Novel therapeutic strategies for injured endometrium: intrauterine transplantation of menstrual blood­derived cells from infertile patients.

BACKGROUND: Menstrual blood-derived cells show regenerative potential as a mesenchymal stem cell and may therefore be a novel stem cell source of treatment for refractory infertility with injured endometrium. However, there have been few pre-clinical studies using cells from infertile patients, which need to be addressed before establishing an autologous transplantation. Herein, we aimed to investigate the therapeutic capacity of menstrual blood-derived cells from infertile patients on endometrial infertility. METHODS: We collected menstrual blood-derived cells from volunteers and infertile patients and confirmed their mesenchymal stem cell phenotype by flow cytometry and induction of tri-lineage differentiation. We compared the proliferative and paracrine capacities of these cells. Furthermore, we also investigated the regenerative potential and safety concerns of the intrauterine transplantation of infertile patient-derived cells using a mouse model with mechanically injured endometrium. RESULTS: Menstrual blood-derived cells from both infertile patients and volunteers showed phenotypic characteristics of mesenchymal stem cells. In vitro proliferative and paracrine capacities for wound healing and angiogenesis were equal for both samples. Furthermore, the transplantation of infertile patient-derived cells into uterine horns of the mouse model ameliorated endometrial thickness, prevented fibrosis, and improved fertility outcomes without any apparent complications. CONCLUSIONS: In our pre-clinical study, intrauterine transplantation of menstrual blood-derived cells may be a novel and attractive stem cell source for the curative and prophylactic therapy for injured endometrium. Further studies will be warranted for future clinical application.

Assessing the performance of the Cell Painting assay across different imaging systems.

Quantitative microscopy is a powerful method for performing phenotypic screens from which image-based profiling can extract a wealth of information, termed profiles. These profiles can be used to elucidate the changes in cellular phenotypes across cell populations from different patient samples or following genetic or chemical perturbations. One such image-based profiling method is the Cell Painting assay, which provides morphological insight through the imaging of eight cellular compartments. Here, we examine the performance of the Cell Painting assay across multiple high-throughput microscope systems and find that all are compatible with this assay. Furthermore, we determine independently for each microscope system the best performing settings, providing those who wish to adopt this assay an ideal starting point for their own assays. We also explore the impact of microscopy setting changes in the Cell Painting assay and find that few dramatically reduce the quality of a Cell Painting profile, regardless of the microscope used.

Assessing the performance of the Cell Painting assay across different imaging systems.

Quantitative microscopy is a powerful method for performing phenotypic screens from which image-based profiling can extract a wealth of information, termed profiles. These profiles can be used to elucidate the changes in cellular phenotypes across cell populations from different patient samples or following genetic or chemical perturbations. One such image-based profiling method is the Cell Painting assay, which provides morphological insight through the imaging of eight cellular compartments. Here, we examine the performance of the Cell Painting assay across multiple high-throughput microscope systems and find that all are compatible with this assay. Furthermore, we determine independently for each microscope system the best performing settings, providing those who wish to adopt this assay an ideal starting point for their own assays. We also explore the impact of microscopy setting changes in the Cell Painting assay and find that few dramatically reduce the quality of a Cell Painting profile, regardless of the microscope used.

Trehalose Suppresses Lysosomal Anomalies in Supporting Cells of Oocytes and Maintains Female Fertility.

Supporting cells of oocytes, i.e., cumulus cells, control oocyte quality, which determines fertilization success. Therefore, the transformation of mature and immature cumulus cells (MCCs and ICCs, respectively) into dysmature cumulus cells (DCCs) with dead characteristics deteriorates oocyte quality. However, the molecular basis for this transformation remains unclear. Here, we explored the link between autophagic decline and cumulus transformation using cumulus cells from patients with infertility, female mice, and human granulosa cell-derived KGN cell lines. When human cumulus cells were labeled with LysoTracker probes, fluorescence corresponding to lysosomes was enhanced in DCCs compared to that in MCCs and ICCs. Similarly, treatment with the autophagy inhibitor chloroquine elevated LysoTracker fluorescence in both mouse cumulus cells and KGN cells, subsequently suppressing ovulation in female mice. Electron microscopy analysis revealed the proliferation of abnormal lysosomes in chloroquine-treated KGN cells. Conversely, the addition of an autophagy inducer, trehalose, suppressed chloroquine-driven problematic lysosomal anomalies and ameliorated ovulation problems. Our results suggest that autophagy maintains the healthy state of the supporting cells of human oocytes by suppressing the formation of lysosomes. Thus, our results provide insights into the therapeutic effects of trehalose on female fertility.

Chemical and biological evaluation of unusual sugars, α-aculosides, as novel Michael acceptors.

The unusual sugars, α-aculosides, which appear in certain antibiotics and have an α,ß-unsaturated ketone structure, were found to be novel and selective Michael acceptors for the thiol function of cysteine residues. A coumarin derivative possessing α-aculoside as a Michael acceptor effectively and irreversibly operated as a fluorescent probe in cells. Furthermore, α-aculosides exhibited cytotoxic activity against several cancer cell lines.

Total synthesis of vineomycin B2.

The first total synthesis of vineomycin B2 (1) has been accomplished. The aglycon segment, a vineomycinone B2 derivative, and the glycon segment, an α-L-acurosyl-L-rhodinose derivative, were prepared via C-glycosylation using an unprotected sugar and powerful chemoselective O-glycosylation using a 2,3-unsaturated sugar, respectively, as the key steps. Furthermore, effective and simultaneous introduction of the two glycon moieties to the aglycon part by concentration-controlled glycosylation led to the total synthesis of 1.

Tumor-suppressive sphingosine-1-phosphate receptor-2 counteracting tumor-promoting sphingosine-1-phosphate receptor-1 and sphingosine kinase 1 - Jekyll Hidden behind Hyde.

Sphingosine-1-phosphate (S1P) is a plasma lipid mediator with multiple roles in mammalian development, physiology and pathophysiology. It is constitutively produced mostly by erythrocytes by the action of sphingosine kinase 1 (SphK1), resulting in high (∼0.5 micromolar) steady-state plasma S1P content and steep S1P concentration gradient imposed between plasma/lymph/tissue interstitial fluid. S1P is also locally produced by activated platelets and tumor cells, in the latter case SphK1 is a downstream target of activated Ras mutant and hypoxia, and is frequently upregulated especially in advanced stages of tumors. Most if not all of the S1P actions in vertebrates are mediated through evolutionarily conserved G protein-coupled S1P receptor family. Ubiquitously expressed mammalian subtypes S1PR1, S1PR2 and S1PR3 mediate pleiotropic actions of S1P in diverse cell types, through coupling to distinctive repertoire of heterotrimeric G proteins. S1PR1 and S1PR3 mediate directed cell migration toward S1P through coupling to G(i) and activating Rac, a Rho family small G protein essential for cell migration. Indeed, S1PR1 expressed in lymphocytes directs their egress from lymph nodes into lymph and recirculation, serving as the target for downregulation by the immunosuppressant FTY720 (fingolimod). S1PR1 in endothelial cells plays an essential role in vascular maturation in embryonic stage, and mediates angiogenic and vascular protective roles of S1P which include eNOS activation and maintenance of barrier integrity. It is likely that S1PR1 and SphK1 expressed in host endothelial cells and tumor cells act in concert in a paracrine loop to contribute to tumor angiogenesis, tumor invasion and progression. In sharp contrast, S1PR2 mediates S1P inhibition of Rac at the site downstream of G(12/13)-mediated Rho activation, thus identified as the first G protein-coupled receptor that negatively regulates Rac and cell migration. S1PR2 could also mediate inhibition of Akt and cell proliferation/survival signaling via Rho-ROCK-PTEN pathway. S1PR2 expressed in tumor cells mediates inhibition of cell migration and invasion in vitro and metastasis in vivo. Moreover, S1PR2 expressed in host endothelial cells and tumor-infiltrating myeloid cells in concert mediates potent inhibition of tumor angiogenesis and tumor growth in vivo, with inhibition of VEGF expression and MMP9 activity. These recent findings provide further basis for S1P receptor subtype-specific, novel therapeutic tactics for individualized treatment of patients with cancer.

Degradation of reactive dyes by ozonation and oxalic acid-assimilating bacteria isolated from soil.

Ozonation and treatment of wastewaters with oxalic acid-assimilating bacterium was attempted for the complete degradation of reactive dyes. Oxalic acid-assimilating bacterium, Pandoraea sp. strain EBR-01, was newly isolated from soil under bamboo grove and was identified to be a member of the genus Pandoraea by physicochemical and biochemical tests including 16S rDNA sequence analysis. The bacterium was grown optimally at pH 7 and temperature of 30 degrees C under the laboratory conditions. Reactive Red 120 (RR120), Reactive Green 19 (RG19), Reactive Black 5 (RB5) and Remazol Brilliant Blue R (RBBR) were used in degradation experiments. At the initial reactive dye concentrations of 500 mg/l and the ozonation time of 80 min, it was confirmed that 75-90 mg/l oxalic acid was generated from reactive dyes by ozonation. Microbial treatment using EBR-01 greatly decreased the amount of oxalic acid in the mixture after 48 h, but it was not removed completely. TOC/TOC(0) of reactive dye solutions was also decreased to 80-90% and 20-40% by ozonation and microbial treatment using EBR-01, respectively. The study confirmed that consecutive treatments by ozone and microorganisms are efficient methods to mineralize reactive dyes.