BACKGROUND:Choosing an effective means to label and trace the distribution, differentiation and migration of celsin vivo help to further explore the specific mechanism of cels that exert a therapeutic effect. OBJECTIVE:To understand the migration and localization of BrdU-labeled human umbilical cord mesenchymal stem cels in brain injury model rats. METHODS:Human umbilical cord blood samples were obtained, and the isolation of human umbilical cord mesenchymal stem cels was carried out. The primary and passage culture were performed. The phenotype of cels was detected by flow cytometry. Passage 3 human umbilical cord mesenchymal stem cels were labeled using BrdU, and the cel proliferation was detected using MTT method. BrdU-labeled cels were injected into brain injury ratsvia the tail vein. At 14 days after transplantation, brain tissues in the injury region were cut into sections and the migration and location of the umbilical cord mesenchymal stem cels were observed under inverted
fluorescence microscope. RESULTS AND CONCLUSION: Cel surface specific markers CD45 and CD34 were detected by flow cytometry, but the cels could not express CD44, CD105 and CD29. Based on the cel growth curve, the cels came into a conditioning period at 1-3 days of seeding and came into a logarithmic phase at 3-5 days. BrdU-positive cels were visible at the injury region after 14 days, indicating that in the rats, transplanted human umbilical cord mesenchymal stem cels migrated from the peripheral blood to the site of brain injury to achieve the effective repair of injured parts. Cite this article:Liu HL, Liu ZJ, Chen XB, Hu WZ, Ding BQ. Migration and localization of umbilical cord mesenchymal stem cels implanted into brain injury model rats. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):31-35.

BACKGROUND:Bone marrow stromal cel s can differentiate into nerve cel s to promote nerve tissue repair, but the exact mechanism has not been ful y elucidated.
OBJECTIVE:To explore the influence of adenovirus-mediatedβnerve growth factor transfection on bone marrow stromal stem cel transplantation fighting against brain injury in rats.
METHODS:(1) Rat bone marrow stromal stem cel s were cultured in vitro, transfected with the adenovirus-mediatedβnerve growth factor and directional y induced usingβ-mercaptoethanol. (2) A total of 210 Sprague-Dawley rats were randomized into induction+tranfection group, induction+non-transfection group, induction+medium group, model group, and sham group (n=42 per group). Rat skul injury models were made, and given corresponding treatments at different time points (12, 24, 36, 48, 72 hours). Neurological function of rats was evaluated based on neurological severity scores on the day that the rats were given transplantation, and 1, 2, 3, 4 weeks after transplantation. (3) Another 75 Sprague-Dawley rats were also divided into five groups (n=15 per group) as above, fol owed by model establishment and corresponding treatments at 24 hours after modeling. Neurological severity scores were recorded at the same day, 1, 2, 3, 4 weeks after transplantation. Five rats from each group were sacrificed to detect levels of malondialdehyde and superoxide dismutase in the rat brain at the same day, 2 and 4 weeks after transplantation, respectively.
RESULTS AND CONCLUSION:If the cel s were transplanted within 48 hours after modeling, the neurological severity scores in the induction+transfection group decreased significantly compared with the induction+non-transfection group and model group at 1 and 2 weeks after transplantation (P<0.05). If the cel s were transplanted at different time, the neurological severity scores in the induction+transfection group were decreased significantly compared with the induction+non-transfection group and model group at 3 and 4 weeks after transplantation (P<0.05). If the cel s were transplanted within 24 hours after modeling, the neurological severity scores in the induction+transfection group decreased significantly compared with the model group at 1 week after transplantation (P<0.05), and the neurological severity scores in the induction+transfection group and induction+non-transfection group both were significantly lower than those in the model group (P<0.05). Two weeks after cel transplantation, the level of superoxide dismutase was significantly higher in the induction+transfection group than the induction+medium group and model group (P<0.05), but the level of malondialdehyde was significantly lower (P<0.05). Al these findings indicate that adenovirus-mediatedβnerve growth factor transfer plays a certain neuroprotective role in bone marrow stromal stem cel transplantation for brain injury in rats.

BACKGROUND:Bone marrow mesenchymal stem cells(BMSCs)differentiating into neural cells is an effective way of cell therapy of nervous system disease.However,the methods used nowadays still need to be improved.OBJECTIVE:To induce the differentiation of rhesus monkey BMSCs into neuron-like cells by using sonic hedgehog factor.METHODS:Rhesus monkey BMSCs differentiating into neuron-like cells was induced by typical retinoic acid and sonic hedgehog factor.Rhesus monkey BMSCs were isolated and cultured by density gradient centrifugation method.Cell growth was observed under an inverted phase contrast microscope and cell growth curve was determined using MTT assay.Flow cytometry was performed to characterize the phenotype of BMSCs,and immunohistochemistry was utilized to assess differentiated cells.Ultra-structure of the differentiated cells was observed by transmission and scanning electron microscopes.RESULTS AND CONCLUSION:Rhesus monkey BMSCs cultured in vitro were identified by flow cytometry,with high homogenicity.Following sonic hedgehog factor disposal for 7 days,differentiated cells were mainly positive for neurone specific enolase,neurofilament protein,Tau and glial fibrillary acidic protein(GFAP).Image statistical analysis found that in sonic hedgehog factor scheme,neural stem cells marker Neetin positive rate was significantly higher compared with the rstinoic acid scheme(P＜0.01).GFAP-positive rate was greater in the retinoic acid scheme than in the sonic hedgehog factor scheme(P＜0.05).Results indicated that sonic hedgehog factor scheme is an effective pathway of rhesus monkey BMSC differentiation into neuron-like cells.

BACKGROUND: Present studies have demonstrated that during neural development, differentiation of neural stem cells (NSCs) was affected by various regulatory factors from surrounding microenvironment. Sonic hedgehog (Shh) is a key induction signal during neural fetal development, and can be an effective inductor to regulate differentiation of neural cells. OBJECTIVE: To investigate the signal transduction pathway of SHH for differentiation of rhesus monkey bone marrow mesenchymal stem cells (BMSCs) into of neuron-like cells by sonic hedgehog factor. METHODS: Rhesus monkey BMSCs were isolated and cultured by conventional density gradient centrifugation. BMSCs in the induction group were treated with L-DMEM containing FGF2, B27 and fetal bovine serum for preinduction of 24 hours, and then with DMEM supplemented with 0.5 μmol/L retinoic acid or 400 μmol/L SHH for 8 days. Non-induced cells served as control group. Following labeling with neuron enolase, positive cells were screened by flow cytometry. RT-PCR and Western-blot were used to detect SHH- and retinoic acid-induced cell membrane receptor and intracellular signal protein changes. RESULTS AND CONCLUSION: SHH specific membrane receptor Ptc, retinoic acid specific receptor RARα, signal protein molecule ptch1 and Smad expressed in normal cells. Ptc expression upregulated in SHH-induced cells. High expression lasted for a long time with induction time, which was significantly stronger compared with the retinoic acid and control groups (P < 0.01). Intracellular ptch1 protein molecule expression showed similar tendency as this, but could not induce upregulation of RARα expression. During induction, retinoic acid-stimulated cells did not activate Ptc pathway. Four days following induction, RARα expression upregulated and lasted till 6 days, but there were no significant differences. No significant change in ptch1 expression was determined. SHH- and retinoic acid-induced cell Smad molecule expression upregulated, but no significant difference was determined. Results verified that SHH-induced scheme participated in cell induction and differentiation by persistently activating its specific receptors. However, there was no significant receptor pathway crossing between retinoic acid-induced and SHH-induced schemes.

Objective To discuss the efficacy of the flexible ureteroscopy combined with Holmium laser lithotripsy for kidney calculi with HIV positive patients.Methods From May 2015 to May 2016,47 cases of patients with renal calculi were treated by the flexible ureteroscopy combined with Holmium laser lithotripsy in our hospital.There were 29 cases male,18 cases female,aged from 22 to 56 years old,average 39 years.There were 38 cases with single stone,9 cases with multiple stones.There were 9 cases with stones on both sides.Flexible ureteroscopy channel sheath was used in surgery.Stones were fragmented by Holmium laser.During one-month follow-up after surgery,stone removal and stone free rate were recorded.Residual stones were re-treated with a secondary lithotripsy or ESWL.Results The flexible ureteroscopy channel sheath was indwelled successfully in all the cases.All stones were detected.The average operation time was 63min (range,42-141min) and the median postoperative stay was 4.5days (range,2-16 d).Among the 47 patients,41 patients underwent first-stage lithotripsy,6 patients underwent second-stage ESWL after lithotripsy,and 1 patient underwent third-stage lithotripsy.The one-month stone free rate was 87.2％ (41/47).The total stone free rate was 97.9％ (46/47) after second-stage lithotripsy.Postoperative fever occurred in 4 cases after lithotripsy.No blood transfusion,systemic infection,ureteral perforation,or ureteral avulsion occurred.The total complication rate was 8.5％ (4/47).The mean number of CD4 +T lymphocytes before lithotripsy was 402/μl,and was 410/μl 3 days after lithotripsy.There was no Statistical differences between them.Conclusions Flexible ureteroscopy combined Holmium laserlithotripsy could be a safe and effective treatment for kidney calculi patients with HIV/AIDS positive.

Objective To assess the safety and efficacy of percutaneous nephrolithotripsy withpneumatic and ultrasonic power in pediatric renal calculi. Methods The clinical data of 44 patientstreated with combination of pneumatic and ultrasonic power during pereutaneous nephrolithotripsywere retrospectively analyzed. The 44 patients had 49 renal calculi. The patients were all under 14years old. The average age was 11 years (range 7-14 years). There were 39 unilateral and 5 bilateralcalculi. Among the 44 patients,metabolic disturbance occurred in 19 cases (43.2%),anatomical dys function occurred in 15 cases (34.1%),urinary tract infection occurred in 14 cases (31.8%). Ante grade percutaneous access was established under ultrasound guidance,a combination of pneumatic andultrasonic lithotripsy were used. The effect was evaluated by postoperative KUB and ultrasonic. Re suits The access was successfully established in all patients. Complete stone clearance was achievedin 36 kidneys in phase Ⅰ,stones from 9 kidneys were completely removed with second lithotripsy.Leftover stone in 2 kidneys were treated by ESWL. Open surgery was performed in 2 kidneys due toexcessive bleeding. The operative time ranged from 52 132 min,average time was 79 min. Two pa tients needed blood transfusion. No severe complications occurred in all patients. Thirty seven pa tients were followed up for 3 18 months. The renal function was not worsened and hydronephrosiswas not aggravated in these patients. Conclusion The percutaneous nephrolithotripsy with pneumatic and ultrasonic power is a safe,effective treatment for pediatric renal calculi.

Background Mitomycin C (MMC) is still irreplaceable until now.While it's current administration method has proved less than effective in the treatment of refractory glaucoma.To construct a MMC sustained release system which can maintain effective concentration and reduce toxicity is important for postoperative scarring regulation after glaucoma filtration surgery.Objective To evaluate the postoperative effect of use PTMC-F127-PTMC thermosensitive hydrogel as a new drug delivery carrier to sustained release MMC in rabbit trabeculactory.Methods Sixty rabbits,aged 10 to 14 weeks,were divided into 5 trabeculectomy groups in accordance with the random number table,including surgery only group,blank PTMC-F127-PTMC group and three sustained groups with 0.1 ml PTMC-F127-PTMC loaded with 0.05,0.10,0.20 g/L MMC injected after surgery.The MMC concentration of anterior chamber aqueous in three sustained release group with 0.1 ml PTMC-F127-PTMC loaded with 0.05,0.1,0.2 g/L MMC injected after surgery were tested by Guangzhou Analysis and Testing Center using high performance liquid chromatograph-mass spectrometer (HPLC-MS).At 3,7 days postoperatively,0.1 ml aqueous humor from 2 random selected rabbits in each group was extracted using 1 ml syringe with 30G needles from corneal limbus.At postoperative 1,3,5,7,10,14,28 days,bleb width and depth were calculated with caliper measurements and height was graded semiquantitatively by slit-lamp examinations,intraocular pressure (IOP) was measured with Tonopen.And corneal endothelial cell densities were examined by corneal endothelial counting before and 28 days after surgery.Sequential sections of the operative region were prepared and stained with hematoxylin and eosin and proliferating cell nuclear antigen (PCNA) after taking off the eyeballs from dead rabbits at 28 days later.Results MMC could be sustained released from PTMC-F127-PTMC hydrogel for more than 20 days.The mean postoperative bleb survival time in trabeculectomy surgery only group,blank hydrogel group and three sustained release groups were (5.3 ± 0.4),(5.5 ± 0.4),(12.2 ± 1.0),(25.1 ± 0.9),(26.7 ± 0.7) days respectively,the difference between each group was significantly (F =123.200,P =0.000).0.05 g/L MMC sustained release group has a better bleb survival time than that of surgery only group and blank hydrogel group (P =0.000).Compared with other groups,0.10 g/L and 0.20 g/L MMC sustained group has the longest bleb survival time (P =0.000),and more obvious IOP downtrending (F=53 010.000,P＜0.01).But the difference between the two groups was not significant.There was no difference in cornea endothelia cells counts between each group and no MMC was detected in aqueous humors.Histopathology test shows that the inflammatory response and fibrosis were lighter in MMC sustained release group,with stronger proliferation inhibition.Conclusions PTMC-F127-PTMC thermosensitive hydrogel can be a new drug delivery carrier to sustained release MMC.Sustain release MMC can extent bleb survival time with low toxicity.

ObjectiveTo investigate the role of p38 and ERK1/2 during rhesus monkeys mesenchymal stem cells differentiated into neuron-like cells.MethodsTo induce the neuronal phenotype,rhesus monkeys mesenchymal stem cells were maintained in sub-confluent cultures in serum-contain medium supplement with Sonic hedgehog.Western blot analysis the change of p38 and ERK1/2 during rhesus monkeys mesenchymal stem cells differentiated into neuron-like cells.Under transmission and scanning electron microscope,ultra-structure of the differentiated cell were observed.ResultsDuring BMSCs differentiated into neuron-like cells by SHH,Mitogen-activated protein kinases (MAPK) involved in their signal transduction,p38 was activated and ERK1/2 was inhibited.P38 inhibitor SB203580 increased induced differentiation time compared with normal induced cells,and inhibited neurite outgrowth.ConclusionActivation of p38 and inhibition of ERK was impacted on differentiation into neuron-like cells from rhesus monkeys mesenchymal stem cells induced by Sonic hedgehog,which may has potential application on neuroprotection of stem cells in Nervous system diseases

Through literature method,expert consultation method,K-means algorithm and empirical analysis method,the paper analyzes the methods for evaluation on the layout of first-aid sites,discusses the indexes for scientific evaluation on the rationality of layout of first-aid sites,conducts visualized and practical calculation based on the first-aid related data of Wuhan city in 2015,verifies the rationality and operability of the evaluation indexes,and provides reference for overall allocation of first-aid resources and rational layout of first-aid sites.

BACKGROUND:Chitosan biological materials can induce bone marrow mesenchymal stem cells to differentiate toward neurons. As a derivative of chitosan, carboxymethyl chitosan has a series of excelent properties. However, whether carboxymethyl chitosan can induce the neuronal differentiation of bone marrow mesenchymal stem cells remains unclear.OBJECTIVE:To investigate the effect of carboxymethyl chitosan thermosensitive hydrogel on the differentiation of bone marrow mesenchymal stem cells into neurons and the possible mechanism.METHODS:Passage 3 bone marrow mesenchymal stem cells from rats were selected and cultured in carboxymethyl chitosan thermosensitive hydrogel extracts in different concentrations (0, 50, 100, 150, 200, 500 g/L). Control cells were cultured in culture medium with no addition of carboxymethyl chitosan thermosensitive hydrogel extracts. MTT assay was performed to investigate the effects of different concentrations of carboxymethyl chitosan thermosensitive hydrogel extracts on bone marrow mesenchymal stem cell proliferation. Western blot assay was used to explore the effect of 150 g/L carboxymethyl chitosan thermosensitive hydrogel extracts on the expression of neuron-specific enolase, Nestin, Vimentin, NF-M, microtubule associated protein 2, glial fibrillary acidic protein, β3-tubulin, Notch1 and Jag1 protein.RESULTS AND CONCLUSION:MTT assay showed that carboxymethyl chitosan thermosensitive hydrogel promoted the cell proliferation, and the proliferation rate reached the peak at the concentration of 150 g/L. Western blot assay showed that the cells induced by 150 g/L carboxymethyl chitosan thermosensitive hydrogel extract had significant increases in neuron-specific enolase, Nestin, Vimentin, NF-M, microtubule associated protein 2, glial fibrillary acidic protein, and β3-tubulin protein expression, and obvious decreases in Notch1 and Jag1 protein expression in comparison with the control group. These results indicate that the carboxymethyl chitosan thermosensitive hydrogel induces rat bone marrow mesenchymal stem cells to differentiate toward neurons, and suppresses the activity of Notch signal pathway in the process of differentiation.