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Mesenchymal stem cell (MSC)-derived exosomes (Exos) were reported to a prospective candidate in accelerating diabetic wound healing due to their pro-angiogenic effect. MSCs pretreated with chemistry or biology factors were reported to advance the biological activities of MSC-derived exosomes. Hence, this study was designed to explore whether exosomes derived from human umbilical cord MSCs (hucMSCs) preconditioned with Nocardia rubra cell wall skeleton (Nr-CWS) exhibited superior proangiogenic effect on diabetic wound repair and its underlying molecular mechanisms. The results showed that Nr-CWS-Exos facilitated the proliferation, migration and tube formation of endothelial cells in vitro. In vivo, Nr-CWS-Exos exerted great effect on advancing wound healing by facilitating the angiogenesis of wound tissues compared with Exos. Furthermore, the expression of circIARS1 increased after HUVECs were treated with Nr-CWS-Exos. CircIARS1 promoted the pro-angiogenic effects of Nr-CWS-Exos on endothelial cellsvia the miR-4782-5p/VEGFA axis. Taken together, those data reveal that exosomes derived from Nr-CWS-pretreated MSCs might serve as an underlying strategy for diabetic wound treatment through advancing the biological function of endothelial cells via the circIARS1/miR-4782-5p/VEGFA axis.
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Humans , Endothelial Cells/metabolism , Exosomes/metabolism , Cell Wall Skeleton/metabolism , Neovascularization, Physiologic , Wound Healing/physiology , MicroRNAs/metabolism , Diabetes Mellitus , Vascular Endothelial Growth Factor A/metabolismABSTRACT
Objective:To evaluate the effects of adipose-derived stem cells (ADSCs) and ADM microparticle on diabetic wound healing.Methods:ADSCs was co-cultured with ADM microparticle in vitro. The models of diabetic nude mice were established by intraperitoneal injection of STZ and the full-thickness skin defects were designed on the back. All 24 diabetic mice were randomly divided into 4 group: experimental groups were transplanted with ADSCs and ADM microparticle and the other groups were transplanted with ADSCs, ADM microparticle and blank control group was set up. On the 7th and 14 th days, the wound healing rate of 3 mice randomly selected from each group was calculated, and the thickness between dermis and epidermis was measured by hematoxylin and eosin staining. The density of neovascularization was measured by immunohistochemical staining. The differences were compared between the groups.Results:Compared to the ADSCs groups, the mice of the experimental groups showed higher cell survival rate. The wound healing rate in the experimental groups was (86.0±2.7)% (7 days) and (98.5±1.1)% (14 days), thicker dermis-epidermis distance was (99.1±1.8) μm (7 days) and (124.3±4.3) μm (14 days) ( P<0.05), and higher density of neovascularization was noted. Conclusions:The transplantation with active ADM microparticle can significantly promote neovascularization and wound healing of diabetic wound.
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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 observe content of cytokine in human stromal vascular fraction gel (SVF-GEL) and effect of SVF-GEL on biological behaviors of epidermal and dermal cells in vitro and clinical efficacy of SVF-GEL.@*Methods@#(1) SVF-GEL was prepared using liposuction aspirates harvested from females who received abdomen liposuction in author′s unit. SVF-GEL (1 mL) and high-glucose Dulbecco′s modified eagle medium (DMEM, 1 mL) were respectively cultured for 24 h with high-glucose DMEM containing 10% fetal calf serum, 10 g/L penicillin, and 10 g/L streptomycin, denoted as SVF-GEL group and negative control group, with 6 samples in each group. Content of epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) in the supernatant was determined by enzyme-linked immunosorbent assay. (2) A number of 5×105 human skin fibroblasts (HSF) and HaCaT cells in logarithmic phase were inoculated and cultured in Transwell chambers for 12 h. All Transwell chambers containing cells were divided into SVF-GEL group (0.5 mL SVF-GEL was added for co-culture) and control group (0.5 mL high-glucose DMEM was added for co-culture), with 9 samples in each group for HSF and HaCaT cells. Scratch assay was performed after culture for 24 h, and residual scratch width was observed at post scratch hour (PSH) 0 (immediately), 24, and 48. Cell migration distance was measured at PSH 24 and 48. After culture for 24, 48, and 72 h, the number of living cell was counted using cell counter. (3) From June 2018 to June 2019, SVF-GEL was applied clinically to treat 15 patients with depressed scars on face, including 2 males and 13 females, aged 19 to 42 years. Survival condition of SVF-GEL and whether complications or not were observed 6 months after surgery. Before surgery and 6 months after surgery, depressed degree, color, and pliability of scar were compared. Vancouver Scar Scale (VSS) was employed to access color, vascularity, and pliability before surgery and 6 months after surgery, and total score was calculated. The number of patients with complete satisfaction or satisfaction was counted six months after surgery. Data were processed with analysis of variance of factorial design, paired samples t test, and Wilcoxon rank sum test.@*Results@#(1) The content of EGF in SVF-GEL group and negative control group was (316.6±12.8) and (3.4±0.6) pg/mL, and the content of VEGF in SVF-GEL group and negative control group was (568.67±12.19) and (4.93±0.16) pg/mL, with statistically significant differences between the two groups (t=48.777, 92.485, P<0.01). (2) Residual scratch widths of HSF and HaCaT in SVF-GEL group and control group were decreased gradually along with time elapse, in which those in SVF-GEL group at PSH 24 and 48 were less than those in control group. At PSH 24 and 48, cell migration distances of HSF and HaCaT in SVF-GEL group were more than those in control group (tHSF=-20.304, -43.516, tHaCaT=-15.060, -8.684, P<0.01). After culture for 24, 48, and 72 h, the number of living cell of HSF and HaCaT in SVF-GEL group was significantly more than that in control group (tHSF=-3.374, -6.809, -18.036, tHaCaT=-4.793, -6.028, -8.141, P<0.05 or P<0.01). (3) Six months after surgery, SVF-GEL grafted into patients survived well without complications, and depressed degree of scar ameliorated obviously with lightened pigmentation and softer texture as compared with before surgery. Compared with those before surgery, VSS scores of color, vascularity, and pliability, and total score of 15 patients with depressed scars on face were obviously decreased 6 months after surgery (Z=-2.06, -2.07, -2.07, t=-15.811, P<0.05 or P<0.01). One patient was satisfied with the clinical outcome, and the rest 14 patients were completely satisfied with the clinical outcomes.@*Conclusions@#SVF-GEL contains cytokines EGF and VEGF, which can enhance cell migration ability and proliferation ability of HSF and HaCaT cells and have obvious effects on depressed scars for clinical application.
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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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BACKGROUND: Many studies have shown that cell-assisted lipotransfer promotes the survival of fat implants, but there are few studies exploring stromal vascular fraction (SVF) -assisted fat transplantation for improving facial skin quality. OBJECTIVE: To evaluate the clinical effect of autologous adipose-derived SVF-assisted autologous fat transplantation in the improvement of facial skin quality. METHODS: From September 2016 to June 2018, in the Plastic and Cosmetic Center, the Affiliated Hospital of Xuzhou Medical University, 20 patients were recruited and then randomly divided into experimental group and control group, namely SVF-assisted fat transplantation and simple fat transplantation, respectively. A VISIA skin detector was used to detect facial skin quality of patients preoperatively and 6 months postoperatively, including spots, wrinkles, texture, pores, ultraviolet spots, brown spots, couperose skin and porphyrin. RESULTS AND CONCLUSION: The effect of improvement in wrinkles and texture in the experimental group was better than that of the control group, and the difference was statistically significant (P < 0.05). In terms of spots, pores and couperose skin, the therapeutic effect in the experimental group was better than that in the control group, but the difference was not statistically significant (P> 0.05). There was no significant difference between the two groups in the improvement of ultraviolet spot and brown spots. The porphyrin was not affected by the subjects. To conclude, SVF can promote the effect of fat transplantation in the improvement of facial skin quality.
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BACKGROUND: Many studies have shown that cell-assisted lipotransfer promotes the survival of fat implants, but there are few studies exploring stromal vascular fraction (SVF) -assisted fat transplantation for improving facial skin quality. OBJECTIVE: To evaluate the clinical effect of autologous adipose-derived SVF-assisted autologous fat transplantation in the improvement of facial skin quality. METHODS: From September 2016 to June 2018, in the Plastic and Cosmetic Center, the Affiliated Hospital of Xuzhou Medical University, 20 patients were recruited and then randomly divided into experimental group and control group, namely SVF-assisted fat transplantation and simple fat transplantation, respectively. A VISIA skin detector was used to detect facial skin quality of patients preoperatively and 6 months postoperatively, including spots, wrinkles, texture, pores, ultraviolet spots, brown spots, couperose skin and porphyrin. RESULTS AND CONCLUSION: The effect of improvement in wrinkles and texture in the experimental group was better than that of the control group, and the difference was statistically significant (P < 0.05). In terms of spots, pores and couperose skin, the therapeutic effect in the experimental group was better than that in the control group, but the difference was not statistically significant (P> 0.05). There was no significant difference between the two groups in the improvement of ultraviolet spot and brown spots. The porphyrin was not affected by the subjects. To conclude, SVF can promote the effect of fat transplantation in the improvement of facial skin quality.
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Objective A database of normal people's external nose was established through 3D measurement. This database was used to customize the external nose for patients with nasal defects and to assist the operator to carry out the whole nose reconstruction surgery, so as to carry out the postoperative evaluation.Methods 3D scanning of the subject's face, measurement of relevant indexes of the nose and establishment of a database, the operator used normal nose database to customize the customized external nose for 17 patients with nasal defects, assisted them in the whole nose reconstruction surgery, and used independent sample t test for data statistics to evaluate the expected effect of surgery. Results There was no statistically significant differences between the postoperative actual data and the preoperative personalized data (P> 0.05) in right root wing distance, left root wing distance, nose length, nasal base width, nose width, right side vertical bisect nasal line, left side vertical bisect nasal line, nose height, medial malleolus spacing, face width, mouth split width, facial height, nasal width index, nasal width index, interondylar-nasal width index and nasal high index. The actual data of nasal deep was statistically different from preoperative personalized data (P < 0.05). Conclusions Analysis showed no significant difference between the actual data nasal surgery and preoperative customization data. 3D measurement of normal human external nasal establishment database to customize the external nose for patients with nasal defects, can assist the surgeon to perform total nasal reconstruction surgery and improve predictability and make surgery more precise. Postoperative assessments can also be performed to compare preoperative and postoperative outcomes.
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Objective@#The purpose is to explore the method and clinical effects of total nasal reconstruction with the assistance of three-dimensional (3D) scanning, 3D printing and monitoring the blood circulation after operation.@*Methods@#3D scanning: Artex Eva 3D scanner was used to record the nose data of 500 volunteers from Xuzhou Medical University and its affiliated hospital from September 2016 to February 2017. A nose database of normal individuals was established, of which male was 138 and female was 362. In addition, 3D facial scanning was performed in patients wish to total nasal reconstruction. 3D printing: The individualized nasal structure was designed, with the assistant of patients′facial characteristics, combined with the normal nose database and the opinion of the patients. Anactual nose model was used as guidance during the operation. Postoperative monitoring: The blood flow and the retraction rate of forehead flap after surgery were measured using Laser Doppler Flowmeter and Geomagic Qualify software. The blood flow values, the temperature and the surface area of the flap were recorded and analyzed.@*Results@#The nasal database of normal people in the Huaihai region successfully established. Overall, the width of the nose takes up a quarter of the width of the faces, and the length is 1/3 of the distance from the hairline to the chin. From February 2017 to June 2018, 7 cases underwent total nasal reconstruction operations were performed by this procedure. The nasal models were all successfully printed out, as the guide of flap taken during the operation. The mean operation time of the cases was (2.45±0.75) h, and the follow-up time was 5-15 months, with an average of 12.5 months. After the operations, the retraction rate of the forehead flap were (21.8±2.72)% in one month, and (29.1±1.82)% in six months. All patients are satisfied with the nasal appearance.@*Conclusions@#Nasal reconstruction with forehead flap based on 3D scanning and 3D printing, provides objective targets for nasal fine-structure in a noninvasive way. The postoperative monitoring of the blood flow promotes the successful completion of the total nose reconstruction.
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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 study the effect of the lentivirus encoding acidic fibroblast growth factor transfecting human adipose-derived stem cells (ADSCs) on the cell cycle and proliferation of ADSCs.Methods ADSCs were isolated and extracted by enzymatic digestion from the liposuction aspirate.ADSCs were cultured,identified and osteogenic induced reagent was used to induce differentiation of ADSCs towards bone cells.To obtain lentivirus encoding FGF-1,the plasmid PWPXLd FGF-1 was co-transfected with plasmid psPAX2,pMD2.G in 293T cells.ADSCs were infected with lentivirus encoding FGF-1.Expression of green fluorescent protein (GFP) in infected FGF-1 was observed by fluorescence microscope and expression of FGF-1 in ADSCs was verified by Western blot analysis.Flow cytometry was used to detect the cell cycle of ADSCs infected with lentivirus encoding FGF-1.EDU assay was performed to examine cell viability.Results Lentivirus encoding FGF-1 was obtained.After ADSCs being infected green fluorescence was found in about 70％ ADSCs,and overexpression of FGF-1 protein was detected in infceted cells by Western blot analysis.The percentage of G2/M phase cells was significantly increased compared with the control group,and the proliferation of ADSCs infected with lentivirus encoding FGF-1 was promoted as compared with the control group.Conclusions FGF-1 can enhance G2/M phase of ADSCs and promote the proliferation of ADSCs.
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Objective To detect the characteristics and in vitro cell compatibility of human acellular dermal matrix (ADM) with the improved method.Methods Cell components of healthy human skins were removed by the improved method and the traditional method respectively.The porosity, degradation time in vitro of the ADM prepared by two methods and the cytotoxicity of the material infiltration liquid with the improved method on the adipose derived stem cells were detected.HE staining was used to detect the residual of the cells, the integrity of collagen and cell biocompatibility.Scanning electron microscopy (SEM) was used to detect the pore size.Results Both the two methods could completely remove the cells, and maintain the integrity of the collagen scaffold;The porosity of ADM with the improved method was higher (93.1±1.02)% than that of traditional method (74.27±2.04)% (P<0.05);There was no significant difference in the cytotoxicity and in vitro degradation time between the two kinds of ADM;While pore diameter of the improved method was significantly higher [(181.21±66.9) μm] than that [(102.38±15.63) μm] in dermal reticular surface with the traditonal method (P<0.05).Conclusions There is no obvious cytotoxicity of the ADM with the improved method, and therefore it is more suitable for cell adhesion growth with higher porosity and larger pore size.
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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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Objective To observe and explore the value of combined detection of serum BNP and TnI in patients with acute heart failure(AHF). Methods 60 AHF patients from February 2015 to April 2016 were selected as observation group, and divided into Level III group(n=32) and Level IV group(n=28) according to NYHA classification;60 healthy patients at the same period were selected as control group.BNP and TnI levels were detected by chemiluminescence in each group, and were compared.Results BNP and TnI of observation group were(2624.4 ±378.6) ng/mL and(0.87 ± 0.22) ng/mL, while in control group were(72.4 ±20.2) ng/mL and(0.13 ±0.02) ng/mL, the differences were significant(P<0.05);BNP and TnI in level III group before treatment were significantly lower than grade IV group(P<0.05), after treatment, the level of BNP and III in group IV and group TnI were significantly lower(P<0.05), and the levels of BNP and TnI were significantly lower than those in III group after treatment(P<0.05). Conclusion BNP combine with TnI levels monitoring play an important role in AHF diagnosis and prognosis evaluation, can help to guide the treatment of AHF.
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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 early diagnosis and disease evaluation value in patients with acute pancreatitis by various serum cytokines measurement.Methods Forty-eight acute pancreatitis patients were divided into two groups based on the results of computed tomography (CT) examination:mild acute pancreatitis group (30 cases) and severe acute pancreatitis group (18 cases).The other 30 normal persons were selected as control group.The various serum cytokines were measured by enzyme-linked immunosorbent assay (ELISA).Results The serum concentrations of interleukin(IL)-1,IL-10 and tumor necrosis factor (TNF)-α in mild acute pancreatitis group were significantly higher than those in control group [(25.00 ± 1.92) ng/L vs.(10.08 ± 2.65) ng/L,(59.78 ± 4.51) ng/L vs.(1.80 ± 0.66) ng/L,(55.31 ± 8.54) ng/L vs.(18.72 ± 7.84) ng/L,P ＜ 0.05].The serum concentrations of IL-1,IL-6,IL-8,TNF-α and platelet activating factor (PAF) in severe acute pancreatitis group were significantly higher than those in mild acute pancreatitis group [(93.27 ± 3.98) ng/L vs.(25.00 ± 1.92) ng/L,(397.84 ± 13.05) ng/L vs.(34.12 ± 4.96) ng/L,(93.32 ±3.40) ng/Lvs.(13.06± 1.86) ng/L,(181.94 ±7.54) ng/Lvs.(55.31 ±8.54) ng/L,(284.53 ±7.88) ng/L vs.(175.25 ±30.15) ng/L,P＜0.05].Conclusion The various serum cytokines measurement has great importance on the early diagnosis of acute pancreatitis and discrimination between the mild acute pancreatitis and severe acute pancreatitis.
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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.