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Micro- and mini-bioreactors are characterized by their miniature working volume and comprehensive monitoring of process data, e.g., biomass, pH, dissolved oxygen, and fluorescence that are on par with conventional bench-top systems. The technical advancements of micro- and mini-bioreactors are supported by single-use material and micro-manufacturing, non-invasive optical sensors, automation such as industrial robotics and the integration of design of experiment software with data acquisition and process control. Owing to the miniature scales, micro-bioreactors typically feature lower turbulence intensity and energy dissipation rate, resulting in different mass transfer, mixing and shear conditions as compared to industrial scale equipment. Mini-bioreactors, nevertheless, are closer to large vessels. Micro- and mini-bioreactors are used mostly in screening and process development nowadays, owing to their combined high throughput and richness of data. They are also the hardware that will enable "precision medicine" in the near future.
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Biomass , Bioreactors , OxygenABSTRACT
As one of the top 10 breakthrough and emerging technologies in the world in 2018, cultured meat has attracted extensive attention due to its advantages of traceable origin, food safety and green sustainable development. Europe and the United States have invested a lot of resources to focus on research about cultured meat, which will affect our domestic meat and food market in the future. At present, the challenge of cultured meat production is how to efficiently simulate the growth environment of animal muscle tissue and realize large-scale production in bioreactor. Although cell tissue engineering has been deeply studied and achieved varying successful application, it is still difficult to obtain large-scale cultured meat production due to the high cost and technical requirements. Therefore, the development of efficient and safe cell culture technology is an urgent problem for large-scale cultured meat production, which can effectively reduce costs and achieve industrial application. In this review, we summarize the research progress of animal cell tissue culture technology used for cultured meat, and highlighted the current challenges and possible strategies in further applications.
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Animals , Bioreactors , Cell Culture Techniques , Meat , Tissue Engineering , United StatesABSTRACT
Industrial fermentation focuses on realizing the uniform of high titer, high yield, and high productivity. Multi-scale analysis and regulation, including molecule level, cell level, and bioreactor level, facilitate global optimization and dynamic balance of fermentation process, which determine high efficiency of biosynthesis, targeted directionality of bioconversion, process robustness, and well-organized system. In this review, we summariz and discuss advances in multi-scale analysis and regulation for fermentation process focusing on the following four aspects: 1) kinetic modeling of metabolic pathways, 2) characteristic of cell metabolism, 3) co-coupling fermentation and purification, and 4) bioreactor design. Integrating multi-scale analysis of fermentation process and integrating multi-scale regulation are expected as an important strategy for realizing highly efficient fermentation by industrial microorganisms.
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Bioreactors , Fermentation , Industrial Microbiology , Kinetics , Metabolic Networks and PathwaysABSTRACT
Objective To evaluate the clinical value of acetic acid with narrow-band imaging ( NBI ) and magnifying endoscopy ( ME ) on diagnosis of small colorectal polyps. Methods In this prospective study, 261 small colorectal polyps from 122 patients were observed by ME, NBI-ME, and acetic acid with NBI-ME, and then received endoscopic treatment. Endoscopic images were stored electronically and randomly allocated to 3 experts and 3 non-experts for diagnosis using Kudo pit pattern. The postoperative pathologic results acted as gold standard to evaluate the diagnostic accuracy of different endoscopic modes for small colorectal polyps. The image definition and interobserver agreement were compared among different endoscopic modes. Results The diagnostic accuracy of ME, NBI-ME, and acetic acid with NBI-ME for small colorectal polyps was 65. 5％ ( 171/261) , 90. 0％ ( 235/261) , and 94. 6％ ( 247/261) , respectively, in the experts group, and 57. 1％ ( 149/261) , 83. 1％ ( 217/261) , and 89. 3％ ( 233/261) , respectively, in the non-experts group. All experts and non-experts diagnosed small colorectal polyps more accurately by acetic acid with NBI-ME than by NBI-ME ( all P<0. 05 ) and ME ( all P<0. 001 ) . The image definition scores of acetic acid with NBI-ME in the experts group and non-experts group were significantly higher than those of NBI-ME and ME ( all P<0. 001) . The results of interobserver agreement showed that Kappa values (95％CI) of ME, NBI-ME, and acetic acid with NBI-ME diagnosis were 0. 578 (0. 508-0. 648), 0. 669 (0. 599-0. 739), and 0. 940 (0. 870-1. 010), respectively, for experts and 0. 476 (0. 406-0. 546), 0. 534 ( 0. 464-0. 604) , and 0. 830 ( 0. 760-0. 900 ) , respectively, for non-experts. Acetic acid with NBI-ME showed good interobserver agreement. Conclusion Acetic acid with NBI-ME has a higher diagnostic accuracy and good reproducibility for colorectal small polyps compared with ME and NBI-ME.
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Pancreas is an integrated organ with both endocrine and exocrine functions. Insulin is secreted from the pancreas islet B cell and is the only hormone that could reduce blood sugar and play an important role in glucose homeostasis. Pancreatic disease can result in diabetes mellitus in some cases. Although acute pancreatitis is the most common disease of pancreas,the relationship between acute pancreatitis and new-onset diabetes has been ignored. However,many researches have shown recently that acute pancreatitis is associated with new-onset diabetes. Blood glucose monitoring is warranted for patients after acute pancreatitis with important significance.
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This study was aimed to observe bronchial asthma patients in remission after treatment of traditional Chi-nese medicine(TCM) with four diagnostic information, syndrome differentiation and changes oflung function indicators, in order to explore the efficacy evaluation with TCM characteristics. TCM tongue manifestation instrument, TCM in-quiry scale, pulse-taking instrument, acoustic diagnostic information collection system, and spirometer were used in thecollection of 33 bronchial asthma cases in remission before and after treatment (1~5 months) basedon four diagnos-tic information and lung function indexes. Single-factor analysis of variance and other methods were used in the analysis of four diagnostic parameters and lung function indexes before and after treatment. The results showed that after TCM treatment, there were significant changes on indexes such as facial complexion, tongue sub-region color, color of tongue coating and other parameters. There was significant difference in the acoustic parameters before and after treatment. After TCM treatment, the frequency of lung system symptom such as nasal obstruction was decreased. There was no significant difference on changes of pulse-taking indexes among asthma patients before and after treat-ment. After treatment, the FEV1.0% of asthma patients was increased,whichindicated that asthma ventilatory function had been improved to some extent. It was concluded that TCM objective test provide an effective basis for the diag-nosis of bronchial asthma on aspects such as TCM syndrome, disease change observation and clinical evaluation.
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Objective To explore the best operation method in the patients with papillary thyroid microcarcinoma.Methods A total of 139 patients with papillary thjroid microcarcinoma were treated at our institute between Jan.2000 and Jan.2010.The data on the clinicopathological characteristics of patients and treatment were collected.Results The mean tumor size was (0.45 ± 0.24) cm.Of the 139 patients,30.2％ had multifocal tumors,19.4％ had bilateral tumors,42.4％ neck lymph node metastases.The number of eases of lymph node metastasis in level Ⅵ,Ⅱa,Ⅲ and Ⅳ was 58(41.7％),3(2.2％),10(7.9％),5(3.6％),respectively.Only one had lymph node metastasis in Level Ⅲ Microscopic extrathyroid extension was associated with neck lymph node metastases in papillary thyroid microcareinoma patients (x2 =38.39,P ＜0.001).No one developed permanent hypoparathyroidism.The median follow-up time was 103 (range,30-154) months,and local recurrence in the thyoid was diagnosed in 2 patients who underwent hemi-or subtotal thyroidectomy.Follow-up of 10 years was done in 16 cases,and the survival rate of 139 patients for 10-year was 100％.Conclusions It suggested that patients with papillary thyroid microcarcinoma has uniform clinicopathologic characteristics and the pattern of lymph node metastasis from those with papillary thyroid carcinoma.Total thyroidectomy plus level Ⅵ dissection is the optimal surgical treatment of papillary thyroid microcarcinoma.
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Objective To determine the clinical implications of internal mammary node biopsy for neoplasm stage,treatment,and prognosis in patients with breast cancer.Methods Internal mammary node biopsy via intercostal space was performed in 229 cases of breast cancer.Anatomical location of internal mammary nodes was recorded.Results Internal mammary node biopsy was successfully finished in 220 patients.There were 56 cases (24.45％ ) with internal mammary nodes metastasis,126 cases (55.02％ ) with axillary nodes metastasis,43 cases (34.13％ ) with regional metastases in both the axillary and internal mammary lymph nodes and 13 cases ( 12.62％ ) with internal mammary node metastasis only.Internal mammary node metastasis rate in patients with the number of positive axillary nodes ≥4 was 49.32％ (36/73).pN stage migration was seen in 56 patients with positive internal mammary nodes.There was no statistic relation between internal mammary nodes metastases and tumor location ( x2 =0.661,P =0.719).70.7％ patients with medial/central tumors and 50.7％ patients with the number of positive axillary nodes ≥4 were free from internal mammary node radiotherapy on account of internal mammary node biopsy.There was no complication such as pneumothorax or haemorrhagia.Conclusions Internal mammary node biopsy from intercostal space is a reliable surgical technique and can improve pN stage in some breast cancer patients.With internal mammary node biopsy,patients with a negative internal mammary node can be prevented from radiation to internal mammary nodal areas.
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ObjectiveTo evaluate the therapeutic effect of endoscopic full-thickness resection (EFTR) for gastric stromal tumors.MethodsA total of 33 patients with gastric stromal tumor orgination from deep muscularis propria layer received EFTR from January 2010 to July 2011.The effectiveness and safety of EFTR were compared with those of other 34 patients with gastric stromal tumor origination from muscularis propria layer who underwent endoscopic submucosal dissection (ESD).ResultsExcept in 2 patients with lesions larger than 3.0 × 3.0 cm,EFTR was successful in others 31 patients,who recovered well and had no recurrence during the follow-up within 12 months.There were no significant differences in resection rate,incidence of complications,body temperature,white blood cell counts or recovery time between 2 procedures (P ＞ 0.05 ).However,the number of clips used in EFTR ( 7.0 ± 3.5 vs.4.9 ± 3.1,P =0.013 ) and postoperative fasting days (3.4 ± 1.5 vs.2.0 ± 1.0,P =0.001 ) were significantly higher than those of ESD procedures.ConclusionEFTR is effective and safe for gastric stromal tumors with no higher risk than ESD,but it is more complex technically.EFTR can be used as an expanding method of ESD in endoscopic treatment of gastric stromal tumors.
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Objective To observe the effects of cinepazide maleate and nimodipine in improving the neurological function in patients with hypertensive cerebral hemorrhage after microtraumatic craniopuncture.Methods Seventy-eight patients with hypertensive cerebral hemorrhage were randomly divided into 2 groups,cinepazide maleate group (39 patients)and nimodipine group(39 patients).After 3 days operated with the microtraumatic craniopuncture,cinepazide maleate group used the amount 160mg cinepazide maleate mixed with sodium chloride injection(500ml,concentration 0.9%),and the nimodipine group uesd nimodipine(4mg)mixed with the same injection.Both the patients of the 2 groups were given intravenous drip once a day,then after continuous 14 days,the general information and the improvement of nerve were observeed.Results The total improvement rate and the improvement rate of nervous symptom was 87.2%and 61.5%respectively,in comparison,the nimodipine group was 64.1%and 39.9%.Conclusion Cinepazide maleate was better than nimodipine in improving chnical symptoms and the neurological deficit of the patients with hypertensive cerebral hemorrhage after microtraumatic craniopuncture.
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Objective To observe the effects of ganglioside and piracetam in improving the neurological function in patients with hypertensive cerebral hemorrhage.Methods Ninety-six patients with hypertensive cerebral hemorrhage Were randomly divided into 2 groups,ganglioside group(48 patients)and piracetam group(48 patients).Ganglioside group used the amount 40mg ganglioside mixed with sodium chloride injection(100ml,concentration 0.9%),and the piracetam group uesd piracetam(20g)mixed with the same injection.Both the patients of the 2 groups were given intravenous drip once a day,then after continuous 3 weeks,the general information and the improvement of nerve were observed.Results The effective rate and excellent rate of ganglioside group were remarkably higher than piracetam group,there was significant difference between the two groups(P＜0.05).Conclusion Ganglioside was better than pimcetam in improving clinical symptoms and the neurological deficit of the patients with hypertensive cerebral hemorrhage.
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Objective To investigate the regional distribution and morphological features of nesfatin-1/NUCB2 in digestive system of the humans, Sprague-Dawley (SD) rats and the institute of cancer research (ICR) mice, so as to lay the foundation for further study of its functions in the digestive system. Methods The specimens were obtained from SD rats and ICR mice as well as 20 patients with digestive disease, who were admitted to the First hospital affiliated to Nanjing Medical University and receired surgercal treatment. The specimens from patients with malignant tumors were obtained 5 cm apart from cancerous tissues and from patients with benign tumors were obtained near the focus. The resected tissues included pancreas, stomach, duodenum, esophagus, liver, small intestine or colon. The distribution of nesfatin-1/NUCB2 was examined with immunohistochemical (IHC) staining and its protein level in each organ was measured using Western blotting. Results The immuinohistocemical study revealed the similar distribution pattern of nesfatin-1/NUCB2 in the digestive system of the patients, SD rats and ICR mice. Nesfatin-1/NUCB2 was found to localize in the center of the pancreatic islets, the lower 1/3 to 1/2 of the gastric mucosal glands, as well as the submucosa of the duodenum. Western blotting examination showed the expression of NUCB2 in all tissues from patients, SD rats and ICR mice, whereas the protein level of the nesfatin-1/NUCB2 was higher in pancreas (0.84±0.03, 0. 84±0.05 and 0. 84±0.04, respectively), stomach (0.86±0.06,0.81±0.02 and 0. 78±0.02, respectively) or duodenum (0.79±0.09,0. 79±0.04 and 0.78±0.05)than that in esophagus (0.43±0.04,0.44 ± 0.02 and 0.47 ± 0.06, respectively), liver (0.42±0.01,0.44±0.04 and 0.43 ± 0.01, objectively), small intestine (0.32±0.04,0. 32 ± 0. 04 and 0.34 ±0.04, respectively) or colon (0. 29±0.01,0.32±0.03 and 0. 28±0.03, respectively)(all P values=0. 000). Conclusion Nesfatin-1/NUCB2 is widely expressed in the pancreatic islets, gastric mucosal glands and duodenum of the patients, SD rats and ICR mice, which indicates that nesfatin-1/NUCB2 may be involved in the regulation of food intake, carbohydrate metabolism and gastrointestinal motility.
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Objective To investigate the effect of different frequencies of endoscopic ultrasonography (EUS) on diagnosis of duodenal submucesal lesions. Methods A total of 210 elevated duodenal lesions detected by routine endoscopy in 201 patients were examined by EUS at different frequencies according to lesion features and image clarity. Endoscopic resection was applied in 27 lesions with informed consents,and pathological findings were compared with the results of EUS. Results The accuracy of EUS was 81.4% (22/27). Ultrasound of low frequency was superior in displaying overview of lesion and was more suitable for lesions larger than 1.5 cm in diameter or lesions of hyperecho. On the other hand,ultrasound of high frequency was more reliable to reveal minute structures and could be employed in scanning lesions smaller than 1.5 cm in diameter. Combination of different frequencies of ultrasound could provide more details in larger lesions. Conclusion Application of appropriate frequency or combined frequencies of ultrasound in examination of elevated duodenal lesions is beneficial to achieve more accurate diagnosis.
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Objective To investigate the effect of taurine on colonic fibrosis in rats with colitis induced by 2,4,6-trinitrobenzene sulphonic acid(TNBS). Methods Thirty-two SD rats were divided into normal control group, model group, low-dose (400 mg/kg) taurine group and high-dose (800 mg/kg) taurine group. Rats in normal group were administrated with 0.9% NaCl solution enema, and the other three groups received TNBS enema. The rats in low-dose and high-dose taurine groups were administrated with 400 mg/kg and 800 mg/kg of taurine daily, respectively, one week before TNBS enema. Morphology and disease activity index (DAI) were evaluated, and the colonic tissues were histologically examined. Colon length and weight of the rats were also measured. The concentrations of hydroxyproline, collagen type Ⅰ, transforming growth factor-betal(TGF-β1), and Smad3 protein and mRNA in colon tissues were tested. Results In comparison with control group, the body weight and colon length were decreased while DAI score and colon weight were increased obviously in model group (P`0.01). All above parameters were improved after intervention of taurine. The fibrotie score in model group (1.88±0.35) was significantly higher than that in control group (0.25±0.46), low-dose (1.25±0.71) and high-dose (0.75±0.47) taurine groups (all P values <0.05). High levels of hydroxyproline, collagen type Ⅰ, TGF-β1 and Smad3 were detected in model group compared with low-dose and high-dose taurine groups (all P values < 0.05). Conclusions Taurine is effective in prevention of colonic fibrosis induced by TNBS in rats, which is mediated by the down regulation of TGF-β1 and the inhibition of TGF-β/ Smad3 pathway. It may be beneficial in treatment of Crohn's disease with colonic fibrosis and strictures.
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Objective To estimate the effect and the intracellular signal transduction pathway of insulin-like growth factor I (IGF-I) on the expression of stem cell factor(SCF) in colonic smooth-muscle cells(SMC). Methods The SMCs isolated from colon of the SD rats by enzymolysis were cultured and identified by α-actin immunocytochemical method. Colonic SMCs were cultured either with 100 μg/L of IGF-I at different time points (0, 8, 16, 24 and 48 hours) or with different concentrations (0,5,10,50,100 and 150 μg/L) of IGF-I for 16 hours. The expressions of SCF in colonic SMCs pretreated with or without speicific phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor LY-294002 or mitogen-activated protein (MAP) kinase (MEK1) inhibitor PD-98059 were examined by Western blotting and quantitative reverse transeription-polymerase chain reaction and immunofluorescence. Results Low expression of SCF was found in colonic SMCs cultured in the bovine serum free medium. There was no effect of 5 or 10 μg/L of IGF- I on the expression of SCF. However, the expressions of SCF mRNA and protein were increased when stimulated with high concentrations (50,100,150 μg/L) of IGF-I. The peak expression of SCF was showed at the 16th hour after stimulating with 100 μg/L of IGF- I that was considered as the most effective concentration in vitro. The expression of SCF was not influenced by LY-294002, but was partly blocked by PD-98059. Conclusions The expression of SCF in colon SMCs may be induced by IGF-Ⅰ through MAP kinase signaling pathway.
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Objective To analyze clinical characteristics of gastroesophageal reflux disease(GERD) in aged patients for improvement of diagnosis and treatemcnt. Methods The reflux disease questionnaire was performed in patients diagnosed as GERD based on Montreal definition and classification as well as Rome Ⅲ criteria.All patients were divided into elderly group (≥65 years) and control group(＜65 years). The incidence of hita[ hernia (HH), the frequencies of esophagitis (based on Los Angeles classification), clinical features, and quality of life were compared between two groups. Results There was no difference between two groups in male/female ratio and morbidity of HH(P＞0.05). In comparison with control group, the frequency of esophagitis graded as LC or LD increased and extra-esophageal symptoms were higher in elderly group (P＜ 0.05), but the lower typical symptoms (heartburn and regurgitation) were seen in the elderly group(P＜0.05). The scores of role physical, bodily pain and role emotional were higher in elderly group than those in control group (P＜0.05). There was no significant differences between two groups in physical function, vitality,social functioning, mental health, and general health. Conclusion The elderly GERD patients often have lower score of typical reflux symptoms (heartburn and regurgitation) and high incidence of severer esophagitis, but their quality of life is not significantly influenced.
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Objective To explore the pattern of cervical nodal metastasis and the clinical significance of total thyroidectomy plus functional neck lymphadenectomy in papillary thyroid carcinoma patients. Methods Clinical and pathological data of 172 patients with papillary thyroid cancer who underwent total thyroidectomy plus functional neck lymph node dissection at Jinan Military General Hospital were retrospectively reviewed, including patient demographics, extent of surgery, parathyroid hormone level,recurrence, tumor pathology, such as tumor size, multifocality, capsular invasion, vascular invasion,extrathyroidal extension, and lymph node status. Results Of the 172 functional neck dissection patients (47 ambilateral), the incidence of lymphonodus metastasis in regions Ⅵ, Ⅳ and Ⅲ was 96. 3% ,78. 5%and 62.1% respectively. Rate of nodal metastasis was higher in patients with extracapsular invasion than in patients with no invasion ( P ＜ 0. 05 ). Serum parathyroid hormone levels significantly decreased immediately postoperatively in total thyroidectomy plus functional neck dissection and remained low for several weeks thereafter ( P ＜ 0. 01 ). The 5-, 10- and 15-year survival rate was ( 98. 83 ± 0. 82) %, (98. 23 ± 1.02 ) % and (96. 42 ± 1.43 )%, respectively. Conclusions Therapeutic neck lymph node dissection for papillary thyroid cancer is recommended for cervical nodal metastasis patients. Total thyroidectomy plus functional neck dissection is important in the treatment of papillary thyroid cancer.
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Objective To establish ulcerative colitis(UC)model in BALB/c mice and to investigate the role of high mobility group box chromosomal protein 1(HMGBI)in pathogenesis of UC.Methods Thirty-two BALB/c mice were randomly divided into UC group(n=24,which were fed with 3%dextran sulfate 80dium solution)and control group(n=8,which were fed with water).The animals were sacrificed at 24.96 and 1 68 hours,respectively,to collect samples of colon and blood.The sernm level of HMGB1 was measured with ELISA and the expression of HMGB1 in colon was determined by Western blotting analysis.Results Histological scoring increased with the induction of the model,and manifestation of colon mucosa at 168h was similar with that of UC in human.The serum level of HMGB1 was slightly higer at 24 h than that of control(5.09±0.61 μg/L vs 4.49±0.53μg/L,P＞0.05),and reached a peak at 96 h (14.74±0.60 μg/L,P＜0.01),decreased at 168 h(9.03±0.78μg/L,P＜0.01).The expression 0.05).significandy increased at 96h(0.76±0.03,P＜0.05)and at 168 h(0.77±0.04,P＜0.05).Conclusion HMGB1 might be involved in pathologic changes of UC at a later stage.
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Objective To assess the effects of zinc protoporphyrin (ZnPP), an inhibitor of the heme oxygenase (HO), on the colonic interstitial cells of Cajal (ICC) of diabetic rats with colonic slow transit. Methods Diabetes mellitus (DM) model was induced by intraperitoneal injection of streptozotocin (STZ) in Sprague-Dawley rats. Twenty four successfully established DM rats were selected, and 16 healthy rats were served as controls. Six weeks later, gastrointestional (GI) dysfunction was observed by charcoal propulsion experiment in 8 DM rats and 8 controls. The rest rats in DM group were divided into 2 groups: DM rats intraperitoneal injected with PBS (n=8) or with 10 μmol/kg of ZnPP (n = 8) every other day for 3 weeks. The rats in control group (n = 8) were intraperitoneally injected with PBS. The levels of HO and c-kit (the special receptor of ICC) expression were detected by Western blotting. The distribution of ICC was observed by immunohistochemistry and the area of c-kit positive cells was counted. Results The GI propulsion rate in DM rats interfered with PBS was significantly declined compared to that in the controls (63.0%± 1.2% vs 71.8%±2.0%, P<0.05). But it was improved in DM rats interfered with ZnPP (72.5± 2.6%, P<0.05), which showed no significant differentee with that in control group (P>0.05). The expression of HO-1 in close and distant colon of DM rats interfered with ZnPP was decreased (P< 0.05). The expression of HO-2 in close colon and the area of c-kit positive cells of DM rats interfered with PBS was reduced compared with that in controls (P<0.05), but both were improved in DM rats interfered with ZnPP (P<0. 05). Conclusion Administration of ZnPP might be able to protect ICC by its blockage of HO-1 in DM rats with gastrointestinal dysfunction.
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Objective To investigate the effect of exogenous insulin-like growth factor-1(IGF-1)on growth of gastric smooth muscle cells(SMCs)and synthesis of stem cell factor(SCF).Methods Gastric SMCs were cultured by enzymolysis and were identified by α-actin immunocytochemical method.After that,SMCs were incubated with different concentrations of IGF-1(0,50,100,150μg/L)or IGF-1 receptor monoclonal antibody(0.50,100,150μg/L)for 24 hours.Proliferation of SMCs was measured by MTT.The expressions of SCF protein and RNA were analyzed by Western blotting and quantitative reverse transcription-polymerase chain reaction,respectively.The SCF content in the culture fluid was detected by enzyme-linked immunosorbent assay(ELISA).Results The proliferation of gastric SMCs and the expression of SCF can be enhanced by IGF-1,and 100μg/L of IGF-1 may be the final effective concentration.Nevertheless,these could be inhibited by IGF-1 receptor monoclonal antibody in a dose-dependent manner.ConclusionElevated expression of SCF may be induced by IGF-1 through proliferation of SMC.