| Diabetes is one of serious diseases that hazard to human health and quality of life.TypeⅠand Ⅱ diabetes are mainly depended on injecting insulin therapy, but the effectof exogenous insulin is not very satisfactory. In recent years, pancreas transplantation iswidely used in clinical application, which can improve the complications of diabetes andrecover to a certain extent glucose homeostasis to a certain extent.However, the surgery isvery complex, and has some risks. Islet cell transplatation is a new method in recent years,which based on molecular biology, immunology, biology and biologicalengineering.Compared in terms of pancreas transplantation, islet transplantation is moreeasy, secure, and less invasive, faster recovery, and at the same time, it can also bemodified in vitro, so islet cell transplantation has become a hot treatment of diabetes.However, fewer donor islet cells can not meet the needs of clinical applications.So westart looking for alternative sources of islet cells. Stem cells are a class of infiniteproliferative and pluripotent capacity cells, which can differentiate into three germ layersof a variety of tissue cells.Under certain conditions, stem cells can be induced into avariety of cells, such as islet cells.However, in recent years, domestic and foreign researchand our results suggest that,although using different induction programs to different stemcells into islet β cell, it will face similar problems, scilicetly it is hard for the cellsdifferentiation into mature, and mostly are islet precursor cells, which express severalsigns simultaneously, and it will take a long time for these cells to aturation whentransplanting in vivo. Therefore, in-depth study of islet cell maturation molecularmechanism to further promote looking for highly efficient and stable islet celldifferentiation and maturation of the new program is still the research focus.In islet cell development process, many temporal and spatial expression oftranscription factors determines the islet cell differentiation and maturation, such asPDX-1、Neurogenin3、NeuroD、Pax4. Recent studies show that, NRSF is involved in theregulation of insulin secretion of mature islet cells. Abderrahmani showed that inNRSF-overexpressing tumor cell lines, the insulin glucose-insulin secretion coupling is in damage mechanisms. Depth study of specific protein complexin I found neuralregulation of islet cell glucose-induced insulin release function, while complexin Iexpression is negatively regulated by the transcriptional NRSF. Martin et produced isletcell-specific transgenic overexpression of NRSF (insulin promoter-NRSF) mice, theresults indicated that these transgenic mice islet β cells decreased, total pancreatic insulincontent decreased twice. Because of its ability impaired insulin secretion, these mice couldnot tolerate high concentrations of glucose. Depth study found involved in the regulationof gene expression exocytosis was significantly inhibited, such as SNAP25、SYTIV、SYTVII、SYTIX、complexin II.These findings suggested that NRSF involved in theregulation of islet cell function. Recently we found that CART gene was also regulated byNRSF, which was mainly expressed in neuroendocrine cells, such as neurons, pituitarycells, islet cells. Studies had shown that CART gene is involved in the regulation ofglucose-stimulated insulin secretion. In addition, we also found that NRSF directlyregulate the human insulin gene expression, human insulin promoter sequence exists inNRSE, NRSF protein with the sequence of human insulin binds to and inhibits thetranscriptional activity of the promoter. Downregulated expression of NRSF will besignificantly increased transcriptional activity of insulin promoter. These findingssuggested that, NRSF in pancreatic β cells disappear entirely possible to establishdifferentiation and maturation, with intact islet cell secretory response required.Amniotic fluid was often been discarded as medical waste. However, there are also a lotof amniotic fluid stem cells in amniotic fluid. In this article, we isolated amniotic fluidstem cells(AFSCs) from the amniotic fluid, which could express stem cells surfacemarkers, such as SSEA-4、Nanog、Oct4. It could aslo be differentiated into osteogenic,neurogenic, hepatic cells.Then we transfected NRSF interference by lentiviral vectorsinto the amniotic fluidstem cells, and found that down-regulation expression of NRSF,the islet related genesPax4and Insulin expression was significantly increased by about4.5times,19.2timesrespectively. We further performed on amniotic fluid stem cells induced in two-stepmethod, and found that in the first7days induction, the cells showed epithelioid cell growth, and in the14days had been part of the growth of spherical cells withcharacteristics similar to islet cells.Then we detected the stem cell surface markers, andfound that the cells have lost the characteristics of stem cells. Instead, it expressed isletcell surface marker, such as Pdx1, Insulin and C-peptide. Using qRT-PCR methodconfirmed that when NRSF silencing expression, the islet related genes expression wassignificantly elevated.Through the siRNA experiments we found that NRSF, CoREST, HDAC1, mSin3Ainhibition of proteins involved in the regulation transcription of the CART. Then, weoverexpressed NRSF and CART gene in HeLa, also confirmed that NRSF and corepressorCoREST, HDAC1, mSin3A plays an important role in the transcription process of theCART gene. Then we went through a luciferase reporter system confirmed that, bybinding with the CART gene promoter and intron of NRSE, recruitment formed similarbut not identical transcriptional complexes to play the regulation precision on the CARTgene. Through this part of the experiment, we could have a good understanding of themechanism that NRSF silencing can promote stem cells into insulin-producing cells. In recent years, with the rapid development of science and technology, nuclear energyhas been widely used in defense, medical, industrial, agriculture and many other fields,such as nuclear power, nuclear weapons research, the food and drug sterilization andtumor radiotherapy. However, nuclear energy brings great benefits as well as seriousnuclear radiation hazards to the world. Nowadays, countries with nuclear weapons areincreased gradually which increase the danger of nuclear war, and bring the society a greatdeal of instability factors. China has many nuclear forces, such as nuclear weapons andnuclear submarines, also we built a large number of nuclear power plants. Therefore, thereis great significance to protect these employees health from radiation injuries. With thedevelopment of medical technology, radiation therapy has been tremendous developmentand application of technology in the treatment of malignant tumors and other diseases.Although current technology has been available for a specific site or tumor selectiveradiation therapy, however, the normal tissues and cells of our body can also be killed atthe same time, which limits the development of the technology of radiotherapy. In additionto these large doses of radiation, there are also a lot of low doses of radiation in theenvironment around us. With the growing dependence on electronic products, computers,mobile phones and microwave ovens bring us great convenience have become anecessity of our life and have become a necessity of our life, but we also suffer from theradiation injury at the same time in such a radiation environment. In2011, Japan’sFukushima nuclear power plant nuclear leakage accident caused injury to some people andhas brought great harm to the surrounding atmosphere and marine environment. Thenuclear accident caused a great panic of people around the world on the nuclear accidentnuclear radiation once again after the Piceno Bailey Soviet nuclear power plant accident.Therefore, further research is needed to investigate high efficiency, low toxicity ofradiation damage repellant and radiation damage repair agent.The hematopoietic system is the system of radiation sensitive. When radiationexposure has occurred, the main pathological damage of radiation damage is the slowrepair of the hematopoietic of bone marrow, which may always be accompanied by radiation damage and may lead to the death of patients with acute radiation sickness.Currently, the only clinical use of hematopoietic stem cell mobilization agent is G-CSF,butits existence medication for a long time, toxic side effects and other defects.In the presentstudy, we report that, the small molecule Me6, which is an alkaloid analogues andpossesses hydrogen bond acceptor complex points.Me6is often used as the transitionmetal introduced into the hydrophobic environment of the ligand.In mouse models, Me6can mobilize HSPCs effectively and rapidly, and increased the total cell number of bonemarrow at about2times after Me6administration. In meanwhile, Me6can also increasethe proportion and number of hematopoietic stem cells of the bone marrow. Me6can alsoincrease the total number of CFU of1.5times. Hemogram analysis showed an increase inperipheral WBCs, neutrophils and lymphocytes starting at12h after administration, thenumber of WBCs, neutrophils, lymphocytes of bone marrow significantly increased about2.6、2.3、3.5times respectively compared with0hour. Me6can also increased the ratio ofHSCs in peripheral blood and spleen at about2.0,3.6times, but did not induce apoptosisin HSCs. To study the effects of Me6on the human umbilical cord blood CD34+hematopoietic stem cell, we found that Me6can effective amplification of humanhematopoietic stem cells and promote the differentiation to megakaryocytes. In irradiatedmice models, Me6can significantly improve the survival rate of lethally irradiated mice,wherein the control group of mice all died, while there were still mice40%survival in theMe6group, at the same time also had a certain elevated survival rate of20%comparedwith the group of G-CSF. After radiation injury in mice，there was a sharp decline in thenumber of WBC, RBC and platelets in peripheral blood. However, the number ofabove-mentioned cells was significantly improved in Me6group at the fourth day afterradiation. At the14th day after the radiation, the cells began to restore and approached tothe normal level at the28th day. By HE staining of bone marrow biopsy, we found thatMe6could promote the repair of radiation injury to bone marrow, which could also beconfirmed by The statistical analysis of the results of the cell number of bone marrow. ofThe cell number of bone marrow in Me6group increased by about3times as comparedwith the control group. Me6could increase the ratio and the number of hematopoieticstem cell in bone marrow and spleen. Me6could also promote the proliferation of hematopoietic stem cell of bone marrow and peripheral blood by the combined with BrdUincorporation assay. The above studies demonstrated that the small molecule Me6canpromote the repair of radiation injury of the irradiated mice’s hematopoietic system.Exposed to radiation, intestinal is vulnerable to ionizing radiation and nuclearradiation injury. Radiation enteropathy is the result of complex interplay among manydose-and time-dependent pathophysiologic processes, including in?ammation, epithelialregeneration, tissue remodeling, and collagen deposition, as well as activation of thecoagulation system and, notably, endothelial dysfunction. Endothelial cells aremechanistically involved in early and delayed radiation responses in many normal tissues,including the intestine.In the study of Me6on irradiated mice intestinal repair, we found that Me6couldpromote the repair of radiation injury to the intestine, which well preserved the integrity ofthe mouse intestinal villi and crypts. The cell number of villus in Me6group is about6times more than that of control group. The length of the villi and crypts in Me6group issignificant longer, approximately2.9times and1.7times than that of the control grouprespectively. In order to better study the cell types involved in intestinal damage repair, weconstructed a GFP chimeric mouse model. We subcutaneous inject the mice with Me6after radiation, the bone marrow-derived cells increased significantly from donor, andmostly of these cell expressed the surface marker of CD105. The ratio of endothelialprogenitor cells (EPCs) in the Me6group was significantly more than that of controlgroup, approximately2.5times. We could found that Me6could aslo promote theproliferation of EPCs of bone marrow by the combined with BrdU incorporation assay.We then observed the proportion of EPCs in the peripheral blood, and found that the ratioof EPCs in Me6group is about3.5times more than that of in control in control group, butthese cells did not exhibit significant proliferation. However, Me6could not reduceintestinal injury of MMP9knockout mice after radiation. Through the above experiment,we could preliminary view that the possibility mechanism of the Me6that could reduceintestinal radiation injury is: Me6promoted the proliferation of EPCs in the bone marrow,and at the same time, the EPCs were mobilized to the peripheral blood by the MMP9 signaling pathway. Then the EPCs got to the radiation injury intestine via the bloodstream,in which the EPCs could proliferate and differentiate into endothelial cells (ECs). The ECscould reduce intestinal radiation injury by improve the microenvironment of the injuredintestine. |
PDF Full Text Request