Effects of preservation time on proliferative potential of human limbal stem/progenitor cells

AIM: To determine the proliferative potential and the maintenance of stem cell activity in stored human limbal tissues, and correlate this with the preservation time, cell viability and the expression of stem cell markers. METHODS: Thirty limbal rims were split into 4 parts and stored in corneal preservation medium at 4 degrees C for 0, 1, 4, or 7 days. The limbal stem cell and mitotic markers P63, CK19, proliferating cell nuclear antigen (PCNA), and Ki67 were determined by immunohistochemical staining. The proliferative potential of limbal epithelial cells was assessed by cell viability, the ability of generating stratified epithelium, and colony forming assay. RESULTS: The stored tissues maintained limbal stratified structure to 7 days and exhibited comparable expression level of stem cell and mitotic markers. The proportion of viable cells decreased with the prolonged preservation time, while colony forming efficiency decreased from the 1st day and disappeared at the 4th day. When inoculated on amniotic membrane, the cells preserved for 1 day formed a stratified epithelium, while the cells from 4 days' preservation formed a discontinuous layer. CONCLUSION: The colony forming efficiency of limbal epithelial stem/progenitor cells decreased rapidly with the increasing preservation time, while the expression level of markers and capacity of forming epithelial monolayer on amniotic membrane decreased gradually. The limbal epithelial stem cells lost their function earlier than the lost expression level of stem cell markers. This may help us to better choose the appropriate preservation grafts for future limbal stem cell transplantation.

Effects of Ovarian Preservation Time on in vitro Maturation and in vitro Fertilization of Oocytes from Slaughtered Sheep

[Objectives] The aim was to study the effects of ovarian preservation time on in vitro fertilization of oocytes from slaughtered sheep. [Methods] The collected ovaries were randomly and evenly divided into four groups. They were preserved in physiological saline containing penicillin（ 100 IU/ml） and streptomycin（ 100 μg/ml） at 15-20 ℃ for 0（ Control）,6,12 and 18 h,respectively. Then,the oocytes were subjected to in vitro fertilization. [Results]The maturation rates,cleavage rates and blastocyst rates of the oocytes preserved for 6 and 12 h showed no significant differences compared with those of the oocytes preserved for0 h（ 72. 03%,70. 87% vs. 73. 68%; 74. 12%,72. 60% vs. 74. 49%; 22. 22%,20. 75% vs. 23. 29%）（ P 〉 0. 05）. There were also no significant differences in maturation rate,cleavage rate or blastocyst rate between the oocytes preserved for 18 and 0 h（ P 〉 0. 05）. [Conclusions] Within a certain rage（ 0-18 h）,storage time of ovary at 15-20 ℃ does not affect the continued development of oocytes from slaughtered sheep.

A tunable XUV monochromatic light source based on the time preserving grating selection of high-order harmonic generation

We accomplish a laboratory facility for producing a femtosecond XUV coherent monochromatic radiation with a broad tunable spectral range of 20 eV-75 eV. It is based on spectral selected single-order harmonics from intense laser driven high harmonic generation in gas phase. The time preserving for the selected harmonic radiation is achieved by a Czerny-Turner type monochromator designed with a conical diffraction grating mount for minimizing the time broadening caused by grating diffraction and keeping a relatively high diffraction efficiency. Our measurement shows that the photon flux of the 23-order harmonic（H23） centered at 35.7 eV is 1×10~9 photons/s approximately with a resolving power E/？E ≈ 36.This source provides an ultrashort tunable monochromatic XUV beam for ultrafast studies of electronic and structural dynamics in a large variety of matters.