Profile of biological characterizations and clinical application of corneal stem/progenitor cells

Corneal stem/progenitor cells are typical adult stem/progenitor cells.The human cornea covers the front of the eyeball,which protects the eye from the outside environment while allowing vision.The location and function demand the cornea to maintain its transparency and to continuously renew its epithelial surface by replacing injured or aged cells through a rapid turnover process in which corneal stem/progenitor cells play an important role.Corneal stem/progenitor cells include mainly corneal epithelial stem cells,corneal endothelial cell progenitors and corneal stromal stem cells.Since the discovery of corneal epithelial stem cells(also known as limbal stem cells)in 1971,an increasing number of markers for corneal stem/progenitor cells have been proposed,but there is no consensus regarding the definitive markers for them.Therefore,the identification,isolation and cultivation of these cells remain challenging without a unified approach.In this review,we systematically introduce the profile of biological characterizations,such as anatomy,characteristics,isolation,cultivation and molecular markers,and clinical applications of the three categories of corneal stem/progenitor cells.

DL0805 derivatives protect the pulmonary arterial cells via the RhoA/ROCK pathway

OBJECTIVE Pulmonary artery hypertension(PAH)is a severe disease characterized by the mean pulmonary artery pressure exceeding 25 mm Hg at rest.PAH could induce right heart failure and has a very high mortality rate.At present,several kinds of drugs have been used in the treatment of PAH.However,most of these drugs aim to relax pulmonary arteries and do not inhibit the injury of vessels.In other words,the drugs available for PAH treatment do not improve the survival rate of PAH patients and cannot satisfy the needs in clinic.To discover and develop novel candidate compounds effective on the treatment of pulmonary artery injury and remodeling will be very important.Based on these background,the present study aimed to study the protective effect of two novel Rho-kinases(Rho-associated coiledcoil forming protein serine/threonine kinase,ROCK)inhibitors,DL0805 derivatives(DL0805-1and DL0805-2),on pulmonary arterial cells and further evaluate the underlying mechanisms and the possibility of DL0805 derivatives become therapeutic drugs for PAH.METHODS The primary cultured pulmonary arterial cells including human pulmonary artery endothelium cells(HPAECs)and human pulmonary artery smooth muscle cells(HPASMCs)were used in this study.HPAECs were injured under hypoxia environment(1%O2)and treated with or without DL0805 derivatives.After 48 h,the proliferation and oxidative stress were observed.CCK8 was used to detect cell viability.DCFH-DA was used as probe for reactive oxygen species(ROS)under fluorescence imaging system.HPASMCs was stimulated by growth factors including platelet-derived growth factor-BB(PDGF-BB)and Fetal Bovine Serum(FBS).The proliferation was observed in the cells treated with or without DL0805 derivatives.HPASMCs treated with or without DL0805 derivatives were further incubated with endothelin(ET-1),the proliferation and cytoskeleton remodeling of cells were detected by immunofluorescence assay.At last,Western blotting(WB)and immunofluorescence assay were employed to analysis the underlying mechanisms in the above experiments.RESULTS 10μmol·L-1DL0805-2 could inhibit the proliferation of HPAECs induced by hypoxia.Each concentration of DL0805-1 and DL0805-2attenuated the production of ROS in HPAECs.Results from WB indicated that DL0805 derivatives decreased the injury of HPAECs induced by hypoxia through the inhibition of the expression of Rho A and the activity of ROCK.On HPASMCs,DL0805 derivatives reduced the proliferation induced by PDGF-BB and FBS and inhibited cytoskeleton remodeling induced by ET-1.Immunofluorescence assay showed that DL0805 derivatives inhibited ROCK activity and down regulated the phosphorylation levels of ROCK substrates.CONCLUSION DL0805derivatives exhibited protective effect on pulmonary arterial cells including endothelium cells and smooth muscle cells.Among the above experiments,DL0805-2 showed stronger potency than DL0805-1.These two compounds might protect the cells through the inhibition of Rho A/ROCK pathway and they probably have the potential in the treatment of PAH and deserve further evaluation.

Hydroxysafflor Yellow A Attenuate LipopolysaccharideInduced Endothelium Inflammatory Injury

Objective： This study observed attenuating effect of hydroxysafflor yellow A （HSYA）, an effective ingredient of aqueous extract of Carthamus tinctorius L, on lipopolysaccharide （LPS）-induced endothelium inflammatory injury. Methods： Eahy926 human endothelium cell （EC） line was used; thiazolyl blue tetrazolium bromide （MTT） test was assayed to observe the viability of EC; Luciferase reporter gene assay was applied to measure nuclear factor- κB （NF- κ B） p65 subunit nuclear binding activity in EC; Western blot technology was used to monitor mitogen activated protein kinase （MAPKs） and NF- κ B activation. Reverse transcription polymerase chain reaction （RT-PCR） method was applied to observe intercellular cell adhesion molecule-1 （ICAM-1） and E-selectin mRNA level; EC surface ICAM-1 expression was measured with flow cytometry and leukocyte adhesion to EC was assayed with Rose Bengal spectrophotometry technology. Results： HSYA protected EC viability against LPS-induced injury （P〈0.05）. LPS-induced NF- κ B p65 subunit DNA binding （P〈0.01） and nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor α （I κ B α） phosphorylation was inhibited by HSYA. HSYA attenuated LPS triggered ICAM-1 and E-selectin mRNA levels elevation and phosphorylation of p38 MAPK or c-Jun N-terminal kinase MAPK. HSYA also inhibited LPS-induced cell surface ICAM-1 protein expression （P〈0.01） and leukocyte adhesion to EC （P〈0.05）. Conclusion： HSYA is effective to protect LPS-induced high expression of endothelium adhesive molecule and inflammatory signal transduction.