Ganoderma lucidum polysaccharides prevent platelet-derived growth factor-stimulated smooth muscle cell proliferation in vitro and neointimal hyperplasia in the endothelial-denuded artery in vivo.

Ganoderma lucidum is used in traditional Chinese medicine to prevent or treat a variety of diseases, including cardiovascular disorders. We previously demonstrated that a glucan-containing extract of Reishi polysaccharides (EORP) has the potent anti-inflammatory action of reducing ICAM-1 expression in lipopolysaccharide (LPS)-treated human aortic smooth muscle cells (HASMCs) and LPS-treated mice. In the present study, we examined whether EORP inhibited platelet-derived growth factor-BB (PDGF)-stimulated HASMC proliferation and the mechanism involved. EORP dose-dependently reduced cell numbers and DNA synthesis of PDGF-treated HASMCs in vitro. EORP also arrested cell cycle progression in the G0/G1 phase, and this was associated with decreased expression of cyclin D1, cyclin E, CDK2, CDK4, and p21(Cip1) and upregulation of the cyclin-dependent kinase inhibitor p27(Kip1). The anti-proliferative effect of EORP was partly mediated by downregulation of PDGF-induced JNK phosphorylation. In in vivo studies, the femoral artery of C57BL/6 mice was endothelial-denuded and the mice were fed a diet containing 100 mg/kg/day of EORP. On day 14, both cell proliferation (proliferating cell nuclear antigen-positive cells) in the neointima and the neointima/media area ratio (0.67 ± 0.03 vs. 1.46 ± 0.30) were significantly reduced. Our data show that EORP interferes with the mitogenic activation of JNK, preventing entry of HASMCs into the cell cycle in vitro and reducing cell proliferation in the neointima and decreasing the neointimal area in vivo. Thus, EORP may represent a safe and effective novel approach to the prevention and treatment of vascular proliferative diseases.

Is there an optimal timing interval between hCG trigger and oocyte vitrification?

OBJECTIVE: Oocyte vitrification has been developed as a promising alternative to slow freezing; however, the clinical outcome is highly operator dependent. From the past study, we know the timing of cryoprotectant exposure and understand that the intervals between the application of liquid nitrogen and thawing solution are crucial factors in the vitrification process. However, the optimal time intervals between hCG trigger and oocyte vitrification and equilibration remain unknown. This study aimed to evaluate the optimal intervals before and during modified vitrification. MATERIALS AND METHODS: This retrospective study included 66 patients undergoing vitrified-thawed oocyte cycles from June 2018 to May 2019. Oocyte in vitro maturation (IVM) is defined as the maturation in vitro of an immature oocyte collected using a standard pick up procedure. Oocytes were grouped into the following intervals: (1) human chorionic gonadotropin (hCG) trigger to oocyte vitrification (<38 h; 38-39 h; >39 h; IVM) (2) oocyte equilibration time (<10 min; 10-12 min; 12-15 min). The vitrification and warming procedures were performed following the steps as shown in the Cryotec method. RESULTS: A total of 390 mature oocytes were vitrified with the Cryotec method. The survival rates were not significantly different among the various intervals after the hCG trigger (97.59%; 95.54%; 100%); however, there was a trend of decreased survival rate in IVM group (66.67%). The oocyte survival rates were not significantly different among the various times of oocyte equilibration (96.77%; 97.33%; 95.42%). CONCLUSIONS: This was the first study to demonstrate no correlation between oocyte survival rate and the time intervals between hCG trigger and oocyte vitrification. Nor did the oocyte survival rate correlate with the various equilibration times during vitrification, as long as the oocyte was mature. In the future, large, prospective, randomized controlled studies will be required to confirm the clinical outcomes.

Metaphase II (MII) human oocytes with smooth endoplasmic reticulum clusters do not affect blastocyst euploid rate.

OBJECTIVE: To investigate whether the rate of euploidy and pregnancy outcomes are affected by smooth endoplasmic reticulum clusters (SERc) and other metaphase II human oocyte dysmorphisms. MATERIALS AND METHODS: Retrospective analysis of the morphologies of metaphase II (MII) human oocytes, which had developed into 590 biopsied blastocysts derived from 109 patients that received preimplantation genetic testing for aneuploidies (PGT-A) cycles between March 2013 and December 2017. The euploid rate of blastocysts that originated from morphologically abnormal or normal oocytes were analyzed. The chromosome status of the blastocysts was determined and analyzed by array comparative genomic hybridization (aCGH) or next generation sequencing (NGS) following trophectoderm biopsy. RESULTS: According to the odds ratios obtained for each oocyte morphotype, no statistically significant relationship was found between oocyte dysmorphisms and euploid rate. Specifically, although SERc-positive oocytes had a higher rate of arrest at two pronuclei, or 2 PN (26.7% vs. 19.4%, p > 0.05), the blastocyst formation rate was not affected as compared with SERc-negative oocytes (40.0% vs. 38.6%, p > 0.05). Among nine euploid embryos derived from oocytes with SERc, three single euploid embryo transfers were performed, of which one resulted in blighted ovum, and two resulted in the births of two healthy, singleton term babies. CONCLUSION: The results presented here suggest that oocyte dysmorphisms do not affect the euploidy rate of the blastocyst. The occurrence of SERc in the oocyte does not seem to impair the developing blastocyst nor does it interfere with good embryo formation rate and euploid rate. Thus, the embryos derived from SERc-positive oocytes could still be considered for embryo transfer if there are no other embryos available.

Clinical application of immunomagnetic reduction for quantitative analysis of beta-subunit of human chorionic gonadotropin in blastocyst culture media to differentiate embryo quality.

BACKGROUND: The most important factor for a successful pregnancy after in vitro fertilization is embryo quality. The aim of this study was to explore the possibility that using the immunomagnetic reduction (IMR) assay to quantitatively measure ß-subunit of human chorionic gonadotropin (ß-hCG) in blastocyst culture media to differentiate embryo quality. METHODS: This was a prospective case-control study including 28 samples of blastocyst culture media. We used single-step blastocyst culture and IMR assay to analyze ß-hCG concentrations in culture media. We also explored the relationship between IMR signals of ß-hCG and morphological grading of blastocysts. RESULTS: ß-hCG concentration-dependent IMR signals were highly correlated with blastocyst morphological quality (Spearman correlation coefficient: 0.731). Receiver-operating characteristic curve analysis showed a cut-off IMR value to differentiate embryo quality of 0.873%, with an area under the curve of 0.947, sensitivity of 0.882 and specificity of 0.818. Furthermore, subanalysis also revealed a positive correlation between ß-hCG concentration-dependent IMR signals and trophectoderm grading, with a Spearman correlation coefficient of 0.576. CONCLUSIONS: An IMR assay can quantitatively measure ß-hCG in blastocyst culture media, and may be a potential clinical tool to assist in the assessment of good blastocyst quality before embryo transfer.

Early blastulation of day 4 embryo correlates with the increased euploid rate of preimplantation genetic screening cycles.

OBJECTIVE: It is known that embryos with faster growing potential, especially in blastocyst development, correlate with the increased euploid rate. Our study investigated the preimplantation genetic screening cycle to analyze the correlation between early blastulation (EB) on day 4 embryo and the euploid rate. MATERIALS AND METHODS: This is a retrospective study examining 273 biopsied blastocysts after preimplantation genetic screening obtained from 54 patients from March 2013 to March 2017. Of the 273 biopsied embryos, 81 had early blastulation on day 4 and were classified as the EB (+) group, while the other 192 had no early blastulation and were classified as the EB (-) group. Euploid rates were compared between the two groups. A total of 34 single euploid embryos were transferred, with 14 from the EB (+) group and 20 from the EB (-) group. Clinical pregnancy was compared between the groups. RESULTS: There is a statistically significant increase in the euploid rate in the EB (+) group (49.4% vs. 34.4%, p = 0.02). The clinical pregnancy rate was also increased in the single euploid embryo transfer group with early blastulation, but did not reach statistical significance (71.4% vs. 50.0%, p = 0.211). CONCLUSIONS: Early blastulation of day 4 embryo correlates significantly with the euploid rate. Early blastulation of day 4 embryo may serve as a potential aid for embryo selection for transfer in preimplantation genetic screening cycles.