| Into the middle of the 20 th century,especially since the 21 st century,the development and use of radioactive substances to human social and economic development is becoming more and more closely.The use of radioactive material benefited human beings as well as threaten human survival. Nuclear leckage, nuclear weapons and the side effects of radiotherapy in anytime threatened human’s health. Divided into two categories: acute and chronic radiation injury. Acute radiation injury syndrome was a kind of systemic diseases involved in the nervous system, hematopoietic system and digestive system, which the clinical symptoms of were diverse, causing the death of patients seriously. Hematopoietic system was sensitive to radiation, which inhibited the activity of hematopoietic stem cells, causing complete blood reduce and lots of complications such as bleeding and infection. When it acted on gut,Radiation caused intestinal epithelial cell necrosis and shedding, which triggered gastrointestinal bleeding, severe diarrhea, blood poisoning and bacteremia. Radiation damaged human body blood brain barrier and made the patient a convulsion, delirium and nervous system symptoms, which causing cerebral edema and meningitis. The skin is the largest organ surrounding the body.when the body suffered radiation, bear the brunt of the skin was damaged. It casued the rash inflammatory change,the superficial erosion and refractory ulcers when radiation affects the deep dermis and subcutaneous tissue. Because radioactive rays damaged tissues and organs function, and caused subcutaneous vascular occlusion.Clinically, the handling and treatment of radiation injury is often very difficult and complicated.MSCs was not hematopoietic stem cells which came from early mesoderm and ectoderm. It was long spindle, adherent growth, widely existed in the body’s bone marrow, umbilical cord and adipose tissue. MSCs expressed CD73, CD90 and CD105, not expressed hematopoietic system surface markers and HLA- DR, and can be induced differentiation in bone, cartilage tissue and adipose tissue in vitro. Currently, humans have built a stable and reliable way of separating and culturing MSCs from bone marrow and umbilical cord and MSCs had low immunogenicity, which laid a foundation for clinical infusion allogeneic MSCs to treat disease. A large number of domestic and foreign scholars have confirmed that MSCs had obvious therapeutic effect on GVHD, myocardial infarction, dementia and spinal cord injury.MSCs participated in the whole process of damage repair, it can promote the release of anti-inflammatory cytokines and inhibit the secretion of inflammatory cytokines to inhibit injury of inflammation, r at the same time it also promoted the secretion of a variety of growth factors and cytokine by paracrine especially VEGF, PDGF EGF and so on. MSCs secreted antimicrobial peptide LL-37 to reduce the growth of pathogenic bacteria. These roles were very conducive to wound repair. When MSCs as seed cells was applied to clinical its biological safety is our prio ity. Study after study has shown the clinical infusion MSCs was safe and reliable. This thesis preliminarily HUstudied MSCs promoted healing combined radiation-wound injury, which laid the foundation for clinical application.First of all,the skin of SD rats was prepared by 8% Na2 S,which was via abdomen anesthesia by 10% chloral hydrate(doses: 0.3 ml / 100 g). beta rays produced by linear accelerator(total exposure: 40 Gy) radiated rats right hip about 2.0X2.5 cm area, and then in which made a round wound about 1.5cm diameter with sterile surgical instruments, required deep into subcutaneous tissue layers.The rats were randomly divided into experimental group and control group.Then the human umbilical cord MSCs after recovery and serial subcultivation was injection into the experimental group rats(5.0 X106/ml/a rat)by tail vein, and the control group rats injected with the same amount of saline solution. The wound healing of experimental group and control group was observed respectively after the injection of 3 d, 5 d, 7 d, 11 d, 14 d, d, 28 d. The content of observation was wound size, presence of blood and osmotic liquid and the wound callus. The result: 1. The experimental group rats had no obvious ooze blood and inflammatory exudation decreased much significantly than control group after 7d injection MSCs. 2 To 28 d, the size of experimental group was much obviously shrinked than the control group and the wound was dry having no apparent ooze blood and inflammatory liquid. The above results show that the human umbilical cord MSCs have indeed promoted healing the combined radiation-wound injury in rats.Secondly, the combined radiation-wound injury rats model was made and randomly divided into 3 groups. Luciferase gene adenovious transfected umbilical cord MSCs and observed transfection effect. Three concentrations of human umbilical cord MSCs(5X106/ ml, 1.0 X107/ ml, 2.0 X107/ ml)was prepared,whichwas injectied respectively into three groups of rats via tail vein.fluorescein(dosage: 2 mg/100 g) was injected in rats by intraperitoneal. IVIS in vivo imaging system was used to observe the distribution of human umbilical cord MSCs in the rat model.The results: 1.By tail vein,human umbilical cord MSCs mainly gathered in the lung after injection of 2 h and lung fluorescence signal intensity gradually weakened after injection of 4h,at most which last 3 d 2.MSCs was gathered in injury location of rats which was injected 2.0X 107 cells By tail vein after injection of 2 h and the fluorescence signal intensity gradually increased. Fluorescent signal was not observed in rats, injury position which was injected 5X106 and 1.0X107 cells.The experimental results showed human umbilical cord MSCs promoted healing the combined radiation-wound injury by migration to the injury location.Finally, the bottom agar was prepared in 6 hole cell culture plate with agarose solution and DMEM medium containing 20% FBS,which was waiting to be solidified at room temperature.Then the top agar was made with hela suspension,human umbilical cord MSCs suspension and agarose solution. The final concentration of human umbilical cord MSCs was 250/hole(low dose group), 500/hole(middle dose group) and 1000/hole(high dose group). At the same time, blank control group and positive control group of hela was set up,of which the concentration was 500/hole.6 hole cell culture plate was placed in 37 ℃ and 5% CO2 incubator.The colony formation of human umbilical cord MSCs and hela was continuously observed with microscope. The result: 1. When the hela was cultured to 7 d,the colony formation was observed obviously with microscope.As time went by, the volume and amount of colony gradually increased.When the hela was cultured to 14 d, the colony was observed with our own eyes.2. The colony was not observed in experimental group of human umbilical cord MSCs and blank control group The results showed that the human umbilical cord MSCs in vitro had not tumorigenicity in a short time, which had certain security.To sum up, this study confirmed that the human umbilical cord MSCs promoted healing combined radiation-wound injury by migration to the radiation injury position and it had no tumorigenicity in vitro in the short period, which laid the foundation of treating radiation injury clinically,especially refractory skin ulcer caused by radiation. |
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