Objective To explore the serious situation of injection abuse, and its influence to average prescription fee. Methods The subjects of this study were health service settings in rural area of 9 provinces/cities in Midwest of China. The treatment prescription indicators of county and village health service settings were calculated. Results Prescription injection rates of health care facility in rural area of Midwest provinces/cities of China (25.8% to 62.2%,mean: 45.1%) were higher than the standard of WHO (13.4% to 24.1%), and the injection abuse situation was serious. Injection bause caused the increase of prescription fee. Excess usage of injection in health service settings was related to the economic level of the on-site county or village, and also related to the size and load of health service facilities. Conclusion Suggestions are proposed to the government health agency according to the results of the study: enhancing the lawmaking, establishing the related policy and effective measure, training the medical personnel, promoting the mass health education, investigating the effective injection management model in rural area, and reducing the rate of injection.
Objective To investigate the effect of cold ischemia on the development of transplant arteriosclerosis (TA) in rat aortic isografts. Methods Aorta grafts from SD and Wister rats were stored in a cold perfusion solution for 0.5 hours and 4 hours respectively before being orthotopically transplanted to Wister recipients. After observation times ranging from 15 to 60 days, the grafts were examined by using histological and electron microscopy techniques. Regional changes in the lumen, intima and media layers were measured by using an image analysis system. Results Partial intima thickings were showed in control isografts at 60 day posttransplantation. Pronounced intima thickings were seen in experimental isografts and control allografts at the same time. The thicking neointimas consist mainly of monocyte/macrophage and smooth muscle cells (SMC). The broken interior elastic lamina (IEL) and necrosis SMC in media were detected in allogenic grafts. Conclusion The damage due to prolonged cold ischemia time is sufficient to cause pronouced graft arteriosclerosis.
ObjectiveTo investigate the protective effects of carboxymethylated chitosan (CMCS) on oxidative stress induced apoptosis of Schwann cells (SCs), and the expressions of brain derived neurotrophic factor (BDNF) and gl ial cell line derived neurotrophic factor (GDNF) in oxidative stress induced SCs.
MethodsTwenty-four 3-5 days old Sprague Dawley rats (weighing 25-30 g, male or female) were involved in this study. The bilateral sciatic nerves of rats were harvested and SCs were isolated and cultured in vitro. The purity of SCs was identified by immunofluorescence staining of S-100. SCs were treated with different concentrations of hydrogen peroxide (H2O2, 0.01, 0.10, and 1.00 mmol/L) for 3, 6, 12, and 24 hours to establ ish the apoptotic model. The cell counting kit 8 (CCK-8) and flow cytometry analysis were used to detect the cell viabil ity and apoptosis induced by H2O2, and the optimal concentration and time for the apoptotic model of SCs were determined. The 2nd passage SCs were divided into 5 groups and were treated with PBS (control), with 1.00 mmol/L H2O2, with 1.00 mmol/L H2O2+50 μg/mL CMCS, with 1.00 mmol/L H2O2+100 μg/mL CMCS, and with 1.00 mmol/L H2O2+200 μg/mL CMCS, respectively. After cultured for 24 hours, the cell viabil ity was assessed by CCK-8, cell apoptosis was detected by flow cytometry analysis, the expressions of mRNA and protein of BDNF and GDNF were detected by real-time quantitative PCR and Western blot.
ResultsThe immunofluorescence staining of S-100 indicated the positive rate was more than 95%. CCK-8 and flow cytometry results showed that H2O2 can inhibit the proliferation of SCs and induce the SCs apoptosis with dose dependent manner, the effect was the most significant at 1.00 mmol/L H2O2 for 24 hours; after addition of CMCS, SCs exhibited the increased proliferation and decreased apoptosis in a dose dependent manner. Real-time quantitative PCR and Western blot analysis showed that 1.00 mmol/L H2O2 can significantly inhibit BDNF and GDNF expression in SCs when compared with control group (P<0.05), 50-200 μg/mL CMCS can reverse the oxidative stress-induced BDNF and GDNF expression in SCs in a dose dependent manner, showing significant difference compared with control group and 1.00 mmol/L H2O2 induced group (P<0.05). There were significant differences among different CMCS treated groups (P<0.05).
ConclusionCMCS has the protective stress on oxidative stress induced apoptosis of SCs, and may promote the BDNF and GDNF expressions of neurotrophic factors in oxidative stress induced SCs.
Objective To set up and to evaluate an acute closed brain injury model in rats. Methods The acute closed brain injury was produced in rats by using an impactor consisting of a stand, a guide tube, a weight and a footplate. Ninetysix SD rats were divided into a control group(n＝32, no impact), a mild injury group(n＝32, impact once at force level of 400 g·cm) and a severe injury group(n＝32, impact once at force level of 800 g·cm) to elucidate the physiological responses, the pathophysiological changes and brain edema after brain injury at different injury levels. Results In the mild injury group and the severe injury group, a sudden rise or reduction of blood pressure, deep and fast breath apnea, and pain reflects inhibition were observed. The responses were more obvious in the severe injury group than in the mild injury group. The water content of the brain increased after 6 hours of injury. The pathological contusion and edema of brain were noted or above the impact force level of 800 g·cm. When the impact force rose to or over 1200g·cm, the animals died of persistent apnea mostly. Conclusion Although the established closed brain injury model with different biomechanical mechanisms as the clinical brain injury, it is in conformity with pathological changes and pathophysiological characteristics of acute clinical brain injury, it can be utilized extensively because of its convenient and practice.
Objective To examine the effects of carbon dioxide (CO2) pneumoperitoneum on local pancreas pathological changes, serum levels of amylase, IL-1, IL-6, and the positive rate of dissolubility adhesion molecule (CD11a/CD18 and CD11b/CD18) expression in rats with severe acute pancreatitis (SAP). Methods Fifty healthy male SpragueDawley (SD) rats were randomly divided into 3 groups: CO2 pneumoperitoneum group (n=20): SAP was induced by injecting 5% sodium taurocholate through retrogradely common biliopancreatic ducts via duodenal papilla， and then CO2 pneumoperitoneum was established at a pressure of 12 mm Hg (1 mm Hg=0.133 kPa) for 30 min; SAP group (n=20): The rats were treated as same as CO2 pneumoperitoneum group, except CO2 pneumoperitoneum； Simple operation group (n=10): Laparotomy was performed and nothing was done to duodenum and pancreas except for moving them softly. The blood samples were collected for examining serum levels of amylase, IL-1, IL-6, and the positive rates of CD11a/CD18 and CD11b/CD18 expression, and histopathologic examination of pancreas was performed. Results Compared with simple operation group, the pancreatic pathologic histology score, serum levels of amylase, IL-1, IL-6, and the positive rates of CD11a/CD18 and CD11b/CD18 expression were significantly higher in CO2 pneumoperitoneum group and SAP group (P=0.000). The levels of IL-1 and IL-6 were significantly lower in CO2 pneumoperitoneum group as compared to SAP group (P=0.000). There was no significant difference between CO2 pneumoperitoneum group and SAP group in pancreatic pathologic histology score (P=0.294), the level of serum amylase (P=0.073), the positive rates of CD11a/CD18 (P=0.155) and CD11b/CD18 expression (P=0.201). Conclusion CO2 pneumoperitoneum has inhibitory effect on the levels of IL-1 and IL-6, rather than the positive rates of CD11a/CD18 and CD11b/CD18 expression in SD rats with SAP.
ObjectiveTo study the effects of the new small molecular oxygen free radical scavenger Tempol on the survival and vasculogenesis of the long random pattern skin flap (LRPSF) and its mechanism.
MethodsEighty-four male Sprague Dawley rats were randomly divided into control and Tempol groups (42 rats in each group). LRPSF of 9 cm×3 cm in size were prepared on the backs of rats in two groups based on the Mcfarlane flap. Rats were administered with Tempol (100 mg/kg) in the Tempol group and with normal saline in the control group by intraperitoneal injection at 15 minutes before operation and at 1-7 day after operation. The rat and the skin flap survival conditions were observed after operation; the survival rate of skin flap was measured, and the vascular structure, vascular volume, and total length of blood vessels were analyzed with Micro-CT three-dimensional imaging after 7 days; HE staining was used to observe the structure of the skin flaps and inflammation, immumohistochemical staining to observe vascular endothelial growth factor (VEGF) expression; water-soluble tetrazolium-1 method was used to measure the content of superoxide dismutase (SOD) and malondialdehyde (MDA), and ELISA to detect the expressions of tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) after 1, 3, and 7 days.
ResultsAll of rats survived after operation, without hemorrhage, edema, and infection. With the extension of time, necrosis occurred in the distal part of the skin flaps in 2 groups, but the necrosis degree of the Tempol group was lower than that of control group; meanwhile, the blood vessel distribution and continuity were better than those of control group. The skin flaps survival rate, vascular volume, and total length of blood vessels of Tempol group were significantly higher than those of control group after 7 days (P<0.05). The clearer skin flaps structure, lighter inflammation reaction and inflammation cell infiltration, and higher VEGF staining intensity were observed in the Tempol group than the control group after 7 days. There was no significant difference in SOD, MDA, and TNF-α, and IL-6 contents between the 2 groups at immediate after operation. SOD significantly increased, but MDA, TNF-α, and IL-6 contents significantly decreased in the Tempol group when compared with control group after 1, 3, and 7 days (P<0.05).
ConclusionTempol can significantly promote the LRPSF survival rates, its mechanism is closely related to the promotion of vasculogenesis and reduction of oxidative stress and inflammation.
Objective To explore the in vitrodifferentiation of the rat mesenchymal stem cells (MSCs ) into the skeletal muscle cells induced by the myoblast differentiation factor (MyoD) and 5-azacytidine. Methods The MSCs were taken from the rat bone marrow and the suspension of MSCs was made and cultured in the homeothermia incubator which contained 5% CO2at 37℃. The cells were observed under the inverted phase contrast microscope daily. The cells spreading all the bottom of the culture bottle were defined as onepassage. The differentiation of the 3rd passage of MSCs was induced by the combination of 5-azacytidine, MyoD, transforming growth factor β1, and the insulin like growth factor 1. Nine days after the induction, the induced MSCs were collected, which were analyzed with the MTT chromatometry, theflow cytometry, and the immunohistochemistry. Results The primarily cultured MSCs grew as a colony on the walls of the culture bottle; after the culture for 5-7 days, the cells were shaped like the fibroblasts, the big flat polygonal cells, the medium sized polygonal cells, and the small triangle cells; after the culture for 12 days, the cells were found to be fused, spreadingall over the bottle bottom, but MSCs were unchanged too much in shape. After the induction by 5-azacytidine, some of the cells died, and the cells grew slowly. However, after the culture for 7 days, the cells grew remarkably, the cell volume increased gradually in a form of ellipse, fusiform or irregularity. After theculture for 14 days, the proliferated fusiform cells began to increase in a great amount. After the culture for 18-22 days, the myotubes increased in number and volume, with the nucleus increased in number, and the newly formed myotubes and the fusiform myoblst grew parallelly and separately. The immunohistochemistry for MSCs revealed that CD44 was positive in reaction, with the cytoplasm ina form of brown granules. And the nucleus had an obvious border，and CD34 was negative. The induced MSCs were found to be positive for desmin and specific myoglobulin of the skeletal muscle. The flow cytometry showed that most of the MSCs and the induced MSCs were in the stages of G0/G1，accounting for 79.4％ and 62.9％，respectively; however, the cells in the stages of G2/S accounted for 20.6％ and 36.1％. The growth curve was drawn based on MTT，which showed that MSCs weregreater in the growth speed than the induced MSCs. The two kinds of cells did not reach the platform stage，having a tendency to continuously proliferate.ConclusionIn vitro,the rat MSCs can be differentiated into the skeletal muscle cells with an induction by MyoD and 5-azacytidine, with a positive reaction for the desmin and the myoglobulin of the skeletal muscle. After the induction, the proliferation stage of MSCs can be increased, with a higher degree of the differentiation into the skeletal muscle.
ObjectiveTo understand the initiation, maturing, and resolution of thrombus by comparing the length and weight of thrombus at different time in the rat inferior vena cava (IVC) stenosis model.
MethodsForty-eight female Sprague Dawley rats, weighing 180-230 g, were selected to prepare the IVC stenosis model by blocking the most of the IVC blood flow. The length and weight of IVC thrombus were measured at different time points, the histological features were observed via HE staining.
ResultsBlood clots formed after 2 hours of modeling, the thrombus length and weight showed no significant difference between at 2 hours and 4 hours after modeling (P>0.05). The thrombus length and weight increased significantly at 6 hours, showing significant differences between at 2 and 4 hours and at 6, 8, 12, 24, and 48 hours (P<0.05); and from 6 to 48 hours, there was no significant difference in the thrombus length and weight (P>0.05), indicating that thrombus was stable, or maturing. Blood clots began to become smaller after 3 days when compared with ones at 48 hours (P<0.05), indicating the start of resolution at 3 days. At 7 days, the thrombus length and weight became further smaller when compared with ones at 3 days (P<0.05). The thrombus completely subsided at 21 days, the IVC recanalized. HE staining showed that thrombus formed after 2 hours of modeling; from 6 to 48 hours, the lumen became hyperemia, and the inflammatory cells, especially neutrophils, could be found. The organization of thrombus could be observed at 3 days and 7 days; thrombus gradually vanished at 21 days.
ConclusionThe time of thrombus initiation, maturing stage, and resolution stage is 6 hours, 6 to 48 hours, and 3 to 21 days after modeling, respectively.
Objective To explore the effect and mechanism of sleeve gastrectomy （SG） for type 2 diabetes mellitus （T2DM） in Goto-Kakizaki （GK） rats. Methods Thirteen male GK rats at 12 weeks of age were randomly divided into SG group （n=7） and sham operation group （SO group， n=6）， receiving SG surgery and sham operation respectively.Body weight， food intake in 24hours， fasting plasma glucose， plasma glucagon-like peptide-1 （GLP-1）， and plasma Ghrelin of rats in 2 groups were measured or tested before operation， 1， 4， 10， and 26 weeks after operation. In 10 weeks after operation， fecal energy content of rats in 2 groups was tested， in addition， oral glucose tolerance test （OGTT） and insulin tolerance test （ITT） were performed to investigate the glucose tolerance and insulin sensitivity. Results ①Body weight：there were no significant difference on body weight between the 2 groups （P＞0.05）. Compared with time point of before operation， the body weight of both 2 groups decreased in 1 week after operation （P＜0.01）， but increased in 10 weeks and 26 weeks （P＜0.01）. ②Food intake in 24 hours：compared with SO group， the food intake of SG group were lower in 4 weeks and 10 weeks after operation （P＜0.05）. Compared with time point of before operation， the food intake of SG group were lower in 1， 4， and 10 weeks after operation （P＜0.05）， but lower only in 1 week in SO group （P＜0.05）. ③Value of fasting glucose：compared with SO group， the value of fasting glucose in SG group were lower after operation （P＜0.01）. Compared with time point of before operation， the value of fasting glucose of SG group were lower after operation （P＜0.01）， but decreased in 1 week only in SO group （P＜0.01）. ④Level of serum GLP-1：compared with SO group， the levels of serum GLP-1 in SG group were higher in 4， 10， and 26 weeks after operation （P＜0.05）. Compared with time point of before operation， the levels of serum GLP-1 in SG group were higher in 4， 10， and 26 weeks after operation （P＜0.05）， but levels of serum GLP-1 in SO group didn’t change significantly （P＞0.05）. ⑤Level of serum Ghrelin：compared with SO group， the levels of serum Ghrelin in SG group were lower at alltime points after operation （P＜0.01）. Compared with time point of before operation， the levels of serum Ghrelin in SGgroup were lower at all time points after operation （P＜0.001）， but levels of serum Ghrelin in SO group didn’t change significantly （P＞0.05）. ⑥Areas under curves （AUC）：the AUC of OGTT and ITT test in SG group were both lower than those of SO group （P＜0.01）. Conclusion SG surgery can induce the level of fasting plasma glucose， and canimprove glucose tolerance and insulin sensitivity with significant changes of levels of plasma GLP-1 and Ghrelin， sugg-esting that SG surgery may be a potential strategy to treat patient with T2DM but without obesity or insulin resistance.
Objective To investigate the effect of the vascular endothelial growth factor (VEGF) gene therapy, the surgical delay, and the combination of the two therapeutic approaches on the survival of the rat over-area abdominal axial skin flap. Methods In 48 male Wistar rats (weight, 400-450 g), a model of the abdominal axial skin flap supplied by the superficial epigastric blood vessel was created. The rats were randomly divided into 6 groups: Group A (the blank group), Group B (the gene-therapy-during-operation group), Group C (the gene-therapy-before-operation group), Group D (themerely-surgical-delay group), Group E (the gene-therapy-during-surgical-delay group), and Group F (the gene-therapy-aftersurgical-delay group). Seven days after operation, the survival rate of the skin flap was measured; the specimens were harvested from the skin flap for a histological investigation of themicrovessels and for an immunohistochemical staining to observe the expression of VEGF165. Results The average survival rate of the skinflap was significantly greater in each of the treated groups than in Group A (Plt;0.05); the rate was the greater in Group E (Plt;0.05), but with no statistically significant difference between the other treated groups (Pgt;0.05). The average number of the microvessels was significantly greater in Groups B, C, E andF than in Groups A and D (Plt;0.05), but with no statistically significant difference between Groups B, C, E and F and between Groups A and D (Pgt;0.05). The lumen diameter of the microvessels was significantly greater in Group D than in Groups E and F (Plt;0.05), and the diameter was significantly greater in Groups D, E andF than in the other groups (Plt;0.05). More deposition of VEGF DNA detected by the immunohistochemical staining was in Groups B, C, E and F than in Groups A and D. There was no newly-formed blood vessel in the rat cornea in the treated groups.Conclusion Both the administration of pcDNA4-VEGF165 and the surgical delay can improve the survival of the rat abdominal axial skin flap, but the mechanism of the effect is different in explanation. The combination of the two therapeutic approaches can achieve a better effect.
