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The management of drainage tube is an important part of nursing work. Patient restraint and tube fixation cannot effectively prevent unplanned extubation (UEX) when the tube is accidentally pulled by violence. The nursing innovation team of Henan Provincial People's Hospital designed a medical drainage tube anti-pull device in order to change the existing technology of preventing drainage tube disconnecting by means of restraint and fixation, and to interfere with the basic cause of drainage tube disconnection, and obtained the national utility model patent (patent number: ZL 2020 2 2843025.1). The design of sleeve and clasp is that when the drainage tube is pulled by accidental violence, the friction fastener clamps the drainage tube mechanically to achieve the purpose of braking the drainage tube and prevent the drainage tube from coming out. Card sleeve ring fracture design can be applied to drainage tubes of different diameters, and the buzzer device at the instant of the snap ring into the card set warning medical staff to the occurrence of risk events, so that the nurse can come in the first place for effective treatment, which is a fuse for surgical drainage tubes and is to timely and effectively prevent UEX.
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The aim of this study was to develop a semi-quantitative immunochromatographic method for rapid detection of Newcastle disease virus (NDV) antibodies by expressing HN protein in rice endosperm bioreactor. The recombinant plasmid pUC57-HN was digested by MlyⅠ and XhoⅠ to retrieve the HN gene, while the intermediate vector pMP3 containing promoter, signal peptide and terminator was digested by NaeⅠ and XhoⅠ. The HN gene and the linearized pMP3 were purified and ligated to form a recombinant plasmid pMP3-HN1. Subsequently, pMP3-HN1 and plant vector pCAMBIA1300 were digested by EcoRⅠ and Hind Ⅲ, and the HN1 gene was cloned into pCAMBIA1300. The recombinant plasmid pCAMBIA1300-HN1 was introduced into Agrobacterium tumefaciens EHA105 by electrotransformation, and the pCAMBIA1300-HN1 was transferred into rice callus by agrobacterium-mediated method. After dark culture, callus screening, differentiation, rooting and transplanting, transgenic rice seeds were obtained 4 months later. PCR identified that the HN gene has been inserted into the rice genome. SDS-PAGE and Western blotting indicated that the HN protein was successfully expressed in the positive rice endosperm. The purity of the HN protein was more than 90% by SP cation exchange chromatography and gel filtration chromatography. According to the national standards for the diagnostic techniques of Newcastle disease HI test (HI≥4log2, positive antibody reaction), a colloidal gold labeled purified HN protein was used to prepare a semi-quantitative test strip by double-antibody sandwich method for rapid detection of NDV antibody. The results showed that the test strip did not cross-react with positive sera against other viruses, and the sensitivity of the test strip reached 1:102 400 for standard positive sera of Newcastle disease. Testing of a total of 308 clinical sera showed that the compliance rate of the test strip with HI test was 97.08%, and the Kappa value was 0.942. In conclusion, high purity recombinant HN protein was obtained from rice endosperm, and a simple, rapid, highly sensitive and highly specific semi-quantitative immunochromatographic strip was developed. The test strip could be used for immune evaluation of the Newcastle disease vaccine.
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Animals , Antibodies, Viral , Chickens , HN Protein/metabolism , Newcastle Disease/prevention & control , Newcastle disease virus/metabolism , Oryza/geneticsABSTRACT
Abstract@#The growth and development of children is related to the future of the country and the nation. In recent years, there have been more and more cohort studies in the field of children s mental health. Bymainly introducing the advantages of cohort studies on children in distress and organizes domestic and foreign cohort studies in the field of mental health of children in distress, this article finds that it is mostly used in depression, anxiety, post traumatic stress disorder, suicidal ideation and attempts, etc, and mainly explores the risk and prevalence of mental health development in children in distress, and identifies long term negative damage. The research aims to promote the long term development and high quality development of such research by analyzing and summarizing the status quo and prospects of cohort research in the field of mental health of children in distress.
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Objective:To probe into the causes, reconstructive strategies, and repair outcomes of asymmetric eyelid configuration after blepharoplasty.Methods:All 73 patients (14 males and 59 females) with asymmetric double eyelid after blepharoplasty were recruited between July 2013 and June 2018 from Department of Plastic and Burns Surgery, West China Hospital, Sichuan University. The patients aged from 18 years to 42 years with the median age of 27 years. The new double eyelid line was designed pre-operation. Releasing subcutaneous adhesion of upper eyelid entirely, trimming inferior orbicularis oculi, adjusting and comparing the attachment position of bilateral levator aponeurosis were performed during surgery. Patients and surgeons marked the appearance of double eyelid both before and after repair operation, results of which were analyzed by t-test.Results:All 73 patients obtained improved double eyelid with primary healing. During follow-up from 8 to 12 months, repaired double eyelid showed satisfactory configuration with smooth natural double eyelid line and symmetric bilateral double eyelid. Of the 73 patients, 3 (4.1%) complaint rough double eyelid line, for whom re-fixation through small incision were adopted and no complication was observed during follow-up time. Scores by patients and surgeons were both significantly better after surgery.Conclusions:Analyzing the causes of asymmetric eyelid after double-eyelid blepharoplasty and repairing it contribute to aesthetic pleasing reconstructed double eyelid.
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Objective: To investigate the expression of vascular endothelial cell growth factor (VEGF) in the immediate replantation of pulp healing in the rats, and to clarify the effect and its mechanism of erythropoietin (EPO) on immediate pulp reconstruction in the rats. Methods: Eighty 4-week-old healthy male Wistar rats were randomly divided into non-tooth extraction group, negative control (normal saline) group, positive control (gentamicin) group and EPO group; there were twenty rats in each group. The teeth in each group were immersed in its corresponding solution for 4 min before replantation. Four rats were killed on the days 3, 7, 14, 21 and 28, respectively, and the specimens were made in the operation area. HE staining was used to observe the pulp revascularization in different time periods. Immunohistochemical staining was used to observe the protein expression levels of VEGF in odontal tissue of the rats in each group in different time periods. Results: The HE staining results showed that compared with non-tooth extraction group, the pulp tissue of replanted teeth of the rats in normal saline group had more inflammation, less root development, less restorative dentin and cementum deposition, and wider apical pores; the inflammation of pulp tissue of replanted teeth of the rats in gentamicin group and EPO group was mild, and the root development was relatively good; there were more deposits of restorative dentin and cementum, and the apical pores were narrowed. The immunohistochemical results showed that compared with non-tooth extraction group, the positive expressions of VEGF in odontal tissue of the rats at the days 3, 7 and 14 in the other groups were strong. Afterwards, the positive expression levels of VEGF were decreased gradually with the prolongation of time. The average optical density (AOD) of VEGF positive area indicated that EPO group > gentamycin group > normal saline group > non-tooth extraction group. Compared with non-tooth extraction group, the protein expression levels of VEGF in odontal tissue of the rats in normal saline group, gentamicin group and EPO group at 3, 7, 14 and 21 d after operation were significanty increased (P0.05). Compared with normal saline group, the protein expression levels of VEGF in odontal tissue of the rats in gentamicin group and EPO group at 3, 7, 14 and 21 d after operation were significantly increased (P0.05). Compared with gentamicin group, the protein expression levels of VEGF in odontal tissue of the rats in EPO group at every time points had no significant differences (P>0.05). Conclusion: EPO can increase the expression of VEGF, induce angiogenesis in pulp tissue, and provide the rich vascular bed for replantated teeth, so as to exert the potential of dental pulp defense and repair, and promote the healing of replanted teeth.
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Objective To study the cell morphology and differentiation efficiency when rabbit bone marrow mesenchymal stem cells (BMSCs) were induced osteogenic differentiation as culturing by autologous platelet-rich plasma (PRP) instead of serum,and to explore a new method of inducing BMSCs osteogenic differentiation.Methods The PRP was prepared by arterial blood of rabbit.Punctured and The bone marrow was sampled from rabbit's iliac bone,and BMSCs were collected,which divided into PRP group,fetal calf serum (FBS) group and serum-free control group,and cultured in 10％ autologous PRP,10％ FBS and serum-free respectively,combined with DMEM-F12 medium.The second generation cells were divided into experimental and control groups.The experimental groups' medium was added dexamethasone,β-sodium glycerophosphate and ascorbic acid,and the control groups went on.The cell morphological difference of each group was Observed between anterior and after inducing differentiation,and compared between each group.Results BMSCs of PRP and FBS groups grew quickly,presented like fusiform form before induction,and increasd in volume,became a triangle,polygonal and round form gradually after osteogenic induction.Cells of PRP and FBS groups aggregated spontaneously and multilayered,and formed calcium nodules and bone-like structure after induced 7 days averagely,which could be stained red by alizarin red S;cells of serum-free groups were induced 14 days averagely,only three samples showed osteogenesis performance.Cells of PRP and FBS groups differentiation efficiency was superior to serum-free groups when inducd 20 days,the difference was statistically significant (P＜0.05),and the difference between efficiency of PRP and FBS groups was not significant (P＞0.05).Conclusions Autologous PRP could be used to proliferate and induce osteogenic differentiation of BMSCs instead of serum.
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Objective:To report one case of bilateral maxillary fourth molars,and to explore the occurrence and treatment method of additional fourth molar.Methods:The case materials of one patient with bilateral maxillary four molars were collected,and the information was record.Combined with reviewing the relevant literatures,the clinical data,complication and treatment of one patient with bilateral maxillary fourth molars were retrospectively analyzed.Results:The patient was diagnosed due to left maxillary posterior teeth area intermittent pain for several months and the increased pain for 5 d;at the same time,there was no abnormal appearance inducing the symptoms detected by speciality check-up.The pantomography showed the four molars in bilateral maxillary,and the fourth molars impacted the third molar root's absorption.The fourth molar crown was smaller and the root was shorter. The 18,28,29 teeth were removed,and the left maxillary posterior teeth area intermittent pain disppeared. Conclusion:The occurrence of additional fourth molar is a relatively rare demal dysplasia.However,there are some complications following with the fourth molar.Early diagnosis and treatment are essential.
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Objective:To repaire the posterior tooth defects in the elderly people with CEREC chairside CAD/CAM restorations,and to evaluate the clinical curative effect 1 year after treatment.Methods:A total of 52 posterior teeth from 48 aged patients were selected.Among them,24 cases were onlays,18 cases were inlays,and 10 cases were crowns.The inlays/onlays/crowns were designed and manufactured by CEREC chairside CAD/CAM system,and then bonded with resin cement.After 12 months,the patients were followed up.The teeth with restorations were assessed by using the modified USPHS criteria including restorations,tooth,and periodontal.At the same time,the patient's satisfaction was evaluated.Results:In all of the 52 restorations,the success rates of inlay,onlay and crown were 94.4％,91.7％ and 80.0％,respectively;among them,one inlay was fractured,one onlay did not close the edge,one onlay's edge integrity was mild defect,one teeth was fractured,one crown's adjacent relationship was not close,and one crown appeared papillitis.Conclusion:The restorations manufactured by CEREC chairside CAD/ CAM system have a good short-term clinical curative effect.It is an effective way to repair the posterior tooth defects in the elderly people.
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Objective To observe the clinical efficacy of propranolol and 595 nm pulsed dye la ser (PDL) in treatment of infantile hemangioma.Methods 26 infants admitted to our hospital from January 2013 to January 2015 with hemangioma underwent oral propranolol 2 mg/(kg · d) treatment after excluding of taboos.The daily doses were divided equally to two parts,taken on the time of 8:00 and 20:00,when the electrocardiograph and pulse oxygen were monitored and recorded persistently.The patients were discharged from the hospital when it was stable,with review of blood routine examination,fasting blood glucose,liver and kidney function,and the change of size,character and color of hemangioma were recorded,and taken photos every two weeks after discharge.The 595 nm PDL was used to treat the hemangioma faded incompletely when the propranolol was terminated.Results The tension and color of all hemangioma decreased in varying degrees in taking propranolol for 72 hours,and evaluated the efficacy as recovery completely 19 cases;signifivantly effective in 3 cases and partial efficacy in 4 cases;the latter 7 cases were further treated with 595 nm PDL.Followed-up for 6-12 months showed that efficacy of recovery reached 100％.10 cases showed heart rate was mild reversibly slow,with no special treatment.5 cases had diarrhea,and healed with symptomatic treatment.No adverse reactions like liver and kidney dysfunction and so on were found.Conclusions Propranolol and 595 nm PDL can effectively treat infantile hemangioma,and thus it can be used as the recommended treatment of infantile hemangioma.
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Objective To explore the operating methods and its related questions of auricular reconstruction with totally expanded skin in combination with laser hair removal for the treatment of adolescent microtia.Methods From Jan.2013 to Dec.2016,30 adolescent microtia patients were treated with totally expanded skin.At the first stage,the 100 ml kidney-shaped expander was implanted under the skin of mastoid.After expanding capacity of 80 ml,the hair on the expanded skin was depilated once a month with reference to the healthy ear;at the second stage,after expanding capacity of 150 ml,the expander was taken out and the fiber capsule was removed;the tautologous rib cartilage was harvested and the scaffolds were sculptured;the cartilage was implanted and the expanded skin flap was used to cover the frontal surface and back surface of the scaffold;at the third stage,the earlobe transposition,conchal excavation and tragus construction were performed at the same time.Results All the patients were followed up for 3 to 24 months;the results showed 1 case of leakage of expander,4 cases of hematoma,2 case of expanded skin burst,and the complications were treated correctly,all patients were satisfied with the appearance;the color,texture,location,size;and height of ear cranial angle were matched with health ear;there was no obvious scar and auricle subunit structure was clear.Conclusions The laser in combination with the large capacity tissue expander in auricular reconstruction is simple,less trauma and less scarring.
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Objective To compare the efficacy of ultrasound-guided lumbar epidural access using paramedian transverse scanning (PMTS) versus paramedian saggital scanning (PMSS) with the needle inplane.Methods Fifty American Society of Anesthesiologists physical status Ⅰ-Ⅲ patients,aged 50-75 yr,weighing 55-85 kg,undergoing lower extremity surgery under combined spinal-epidural anesthesia,were divided into PMSS group (n=25) and PMTS group (n=25) using a random number table.The realtime ultrasound-guided lumbar epidural access (L3,4) was performed using PMTS and PMSS in PMTS and PMSS groups,respectively.The visibility of ligamentum flavum,posterior and anterior dura maters,posterior epidural space on the prepuncture ultrasound images,imaging quality score,time for puncture and depth of puncture were recorded.The development of air ultrasonic contrast sign and backflow of cerebrospinal fluid from the spinal needle were recorded.The development of adverse reactions such as paresthesia and hypokinesia was also recorded on 2 days after operation.Results Compared with group PMSS,the time for puncture was significantly shortened,the depth of puncture was shallower (P＜0.05),and no significant change was found in the visibility of spinal structure,imaging quality score or air ultrasonic contrast sign and incidence of backflow of cerebrospinal fluid in group PMTS (P＞0.05).No significant change was found in adverse reactions such as paresthesia or hypokinesia between the two groups (P＞0.05).Conclusion PMTS provides clear imaging and simple and convenient operation in guiding lumbar epidural access with the needle in-plane when compared with PMSS,and it is worthy of clinical application.
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Objective To evaluate the changes in the expression of neuroligin1 in excitatory postsynaptic membrane of the spinal dorsal horn in a rat model of inc isional pain.Methods Forty-eight pathogen-free healthy adult male Sprague-Dawley rats,aged 6-8 weeks,weighing 280-320 g,were divided into control group (group C,n =12) and incisional pain group (group Ⅰ,n=36) using a random number table.A1 cm long incision was made in the plantar surface of the right hindpaw in group Ⅰ.Cumulative pain score (CPS) was assessed and mechanical paw withdrawal threshold to von Frey stimuli was measured at 3 hand 1 and 3 days after operation (T1,2,3).The animals were then sacrificed and their lumbar segments (L3-6) of the spinal cord were removed for detection of the expression of neuroligin1,postsynaptic density-95 protein (PSD-95),glutamate receptor 1 (GluR1) and GluR2 in the postsynaptic membrane of spinal dorsal horn (by Western blot) and co-expression of neuroligin1 with PSD-95 in spinal dorsal horn (by co-immuno-precipitation).Results Compared with group C,CPS was significantly increased and mechanical paw withdrawal threshold was decreased at T1-3,and the expression of neuroligin1 and GluR1 in the postsynaptic membrane of spinal dorsal horn at T1,2 and co-expression of neuroligin1 with PSD-95 at T1 were up-regulated in group Ⅰ (P＜0.01 or 0.05).Conclusion The development and maintenance of incisional pain is related to the signaling pathway regulated by neuroligin1 in excitatory postsynaptic membrane of the spinal dorsal horn of rats.
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Objective To evaluate the effect of pretreatment with botulinum toxin A injected intrath?ecally or locally at the incision site on the neurokinin?1 ( NK?1) receptor internalization in the spinal dorsal horn in a rat model of incisional pain. Methods Male Sprague?Dawley rats, weighing 280-300 g, aged 6-8 weeks, were used in the study. The experiment was performed in two parts. ExperimentⅠ Twenty?seven rats with no sign of nerve injury at day 7 after successful catheterization were selected and divided into 3 groups (n=9 each) using a random number table: control group (C1 group), incisional pain group (IP1 group) and intrathecal botulinum toxin A group (BoNT∕A1 group). At 24 h before operation, botulinum tox?in A 0.5 U ( in 10μl of normal saline) was injected intrathecally in group BoNT∕A1, and normal saline 10μl was injected intrathecally in group IP1. ExperimentⅡ Twenty?seven rats were selected and divided into 3 groups (n=9 each) using a random number table: control group (group C2), incisional pain group (IP2 group) and locally injected botulinum toxin A at the incision site group (BoNT∕A2 group). At 24 h before op?eration, botulinum toxin A 2 U ( in 0.4 ml of normal saline) was injected subcutaneously at the incision site and into the plantar surface, and normal saline 0.4 ml was injected subcutaneously at the incision site and into the plantar surface in group IP2. Six rats in each group were selected, and the cumulative pain score (CPS) was recorded, and the mechanical paw withdrawal threshold ( MWT) in the right hindpaw was measured be?fore administration, before operation, and at 3 h and 1, 3, 5 and 7 days after operation. At 3 h after opera?tion, 3 rats in each group were selected and sacrificed, and the lumbar segment ( L4,5 ) of the spinal cord was removed for determination of the expression of NK?1 receptors in the spinal dorsal horn by immunofluores?cence. Results ExperimentⅠ Compared with group C1, the CPS was significantly increased at 3 h and 1, 3, 5 and 7 days after operation, the MWT was significantly decreased at 3 h and 1 and 3 days after opera?tion, and the expression of NK?1 receptors in the spinal dorsal horn was significantly up?regulated in group IP1, and the CPS was significantly increased at 3 h and 1, 3 and 5 days after operation, the MWT was sig?nificantly decreased at 3 h after operation ( P0.05). Compared with group IP1, the CPS was significantly decreased, and the MWT was significantly increased at 3 h and 1, 3, and 5 days after oper?ation, and the expression of NK?1 receptors in the spinal dorsal horn was significantly down?regulated in group BoNT∕A1 (P0.05) . Compared with group IP2, the CPS was significantly decreased at 3 h and 1, 3, and 5 days after operation, the MWT was signifi?cantly increased at 3 h and 1 and 3 days after operation, and the expression of NK?1 receptors in the spinal dorsal horn was significantly down?regulated in group BoNT∕A2 (P<0.05). Conclusion Pretreatment with botulinum toxin A injected intrathecally or locally at the incision site can inhibit the internalization of NK?1 re?ceptors in the spinal dorsal horn in a rat model of incisional pain.
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Objective To evaluate the clinical efficacy of “Shamrock” ultrasound images?guided lumbar sympathetic ganglion blockade ( LSGB) . Methods Sixty patients of both sexes, aged 18-60 yr, weighing 50-70 kg, of American Society of Anesthesiologists physical statusⅠorⅡ, undergoing unilater?al LSGB, were divided into groupⅠ ( n=30) and group Ⅱ ( n=30) using a random number table. In group Ⅰ, unilateral LSGB was performed at the L2 level under ultrasound guidance with paramedian trans?verse scanning. In groupⅡ, unilateral LSGB was performed at the L2 level under ultrasound guidance with“Shamrock” ultrasound images. After final needle position was confirmed, 2% lidocaine 6?0 ml was ad?ministered in each patient. At 20 min before and after LSGB, the visual analogue scale scores and skin temperature of the big toe of the affected foot were recorded, and the successful blockade and visibility of important paravertebral structures on ultrasound images were recorded during puncture. Results The visu?al analogue scale scores were significantly lower, and the skin temperature on the affected side was signifi?cantly higher after LSGB than before LSGB in both groups ( P<0?05) . The important paravertebral struc?tures such as erector spinae, quadratus lumborum, psoas major, transverse process of L2 vertebrae, and the curved edge of L2 vertebrae were visible in both groups. The visibility rate of the inferior vena cava or ab?dominal aorta on ultrasound images and the success rate of blockade were significantly lower in group Ⅰthan in group Ⅱ (P<0?01). Conclusion Compared with paramedian transverse scanning, LSGB has some advantages such as real?time monitoring, higher success rate of blockade, better efficacy and avoiding damage to great vessels when performed under “Shamrock” ultrasound image guidance.
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BACKGROUND:Early vascularization is crucial for wound healing. A high-porosity, macrovoid alogeneic skin leads to the rapid vascularization and celular infiltration. OBJECTIVE: To obtain a new alogeneic skin product with high porosity, good cel permeability and good histocompatibility using an improved preparation method of human acelular dermal matrix. METHODS: Cel components of healthy human skins were removed by the improved method and the traditional method, respectively. The improved method was to remove the subcutaneous fat, eliminate the epidermis (1 mol/L NaCl solution at 37℃ for 24 hours) folowed by shaking processing (2% NaOH at 45℃ for 4 hours), and then, the solution was neutralized with PBS rinsing, dried and stored at 4℃ for standby. We detected the porosity and degradation time in vitroof the acelular dermal matrices prepared by two methods and the cytotoxicity of the material infiltration liquid on the adipose-derived stem cels. Hematoxylin-eosin staining was used for the detection of the cel residual, the integrity of colagen and cel biocompatibility. Scanning electron microscopy was used to detect the pore size. RESULTS AND CONCLUSION: Both the two methods could completely remove the cel components, and maintain the integrity of the colagen scaffold. The porosity of acelular dermal matrix with the improved method was (93.22±0.99)%, which was significantly higher than that with the traditional method [(74.28±2.06)%;P 0.05). No obvious cytotoxicity of the acelular dermal matrix prepared with the improved method was detected. At 3-7 days of co-culture, the adipose-derived stem cels cultured on the acelular dermal matrix prepared with the improved method could penetrate the basement membrane to the deep dermis, while there was no obvious cel invasion and growth in the deep dermis prepared by the traditional method. Compared with the traditional method, the improved method is more suitable for cel infiltration and growth with higher porosity and larger pore size.
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Peripheral arterial disease (PAD) shows increasing morbidity and mortality. Clinical manifestations of PAD, such as intermittent claudication, rest pain and nonhealing ulcer, contribute to impaired quality of life, and ischemic stroke caused by PAD can be life-threatening. Unfortunately, PAD patients often receive suboptimal treatment, and pathogenesis of the disease is still unclear. Over the past decade, the evolving technology and interdisciplinary collaboration have enabled improvement of diagnosis and treatment for PAD. This review makes a brief summary of the current status and progress in genetics research on PAD, which included candidate gene studies, linkage analyses, genome-wide association studies, and applications and development prospects of epigenetics, mitochondrial DNA and other new technologies.
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Animals , Humans , DNA, Mitochondrial , Genetics , Genetic Predisposition to Disease , Genetics , Genome-Wide Association Study , Methods , Genotype , MicroRNAs , Genetics , Peripheral Arterial Disease , Genetics , Therapeutics , Polymorphism, Single Nucleotide , Risk FactorsABSTRACT
Objective To explore the effects of recombinant plasmids of pGPU6/GFP/NeoshRNA-CTGF (shRNA-CTGF) on the type Ⅰ collagen (COL-Ⅰ) protein expression in keloid,through RNA interference on connective tissue growth factor (CTGF) in vivo and in vitro.Methods Recombinant plasmids were designed and constructed by specific shRNA-CTGF; After transfeced human keloid fibroblast with shRNA-CTGF in vitro,RT-PCR was used to detect the CTGF mRNA level,and Western blot to detect the secretion of COL-Ⅰ.After transfected the keloid of nude mice with shRNA-CTGF in vivo,RT-PCR was used to detect the CTGF and COL-Ⅰ mRNA level,and Western blot was used to detect the protein expression of COL-Ⅰ.Results Recombinant plasmids of CTGF were successfully constructed,and the CTGF gene expression was significantly decreased in vivo and in vitro by 86.8％ and 54.1 ％,respectively; Down-regulation of CTGF in vitro significantly inhibited the mRNA and protein level of COL-Ⅰ by 76.8％ and 65.6％,respectively; Down-regulation of CTGF in vivo significantly reduced the COL-Ⅰ mRNA and protein level by 52.7％ and 48.0％,respectively.Conclusions CTGF gene expression is successfully down-regulated by the recombinant plasmid of shRNA-CTGF in vivo and in vitro.shRNA-CTGF significantly reduces the COL-Ⅰ protein level in keloid.It implies that CTGF gene is a potential target in the therapy of pathological scar.
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BACKGROUND:A great amount of mesenchymal stem cels can be successfuly derived from fat tissue and induced to differentiate into osteoblasts, chondrocytes, adipocytes and myocardial cels. OBJECTIVE:To establish the method of isolating, culturing and osteogenic differentiation of adipose-derived stem celsin vitro, and to explore the potential of adipose-derived stem cels as seed cels for bone tissue engineering. METHODS:Colagenase enzymatic digestion was used to isolate adipose-derived stem cels from human fat tissue which were then culturedin vitro. Flow cytometry was used to detect cellsurface markers. cellcounting kit-8 assay was performed to examine cellviability. Adipose-derived stem cels were induced by osteogenic induced reagent to differentiate into bone cels. In addition, we also performed BCIP/NBT method to detect alkaline phosphatase activity. Alizarin red staining was used to detect the formation of calcium node. RT-PCR was performed to examine alkaline phosphatase and osteopontin expression. RESULTS AND CONCLUSION:We successfuly obtained adipose-derived stem cels from fat aspirated liquid. Adipose-derived stem cels obtained could passage stably and proliferate highly. Flow cytometry data showed the expression of stem cellsurface markers. Adipose-derived stem cels showed typical osteoblast morphology after osteogenic differentiation. Alkaline phosphatase staining was positive and alizarin red staining displayed the formation of calcium node. Furthermore, we found that alkaline phosphatase and osteopontin mRNA was expressed after differentiation 0, 3, 7, 14, 21, 28 days. These findings indicate that adipose-derived stem cels can be obtained from fat tissue through enzymatic digestion, differentiate towards bone cels, and express alkaline phosphatase and osteopontin, which can become potential seed cels for bone tissue engineering.
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Objective To explore the effect of epidermal growth factor (EGF) on tube formation of HUVEC induced by the secretion of angiogenesis factors of adipose-derived stem cells (ADSCs).Methods ADSCs were primarily cultured by enzyme digestion method.The flow cytomertry was performed to detect the expression of cell surface marker.ELISA was used to detect the expression of VEGF,HGF,and SDF-1 after given different doses of EGF.Tube formation assay was used to examine the effect of EGF on the tube formation induced by ADSCs.Results ADSCs were successfully isolated and cultured from human liposuction tissue and specific markers were expressed on ADSCs.EGF promoted the secretion of angiogenesis factors VEGF,HGF,and SDF-1,which were secreted by ADSCs.EGF pretreatment increased the ability of tube formation of HUVECs induced by ADSCs.Conclusions ADSCs induce the secretion of angiogenesis factors in vitro,and thus increase the ability of tube formation of HUVECs.EGF promotes the secretion ability of ADSCs,and the best concentration is 15 mg/L.
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Objective To investigate the specific silencing of connective tissue growth factor (CTGF) in a nude mouse keloid model,using RNA interference (RNAi) technique,and to provide the basis for gene therapy of keloid.Methods The nude mouse keloid model was established,and then transfected in vivo with well-amplifiating plasmid.The mRNA expression levels of CTGF mRNA and type Ⅰ collagen mRNA were detected with reverse transcription-polymerase chain reaction (RT-PCR).The distribution and protein expression levels of CTGF and type Ⅰ collagen were determined quantitatively using immunohistochemistry.Results The expression of CTGF at mRNA and protein levels was decreased in the experiment group,and the expression of type Ⅰ collagen at mRNA and protein levels was also decreased after transfection,as compared with negative control group and blank group,with significant difference between groups (P＜0.05).Moreover,the expression of type Ⅰ collagen and CTGF was positively correlated (r=0.979).Conclusions Keloid type Ⅰ collagen can be decreased through in vivo inhibiting CTGF expression.The transfection of CTGF gene in vivo may have effects on type Ⅰ collagen generation,and thus inhibit the keloid growth.