accumulation of iron ions which induced ferroptosis in ovarian tissue. And our results show that ferrostatin-1 was able to suppress the ovarian injury and granulosa cell death caused by cisplatin (Cis) in vivo and in vitro. At the same time, we observed significant changes in the expression levels of Acyl-CoA synthetase long-chain family member 4 (Acsl4) and glutathione peroxidase 4 (GPX4). Similarly, Rosiglitazone, an inhibitor of Acsl4, administration alleviated the ovary damage of the mice undergoing chemotherapy. Further mechanistic investigation showed that cisplatin increased the expression level of specificity protein 1 (SP1), and SP1 could bind to the promoter of Acsl4 to increased Acsl4 transcription. Overall, ferroptosis plays an important role in Cis induced ovarian injury, and inhibition of ferroptosis protects ovarian tissues from damage caused by cisplatin, and for the first time, we have identified the potential of Fer-1 and Rosi to protect ovarian function in female mice undergoing chemotherapy.

Automatic bifurcation detection utilizing pullback characteristics of bifurcation in intravascular optical coherence tomography.

Bifurcation detection in coronary arteries is significant since it influences the treatment strategy selection and optimization. Bifurcations are also reliable landmarks for image registration. Intravascular optical coherence tomography (IVOCT) is a high-resolution imaging modality that is very useful in percutaneous coronary intervention stenting optimization. We present a bifurcation identification method utilizing pullback characteristics for IVOCT, which can effectively identify the bifurcations with a small size. The longitudinal view of the pullback will appear as an outward discontinuity in the bifurcation area. By detecting this discontinuity, bifurcation can be identified with high accuracy. We also use the normal vectors method to extract the ostium of bifurcation. We compare the proposed method with the widely-used distance transformation method by clinical 5302 IVOCT images from 22 pullbacks. The average metrics of true positive rate (TPR), true negative rate (TNR), positive predictive value (PPV), and negative predictive value (NPV) for the proposed method are 86.97%, 98.50%, 85.56%, and 98.67%, respectively. TPR, PPV, and NPV by the proposed method are improved by 40.24%, 9.31%, 3.90%, and TNR is on par compared with the distance transformation method. Especially in the small bifurcation identification, TPR of the proposed method is 64.71% higher than the distance transformation method with a bifurcation area ratio less than 0.2.

Analysis and reduction of noise-induced depolarization in catheter based polarization sensitive optical coherence tomography.

In catheter based polarization sensitive optical coherence tomography (PS-OCT), a optical fiber with a rapid rotation in the catheter can cause low signal-to-noise ratio (SNR), polarization state instability, phase change of PS-OCT signals and then heavy noise-induced depolarization, which has a strong impact on the phase retardation measurement of the sample. In this paper, we analyze the noise-induced depolarization and find that the effect of depolarization can be reduced by polar decomposition after incoherent averaging in the Mueller matrix averaging (MMA) method. Namely, MMA can reduce impact of noise on phase retardation mapping. We present a Monte Carlo method based on PS-OCT to numerically describe noise-induced depolarization effect and contrast phase retardation imaging results by MMA and Jones matrix averaging (JMA) methods. The peak signal to noise ratio (PSNR) of simulated images processed by MMA is higher than about 8.9 dB than that processed by JMA. We also implement experiments of multiple biological tissues using the catheter based PS-OCT system. From the simulation and experimental results, we find the polarization contrasts processed by the MMA are better than those by JMA, especially at areas with high depolarization, because the MMA can reduce effect of noise-induced depolarization on the phase retardation measurement.

Automatic lumen segmentation using uniqueness of vascular connected region for intravascular optical coherence tomography.

We present an automatic lumen segmentation method using uniqueness of connected region for intravascular optical coherence tomography (IVOCT), which can effectively remove the effect on lumen segmentation caused by blood artifacts. Utilizing the uniqueness of vascular wall on A-lines, we detect the A-lines shared by multiple connected regions, identify connected regions generated by blood artifacts using traversal comparison of connected regions' location, shared ratio and area ratio and then remove all artifacts. We compare these three methods by 216 challenging images with severe blood artifacts selected from clinical 1076 IVOCT images. The metrics of the proposed method are evaluated including Dice index, Jaccard index and accuracy of 94.57%, 90.12%, 98.02%. Compared with automatic lumen segmentation based on the previous morphological feature method and widely used dynamic programming method, the metrics of the proposed method are significantly enhanced, especially in challenging images with severe blood artifacts.

Inhibiting bridge integrator 2 phosphorylation leads to improved oocyte quality, ovarian health and fertility in aging and after chemotherapy in mice.

Female ovaries degenerate about 20 years earlier than testes leading to reduced primordial follicle reserve and a reduction in oocyte quality. Here we found that bridge integrator 2 (BIN2) is enriched in mouse ovaries and oocytes and that global knockout of this protein improves both female fertility and oocyte quality. Quantitative ovarian proteomics and phosphoproteomics showed that Bin2 knockout led to a decrease in phosphorylated ribosomal protein S6 (p-RPS6), a component of the mammalian target of rapamycin pathway and greatly increased nicotinamide nucleotide transhydrogenase (NNT), the free-radical detoxifier. Mechanistically, we find that phosphorylation of BIN2 at Thr423 and Ser424 leads to its translocation from the membrane to the cytoplasm, subsequent phosphorylation of RPS6 and inhibition of Nnt translation. We synthesized a BIN2-penetrating peptide (BPP) designed to inhibit BIN2 phosphorylation and found that a 3-week BPP treatment improved primordial follicle reserve and oocyte quality in aging and after chemotherapy-induced premature ovarian failure without discernible side effects.

Three-dimensional spatial reconstruction of coronary arteries based on fusion of intravascular optical coherence tomography and coronary angiography.

We present a three-dimensional (3D) spatial reconstruction of coronary arteries based on fusion of intravascular optical coherence tomography (IVOCT) and digital subtraction angiography (DSA). Centerline of vessel in DSA images is exacted by multi-scale filtering, adaptive segmentation, morphology thinning and Dijkstra's shortest path algorithm. We apply the cross-correction between lumen shapes of IVOCT and DSA images and match their stenosis positions to realize co-registration. By matching the location and tangent direction of the vessel centerline of DSA images and segmented lumen coordinates of IVOCT along pullback path, 3D spatial models of vessel lumen are reconstructed. Using 1121 distinct positions selected from eight vessels, the correlation coefficient between 3D IVOCT model and DSA image in measuring lumen radius is 0.94% and 97.7% of the positions fall within the limit of agreement by Bland-Altman analysis, which means that the 3D spatial reconstruction IVOCT models and DSA images have high matching level.

The 5.8S pre-rRNA maturation factor, M-phase phosphoprotein 6, is a female fertility factor required for oocyte quality and meiosis.

OBJECTIVES: M-phase phosphoprotein 6 (MPP6) is important for 5.8S pre-rRNA maturation in somatic cells and was screened as a female fertility factor. However, whether MPP6 functions in oocyte meiosis and fertility is not yet known. We aimed to address this. MATERIALS AND METHODS: Mouse oocytes with surrounded nucleus (SN) or non-surrounded nucleus (NSN) were used for all experiments. Peptide nanoparticle-mediated antibody transfection was used to deplete MPP6. Immunofluorescence staining, immunohistochemistry and live tracker staining were used to examine MPP6 localization and characterize phenotypes after control or MPP6 depletion. High-fidelity PCR and fluorescence in situ hybridization (FISH) were used to examine the localization and level of 5.8S rRNAs. Western blot was used to examine the protein level. MPP6-EGFP mRNA microinjection was used to do the rescue. RESULTS: MPP6 was enriched within ovaries and oocytes. MPP6 depletion significantly impeded oocyte meiosis. MPP6 depletion increased 5.8S pre-rRNA. The mRNA levels of MPP6 and 5.8S rRNA decreased within ageing oocytes, and MPP6 mRNA injection partially increased 5.8S rRNA maturation and improved oocyte quality. CONCLUSIONS: MPP6 is required for 5.8S rRNA maturation, meiosis and quality control in mouse oocytes, and MPP6 level might be a marker for oocyte quality.

Single xenotransplant of rat brown adipose tissue prolonged the ovarian lifespan of aging mice by improving follicle survival.

Prolonging the ovarian lifespan is attractive and challenging. An optimal clinical strategy must be safe, long-acting, simple, and economical. Allotransplantation of brown adipose tissue (BAT), which is most abundant and robust in infants, has been utilized to treat various mouse models of human disease. Could we use BAT to prolong the ovarian lifespan of aging mice? Could we try BAT xenotransplantation to alleviate the clinical need for allogeneic BAT due to the lack of voluntary infant donors? In the current study, we found that a single rat-to-mouse (RTM) BAT xenotransplantation did not cause systemic immune rejection but did significantly increase the fertility of mice and was effective for more than 5 months (equivalent to 10 years in humans). Next, we did a series of analysis including follicle counting; AMH level; estrous cycle; mTOR activity; GDF9, BMP15, LHR, Sirt1, and Cyp19a level; ROS and annexin V level; IL6 and adiponectin level; biochemical blood indices; body temperature; transcriptome; and DNA methylation studies. From these, we proposed that rat BAT xenotransplantation rescued multiple indices indicative of follicle and oocyte quality; rat BAT also improved the metabolism and general health of the aging mice; and transcriptional and epigenetic (DNA methylation) improvement in F0 mice could benefit F1 mice; and multiple KEGG pathways and GO classified biological processes the differentially expressed genes (DEGs) or differentially methylated regions (DMRs) involved were identical between F0 and F1. This study could be a helpful reference for clinical BAT xenotransplantation from close human relatives to the woman.

Low doses of decitabine improve the chemotherapy efficacy against basal-like bladder cancer by targeting cancer stem cells.

Low dose treatment with the DNA methylation inhibitor decitabine has been shown to be applicable for the management of certain types of cancer. However, its antitumor effect and mechanisms are context dependent and its activity has never been systematically studied in bladder cancer treatment. We used mouse models, cultured cell lines and patient-derived xenografts to demonstrate that low dose decitabine treatment remarkably enhanced the effects of cisplatin and gemcitabine on basal-like bladder cancer both in vivo and in vitro. Genetic lineage tracing revealed that the stemness of a bladder cancer stem cell population was inhibited by decitabine treatment in mice. These effects were accompanied by decreases in genome-wide DNA methylation, gene re-expression, and changes in key cellular regulatory pathways such as STAT3 signaling. These results indicate that this DNA-demethylating reagent is a promising therapeutic approach for basal-like bladder cancer treatment.

The ubiquitin ligase KBTBD8 regulates PKM1 levels via Erk1/2 and Aurora A to ensure oocyte quality.

Tight control of energy metabolism is essential for normal cell function and organism survival. PKM (pyruvate kinase, muscle) isoforms 1 and 2 originate from alternative splicing of PKM pre-mRNA. They are key enzymes in oxidative phosphorylation and aerobic glycolysis, respectively, and are essential for ATP generation. The PKM1:PKM2 expression ratio changes with development and differentiation, and may also vary under metabolic stress and other conditions. Until now, there have been no reports about the function and regulation of PKM isozymes in oocytes. Here, we demonstrate that PKM1 or PKM2 depletion significantly disrupts ATP levels and mitochondrial integrity, and exacerbates free-radical generation and apoptosis in mouse oocytes. We also show that KBTBD8, a female fertility factor in the KBTBD ubiquitin ligase family, selectively regulates PKM1 levels through a signaling cascade that includes Erk1/2 and Aurora A kinases as intermediates. Finally, using RNA sequencing and protein network analysis, we identify several regulatory proteins that may be govern generation of mature PKM1 mRNA. These results suggest KBTBD8 affects PKM1 levels in oocytes via a KBTBD8→Erk1/2→Aurora A axis, and may also affect other essential processes involved in maintaining oocyte quality.

The m6A methyltransferase METTL3 promotes bladder cancer progression via AFF4/NF-κB/MYC signaling network.

N6-methyladenosine (m6A) is the most abundant modification in eukaryotic messenger RNAs (mRNAs), and plays important roles in many bioprocesses. However, its functions in bladder cancer (BCa) remain elusive. Here, we discovered that methyltransferase-like 3 (METTL3), a major RNA N6-adenosine methyltransferase, was significantly up-regulated in human BCa. Knockdown of METTL3 drastically reduced BCa cell proliferation, invasion, and survival in vitro and tumorigenicity in vivo. On the other hand, overexpression of METTL3 significantly promoted BCa cell growth and invasion. Through transcriptome sequencing, m6A sequencing and m6A methylated RNA immuno-precipitation quantitative reverse-transcription polymerase chain reaction, we revealed the profile of METTL3-mediated m6A modification in BCa cells for the first time. AF4/FMR2 family member 4 (AFF4), two key regulators of NF-κB pathway (IKBKB and RELA) and MYC were further identified as direct targets of METTL3-mediated m6A modification. In addition, we showed that besides NF-κB, AFF4 binds to the promoter of MYC and promotes its expression, implying a novel multilevel regulatory network downstream of METTL3. Our results uncovered an AFF4/NF-κB/MYC signaling network operated by METTL3-mediated m6A modification and provided insight into the mechanisms of BCa progression.

SOX2 Is a Marker for Stem-like Tumor Cells in Bladder Cancer.

It has been reported that functionally distinct cancer stem cells (CSCs) exist in human bladder cancer (BCa). Here, we found that Sox2, a transcription factor that is well characterized as a marker for stem cells, is upregulated in both mouse and human BCa. Sox2 expression is absent in normal urothelial cells, but it begins to be expressed in pre-neoplastic bladder tumors and continues to be expressed in invasive mouse BCa. Using s as a reporter of Sox2 transcriptional expression, we demonstrated that Sox2-expressing cells mark a subpopulation of tumor cells that fuel the growth of established BCa. SOX2-positive cells also expressed other previously reported BCa CSC markers, including Keratin14 (KRT14) and CD44v6. Ablation of Sox2-expressing cells within primary invasive BCa led to enhanced tumor regression, supporting the essential role of SOX2-positive cells in regulating BCa maintenance and progression. Our data show that Sox2 is a marker of bladder CSCs and indicate it as a potential clinical target for BCa therapy.

Development of monoclonal antibodies against Sj29 and its possible application for schistosomiasis diagnosis.

OBJECTIVE: Timely Schistosoma japonicum detection improves outcomes in schistosomiasis. Here, we established a double antibody sandwich ELISA to detect Schistosoma japonicum. METHODS: Sj29 polyclonal and monoclonal antibodies were developed and identified. A Sj29 double antibody sandwich ELISA was evaluated. RESULTS: Assay sensitivity for detecting Schistosoma japonicum circulating antigen Sj29 was 76.7% (23/30), 54.5% (18/33) and 50.0% (18/36) in patients with acute, chronic and advanced schistosomiasis. No false positives or cross-reactivity was observed in healthy controls or patients with clonorchiasis, paragonimiasis, or ancylostomiasis, respectively. By contrast, false positives (5.7%) and cross-reactivity (6.5%-10%) were detected using an AWA-ELISA. The circulating antigen positive rates decreased significantly faster than that of the antibody detection after 6 months treatment (22.2%, 4/18 and 88.9%, 16/18). Chi-Square Tests revealed that Sj29 sandwich ELISA had lower sensitivity than AWA indirect ELISA in the detection of S. japonicum infected patients (p<0.05). Although our assay detection specificity in patients infected with other parasites or healthy controls appeared higher, the difference between the assays was insignificant. However, our assay showed significantly better results in monitoring praziquantel therapeutic effects (p=0.001), with antigen-positive rates decreasing significantly faster than antibody detection rates after 6 months of treatment (22.2%, 4/18 versus 88.9%, 16/18). CONCLUSIONS: Sj29 double antibody sandwich ELISA was established. The specificity of this method for detecting healthy sera was 100%. Meanwhile, Sj29 sandwich ELISA may have a potential diagnostic capability to distinguish current from past infections and assess drug treatment responses.

Conditional ablation of TGF-ß signaling inhibits tumor progression and invasion in an induced mouse bladder cancer model.

The role of transforming growth factor-ß (TGF-ß) signaling in cancer progression is still under debate. To determine the function of TGF-ß signaling in bladder cancer progression, we conditionally knocked out the Tgfbr2 in mouse model after a N-butyl-N-4-hydroxybutyl Nitrosamine induced bladder carcinogenesis. We found the ablation of TGF-ß signaling could inhibit the cancer cell proliferation, cancer stem cell population and EMT, hence suppressed the invasive cancer progression, which is similar with the result of TGF-ß receptor I inhibitor treatment. These findings recognize the roles and mechanisms of TGF-ß signaling in bladder cancer progression in vivo for the first time.

Preliminary study of differentiating smears from cancerous and non-cancerous nasopharyngeal tissue using confocal Raman spectroscopy.

PURPOSE: Current practice for diagnosing nasopharyngeal carcinoma (NPC) is based on invasive tissue biopsy. This study aims to explore the feasibility of using Raman spectroscopy to differentiate cancerous and non-cancerous nasopharyngeal tissue smears, expecting to realize minimal invasive diagnosis using smears from in vivo mucosa tissue by Raman spectroscopy. METHODS: Biopsy tissue smears were acquired from 74 patients with pathologically diagnosed nasopharyngeal diseases and measured using confocal Raman spectroscopy. RESULTS: Both fingerprint region and high wavenumber Raman spectra were acquired with distinguish features. Multivariate statistical analysis was used to differentiate cancerous and non-cancerous groups, achieving a diagnostic sensitivity of 87.2 and specificity of 85.7 % for differentiating NPC from nasopharyngeal non-cancerous smears. CONCLUSIONS: This work indicates that the method has a unique advantage in microanalysis for tissue smears which may provide a promising minimal invasive (or noninvasive) diagnosing tool for cancer diagnosis.