Objective:To investigate the effects of vitrification and transplantation on mouse ovarian tissues.Methods:Female ICR mice were divided into three groups: fresh-control group ( in vitro oocyte maturation and fertilization, n=9), fresh-transplanted group (ovarian tissue transplantation, in vitro oocyte maturation and fertilization, n=9), frozen-transplanted group (ovarian tissue vitrification-thawed-transplantation, in vitro oocyte maturation and fertilization, n=9). Additionally, frozen-thawed group (ovarian tissue vitrification-thawed, n=6) and ovariectomy group (ovariectomy, n=6) were also set up, to more directly explain the effects of vitrification and freezing on the number of follicles and the changes of endocrine function after ovarian transplantation. We determined the number of follicles by hematoxylin-eosin (HE) staining, neovascularization by CD31 immunohistochemical staining, tissue fibrosis by Masson staining, and serum sex hormone levels by enzyme-linked immunosorbent assay (ELISA) three weeks following ovarian tissue transplantation. In addition, we counted the number of oocytes obtained, oocytes in vitro fertilized, as well as blastocysts. Results:The number of total follicles in both the fresh-transplanted group and the frozen-transplanted group significantly decreased compared with the fresh-control group (all P<0.001) and the frozen-thawed group (all P<0.001). The CD31-positive rate of ovarian tissues in the fresh-transplanted group was significantly higher than that in the fresh-control group ( P=0.044). Although the CD31-positive rate was higher in the frozen-transplanted group than in the fresh-control group, there was no statistical distinction ( P=0.162). The fibrosis area percentage of ovarian tissues in both the fresh-transplanted group and the frozen-transplanted group increased significantly compared with the fresh-control group ( P=0.004; P=0.005). Serum estradiol level in the fresh-transplanted group and the frozen-transplanted group was significantly lower than that in fresh-control group ( P=0.005; P=0.001), significantly higher than that in the ovariectomy group ( P=0.011; P=0.035). Serum follicle-stimulating hormone (FSH) level in the fresh-transplanted group and the frozen-transplanted group was significantly higher than that in the fresh-control group ( P=0.040; P=0.012), significantly lower than that in the ovariectomy group ( P=0.001; P=0.004). In comparison to the fresh-control group, the number of oocytes retrieved in the fresh-transplanted group and the frozen-transplanted group decreased significantly (all P<0.001). Furthermore, the number of oocytes retrieved in the fresh-transplanted group was higher than that in the frozen-transplanted group, and yet there was no statistical difference ( P=0.272). And the number of oocytes in vitro fertilized and blastocysts in the fresh-transplanted group and the frozen-transplanted group were significantly lower than those in the fresh-control group (all P<0.001). The number of total follicles, CD31-positive rate, fibrosis area rate, serum estradiol, and FSH levels, the number of oocytes in vitro fertilized and blastocysts were no substantially distinction between the fresh-transplanted group and the frozen-transplanted group (all P>0.05). Conclusion:?After ovarian tissue vitrification-thawed and transplantation, follicular growth, endocrine function, and fertility were restored in the mouse model, confirming that ovarian tissue vitrification is an effective method for female fertility preservation. Both vitrification and transplantation could cause follicles to be lost and fertility to decrease. And post-transplantation stage is the primary stage of follicle loss during ovarian tissue frozen-thawed-transplantation, and transplantation is the predominant factor affecting the effectiveness of ovarian tissue frozen-thawed-transplantation.

Objective:To investigate the effects of vitrification and transplantation on mouse ovarian tissues.Methods:Female ICR mice were divided into three groups: fresh-control group ( in vitro oocyte maturation and fertilization, n=9), fresh-transplanted group (ovarian tissue transplantation, in vitro oocyte maturation and fertilization, n=9), frozen-transplanted group (ovarian tissue vitrification-thawed-transplantation, in vitro oocyte maturation and fertilization, n=9). Additionally, frozen-thawed group (ovarian tissue vitrification-thawed, n=6) and ovariectomy group (ovariectomy, n=6) were also set up, to more directly explain the effects of vitrification and freezing on the number of follicles and the changes of endocrine function after ovarian transplantation. We determined the number of follicles by hematoxylin-eosin (HE) staining, neovascularization by CD31 immunohistochemical staining, tissue fibrosis by Masson staining, and serum sex hormone levels by enzyme-linked immunosorbent assay (ELISA) three weeks following ovarian tissue transplantation. In addition, we counted the number of oocytes obtained, oocytes in vitro fertilized, as well as blastocysts. Results:The number of total follicles in both the fresh-transplanted group and the frozen-transplanted group significantly decreased compared with the fresh-control group (all P<0.001) and the frozen-thawed group (all P<0.001). The CD31-positive rate of ovarian tissues in the fresh-transplanted group was significantly higher than that in the fresh-control group ( P=0.044). Although the CD31-positive rate was higher in the frozen-transplanted group than in the fresh-control group, there was no statistical distinction ( P=0.162). The fibrosis area percentage of ovarian tissues in both the fresh-transplanted group and the frozen-transplanted group increased significantly compared with the fresh-control group ( P=0.004; P=0.005). Serum estradiol level in the fresh-transplanted group and the frozen-transplanted group was significantly lower than that in fresh-control group ( P=0.005; P=0.001), significantly higher than that in the ovariectomy group ( P=0.011; P=0.035). Serum follicle-stimulating hormone (FSH) level in the fresh-transplanted group and the frozen-transplanted group was significantly higher than that in the fresh-control group ( P=0.040; P=0.012), significantly lower than that in the ovariectomy group ( P=0.001; P=0.004). In comparison to the fresh-control group, the number of oocytes retrieved in the fresh-transplanted group and the frozen-transplanted group decreased significantly (all P<0.001). Furthermore, the number of oocytes retrieved in the fresh-transplanted group was higher than that in the frozen-transplanted group, and yet there was no statistical difference ( P=0.272). And the number of oocytes in vitro fertilized and blastocysts in the fresh-transplanted group and the frozen-transplanted group were significantly lower than those in the fresh-control group (all P<0.001). The number of total follicles, CD31-positive rate, fibrosis area rate, serum estradiol, and FSH levels, the number of oocytes in vitro fertilized and blastocysts were no substantially distinction between the fresh-transplanted group and the frozen-transplanted group (all P>0.05). Conclusion:?After ovarian tissue vitrification-thawed and transplantation, follicular growth, endocrine function, and fertility were restored in the mouse model, confirming that ovarian tissue vitrification is an effective method for female fertility preservation. Both vitrification and transplantation could cause follicles to be lost and fertility to decrease. And post-transplantation stage is the primary stage of follicle loss during ovarian tissue frozen-thawed-transplantation, and transplantation is the predominant factor affecting the effectiveness of ovarian tissue frozen-thawed-transplantation.

Objective:To study the distribution and drug resistance of Carbapenem-Resistant Organism (CRO) and to analysis the risk factors of CRO 30-day mortality.Methods:A total of 181 patients with CRO infection diagnosed in Department of Hematology, Fujian Medical University Union Hospital from January 2018 to June 2020 were retrospectively investigated. The clinical and laboratory data of the patients were collected, the prognosis of patients diagnosed with CRO infection in day 30 was followed up, and the risk factors of prognosis were analyzed. The clinical significance of Carbapenem-Resistant Enterobacteriaceae (CRE) active screening was further evaluated in the CRE subgroup.Results:Among the total of 181 CRO isolates, 47.2% were CRE, 37.0% were Pseudomonas aeruginosa, and 32.6% were Klebsiella pneumoniae, which were highly resistant to carbapenem and had high MIC value, 76.8% (139/181) of CRO were MIC of imipenem resistance≥16 μg/ml. The main sources of isolates were blood and sputum. The 30-day all-cause mortality rates of patients with CRO or CRE infection were (41.4±3.7) % and (44.7±5.4) %, respectively. The COX multivariate regression analysis showed that the level of procalcitonin >0.2 ng/ml and the MIC value of imipenem resistance ≥ 16 μg/ml were independent risk factors for 30-day mortality of CRO infected patients. The CRE subgroup analysis showed that MIC value of imipenem resistance ≥16 μg/ml were independent risk factors for 30-day mortality of CRE infected patients. The 30-day cumulative survival rate of patients with CRE active screening was higher than the patients without CRE active screening [ (68.0±9.3) % vs (50.0±6.5) %, P=0.21]. Conclusion:The high MIC value of imipenem resistance isolates seriously affects the prognosis of patients with CRO infection in the hematology department, and the mortality rate was high. CRE active screening is expected for early prevention, early diagnosis, and early treatment for high-risk patients.

AIM: To clone mouse pdx-1 gene and construct its eukaryotic expression vector for expression of pdx-1 in mouse embryonic stem cells.METHODS: Mouse pdx-1 cDNA fragment was amplified with polymerase chain reaction (PCR) from mouse pancreatic cDNA. The purified fragment was recombinated with a eukaryotic expression vector carrying enhanced green fluorescent protein, pEGFP-N1. The pdx-1 cDNA fragment was inserted into the multi-clone sites of the vector to construct a new plasmid, pEGFP/pdx-1. E.colli strain DH5? was transfected with the new recombinant plasmid to expand it. Plasmid DNA extracted from the expanded DH5? was identifed by cutting with Hind Ⅲ, BamHⅠ nuclease and by DNA sequencing. Identified plasmid DNA was transfected into mouse embryonic stem cell line MESPU13 by carrying with liposome. RESULTS: A 876 bp cDNA fragment was amplified from mouse pancreatic cDNA by PCR and it was inserted into the vector pEGFP-N1 correctly. The fragment was defined to be pdx-1 gene by nuclease digestion and DNA sequencing. Mouse embryonic stem cell line MESPU13 was transfected with the new recombinant plasmid DNA. The green fluorescent protein report gene and pdx-1 gene expressed in transfected mouse embryonic stem cells within 24 h. CONCLUSION: Mouse pdx-1 gene is cloned and its recombinant eukaryotic expression vector carrying green fluorescent protein is constructed successfully. It provides a useful tool for further research on the function of pdx-1.