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Objective:To observe any effect of exosomes derived from umbilical cord mesenchymal stem cells on pain, cartilage repair and the expression of transcriptional activator 3 (ATF-3) and growth related protein 43 (GAP-43) in the dorsal root ganglia (DRG), as well as to explore the mechanism of their relieving pain.Methods:Fifty-four male Sprague-Dawley rats were randomly divided into a sham-operation group, a monoiodoacetate group and an exosome group, each of 18. The knee cavities of the left hind limbs of all of the rats except those in the sham-operation group were injected with 50μl of monoiodoacetate to establish an arthritis pain model. The sham-operation group received only 50μl of saline solution as controls. Two weeks after the modelling, the knee joint cavities of the exosome group were injected with 50μl of exosomes, while the other two groups were injected with 50μl of normal saline. The rats′ mechanical and thermal pain thresholds were measured 1 day before the modeling, 7 and 14 days after the monoiodoacetate injection, as well as 7, 14 and 28 days after the exosome injection. Western blotting was used to detect the expression of ATF-3 and GAP-43 in the rats′ DRG, while hematoxylin and eosin staining was used to detect any cartilage repair.Results:Compared with the monoiodoacetate group, the latency of the mechanical and thermal pain thresholds had increased significantly in the exosome group 7 days after the exosome injection. The difference remained significant until the 28th day after the injection. The expression of ATF-3 protein decreased significantly and that of the GAP-43 protein increased significantly. Significant differences were observed in the average Osteoarthritis Research Society International (OARSI) knee cartilage score.Conclusions:Exosomes can alleviate the pain induced by monoiodoacetate adjuvant. The analgesic mechanism may be related to reducing nerve injury and promoting nerve and cartilage repair, with the nerve repair earlier than cartilage repair.
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Objective To establish a method of quick three-dimensional (3D) reconstruction of pubic symphysis based on magnetic resonance imaging. Methods The pelvis images of adult male were generated on a 3.0 T scanner using a T1 Gradient Echo FLASH-3D (T1- FL3D) sequence and imported the images into medical image control system. Segmentation of binaryzation threshold was conducted and pelvic soft tissue image was extracted by regional growth, 3D structure model of pubic symphysis was obtained by Boolean operation. The 3D structure model of pubic symphysis was established by the noise reduction of reverse engineering software. And compared with the 3D reconstruction model pubic bone CT scan. Results The morphological characters of the MRI pubic symphysis 3D model, such as the ridges and furrows on the symphysial surface, lower extremity, dorsal margin (beveling), margin (beveling) and pubic tubercle, were highly consistent with the morphological characters of the 3D model established by CT scan. Conclusion MRI scan can be used to reconstruct the 3D structure of pubic symphysis quickly and effectively, and it can provide a safe radiation-free 3D visualization imaging technique for forensic age estimation for the living.
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Objective To investigate changes of oxidative stress in the hearts and their roles in diabetic cardiomyopathy of rats.Methods Thirty six rats with diabetic cardiomyopathy were induced by streptozocin(STZ),and were randomly divided into three groups:(1) normal control (NC) group,normal control rats; (2) Diabetes maintain (DM) group,diabetic rats received protamine zinc insulin (PZI) 2 ～4 U/2 days; and (3) Diabetes treatment (DT) group,diabetic rats received PZI 9 ～ 12 U/kg body weigh/day.After 12 weeks,rats were killed,and the parameters including blood glucose,glycated hemoglobin a1c (HbA1c),total cholesterol (TC),and triglyceride (TG) were measured.Maleic dialdehyde (MDA),the total antioxidant capacity of the hearts,and activities of antioxidant enzymes in hearts including total superoxide dismutase (TSOD),catalase (CAT),and glutathione peroxidase (GSH-Px) were measured by chromatometry.The pathologic changes of heart tissues were studied by hematoxylin eosin (HE),periodic acid Schiff (PAS),and nitrotyrosine (NT) staining.Results After 12 weeks,parameters including blood glucose,HbA1c,TC,and TG were increased significantly in DM group relative to NC group or DT group.The level of heart MDA was elevated obviously in the DM group [NC group (7.82 ±0.62) nmol/mgprot,DM group (12.81 ± 1.35) nmol/mgprot,and DT group (8.20 ±0.85) nmol/mgprot].The total antioxidant capacity and activities of TSOD [(235.09 ± 14.94) U/mgprot],GSH-Px [(56.59 ± 4.96) U] were decreased significantly in the DM group.The level of CAT [(13.84 ± 2.88) U/mgprot] was increased obviously in the DM group.The cardial myocyte hypertrophy,myocyte steatosis,and PAS positive material could be found in the hearts of rats in DM group with HE and PAS staining.The expression of NT was much stronger in DM group than NC or DT groups.NT was less expressed in DT group,and was almost not expressed in the NC group.Conclusions Oxidative stress could be caused by diabetes in diaetic hearts and might play an important role in the etiology of diabetic cardiomyopathy.
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Objective To explore the application probability of AFLP method (Amplified Fragment Length Polymorphism) for the genetic study of complicated samples and to make clear the analytical approach on polymorphic fragments.Methods An AFLP fragement sequence,resolved between murines with high and low metastatic hepatocarcinoma,was retrieved from a reference paper and was annotated on NCBI database by different BLAST programs. Results The retrieved information varied along with the different BLAST programs, nevertheless BLASTX program provided more comprehensive information and showed that the differentiated fragment exhibited high homology with terminase large subunits from Burkholderia phage Bups phil and Mycobacterium tuberculosis H37Ra. However, there is no detailed information for the latter,key words search returned the relationship between terminase and the formation and metastasis of tumors and provided the further research clues. Conclusions Whole genome screening in parallel of AFLP markers between genetically differentiated paired materials would readily produce informatic fragments. Further analysis of the sequenced fragments by informatic methods would facilitate the subsequent research.
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Aim To study the application status of odds ratio for medical animal experiments.Methods Odds ratio and seven kinds of animal models were used as retrieval strategy to search medical animal experiment related papers in several Chinese and English databases.Papers relating to each kind of animal model and using odds ratio in abstract and text were counted. Data from different databases were compared. Calculation of odds ratio was exemplified and the significance of different odds ratio values was illustrated in this paper.Results Few medical animal experiments cited odds ratio as statistics.Conclusions The importance of odds ratio has not been fully recognized in Chinese references.
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Aim To investigate the ability of chymase inhibitors o n histamine release from human colon mast cells. Methods Human ma st cells were dispersed from colon tissue with collagenase and hyaluronidase, an d were challenged with stimulus for 15 min at 37℃.A glass fibre-based fluorome tric assay was used to measure histamine in the supernatants of dispersed mast c ells.Results chymase inhibitors ZIGPFM, TPCK and ? 1-antitry psin failed to induce significant histamine release from colon mast cells. All t he chymase inhibitors were able to inhibit anti-IgE induced histamine release i n a concentration dependent manner with a maximum of 37%, 26% and 36.8% inhibit ion being achieved with 1 mmol?L -1 of ZIGPFM, 80 mmol?L -1 of TPCK , 30 mmol?L -1 of ? 1-antitrypsin, respectively. Preincubation of inhib itors of ZIGPFM and TPCK with cells for 20 min at 37℃ before challenging with a nti-IgE was able to slightly enhance their inhibitory actions. All the chymase inhibitors were able to inhibit calcium ionophore induced histamine release, th e maximum inhibition was 23.6%～35%.And the extent of inhibition by TPCK was in creased when colon mast cells were preincubated for 20 min before calcium ionoph ore being added. However, the same treament failed to improve the action of ZIGP FM. Conclusion In the current study, we found that inhibitors o f chymase were able to inhibit anti-IgE and calcium ionophore induced histamine release from human colon mast cells, which may indicate a potential of a novel therapy for the treatment of inflammatory bowel disease or other mast cell relat ed diseases.
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Objective:To investigate the ability of chymase inhibitors on tryptase release from human colon mast cells.Methods:Human mast cells were dispersed from colon tissue with collagenase and hyaluronidase,and were challenged with stimulus for 15 min at 37℃.Tryptase assay performed following previous procedures.In brief,a 96-well microtitre plate was coated with antiserum to human tryptase.The tryptase levels in the samples were detected with a monoclonal antibody specific to tryptase and the reaction was visualized by addition of OPD.Results:At 15 min and 35 min following incubation,anti-IgE and calcium ionophore were able to provoke significant tryptase release from human colon mast cells.Chymase inhibitors ZIGPFM,TPCK and ?1-antitrypsin had no stimulatory effect on colon mast cells at both 15 min and 35 min incubation periods.All the chymase inhibitors were able to inhibit anti-IgE induced tryptase release in a concentration dependent manner with a maximum of 37%,40% and 36.6% inhibition being achieved with 1 ?mol/mL of ZIGPFM,80 ?mol/mL of TPCK,30 ?mol/mL of ?1-antitrypsin,respectively.Preincubation of inhibitors of ZIGPFM and TPCK with cells for 20 min at 37℃ before challenging with anti-IgE was able to slightly enhance their inhibitory actions.Amastatin,a specific inhibitor of aminopeptidase,had no effect on anti-IgE induced tryptase release.All the chymase inhibitors were able to inhibit calcium ionophore induced tryptase release,the maximum inhibition were 23%-35.3%.And the extent of inhibition by ZIGPFM was increased when colon mast cells were preincubated for 20 min before calcium ionophore being added.However,the same treament failed to improve the action of TPCK.Conclusion:We found for the first time that inhibitors of chymase were able to inhibit anti-IgE and calcium ionophore induced tryptase release from human colon mast cells,which may indicated a potential of a novel therapy for the treatment of inflammatory bowel disease or other mast cell related diseases.
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AIM: To investigate the ability of Chinese cobra snake venom-metalloproteinase(MT) to induce the histamine release from human mast cells and its potential mechanisms.METHODS: MT was purified from the snake venom by using heparin agarose and Superdex75 chromatography.Mast cells were dispersed from human lung, colon and tonsil tissues after digestion with collagenase and hyaluronidase.The dispersed mast cells were then challenged with MT,stimulus and control in LP4 tubes for 15 min at 37 ℃.A glass fibre-based fluorometric assay was used to measure histamine in the supernatants of dispersed mast cells.RESULTS: MT induced a dose-dependent release of histamine from human colon,lung and tonsil mast cells.As low as 0.03(mg/L) of MT was able to stimulate significant histamine release from human colon mast cells,but a minimum of 0.3 or 30 mg/L of MT was required to stimulate a similar level of histamine release from lung or tonsil mast cells,respectively.The release of histamine from colon and lung mast cells in response to MT was maximized at 12 min following the addition of the stimulus.This was quite different from the picture of the peak histamine release from tonsil mast cells,in which histamine release was maximized at 8 min following the addition of MT.Pretreatment of cells with metabolic inhibitors and pertussis toxin reduced dramatically histamine release from human colon,lung and tonsil mast cells by MT.In exogenous Ca~(2+) and Mg~(2+) free experiments,the release of histamine induced by MT was significantly decreased.CONCLUSION: Cobra snake venom MT induces human mast cells to release histamine through a G-protein-related mechanism,which may contribute to the pathogenesis of venomous snake bite.
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AIM: To investigate the changes of oxidative stress in the kidneys and their roles in nephropathy in diabetic rats. METHODS: Diabetic rats were induced by streptozotocin (STZ). 36 rats were divided into three groups randomly: (1) NC group, normal control rats; (2) DM group, diabetic rats received protamine zinc insulin (PZI) 2U-4U/2 d; (3) DT group, diabetic rats received PZI 9-12 U/kg body weigh/day. 12 weeks later, rats were killed, blood glucose, blood cholesterol, serum creatinine, blood urea nitrogen, HbA1c, urinary creatinine, and urinary protein for 24 h were measured. The activities of antioxidant enzymes in renal cortex, including total superoxide dismutase (TSOD), Cu-Zn superoxide dismutase (Cu-Zn SOD), Mn superoxide dismutase (Mn SOD), catalase (CAT), glutathione peroxidase (GSH-Px), and maleic dialdehyde (MDA) were measured by chromatometry. RT-PCR was performed to detect the expression of different antioxidant enzymes mRNA. RESULTS: For all the targets we measured, there was no significant difference between NC and DT groups. Compared with the other two groups, the levels of blood glucose, cholesterol, trigalloyl glycerol, HbA1c in DM group increased significantly. The activities of TSOD, Cu-Zn SOD and CAT decreased significantly. The activity of GSH-Px increased significantly. There was no significant difference among the activities of Mn SOD in all three groups. The level of MDA in DM group was much higher than that in NC or DT group. The relative expression levels of GSH-Px and Cu-Zn SOD mRNA in DM group were higher than those in other two groups, while the relative expression level of CAT decreased. Mn SOD mRNA was expressed without significant difference in all groups. Compared with NC or DT group, urinary protein in DM group increased significantly, while creatinine clearance rate decreased. CONCLUSIONS: Hyperglycemia affected the expression of antioxidant enzymes. Oxidative stress was caused by hyperglycemia in diabetic rats and may be an important factor in the etiology of diabetic nephropathy.