Objective To establish a purified model of rat retinal ganglion cells （RGCs） cultured by serum-free medium，and provide a good cell model to investigate the damage of RGCs in glaucoma，retinal ischemia，and degenerative retinopathy. Methods Two monoclonal antibodies，anti-rat SIRP（OX-41）against rat macrophage and antibody against rat Thy-1（OX-7），were used to purify and characterize RGCs from 1-3-day old Sprague-Dawley（SD）rats by means of two-step filtration.Purified RGCs were cultured in serum-free neurobasal medium containing B27 and ciliary neurotrophic factor（CNTF） meeting the neuronal cellrsquo;s special requirements.Photomicrographs illustration，immunfluorescence staining of Thy-1，calcein-acetoxymethyl ester（calcein-AM）fluorescence images were used to observe and identify cultured retinal cells and purified RGCs. Results Among the primary cultured rat retinal cells，91% were retinal neurons.Protuberances of RGCs were seen after cultured for 24 hours.At the4th to 8th day，many cells had uniform configuration，large body，and long protuberances. At the 14th day，over 60% cells maintained viability.Immunoflurescence staining of Thy-1 showed the purity of RGCs was about 90%. The results of calcein-AM staining，which stained the living cells only，showed large cell body of RGCs and most of RGCs had a protuberance whose length was twice longer than the diameter of the cells. Conclusion RGCs cultured by serum-free medium has uniform size，good configuration，and high purity，which is adapt to the research of damage of RGCs caused by various factors and to evaluate the protective effects of neuroprotective agents. (Chin J Ocul Fundus Dis, 2006, 22: 200-203)
ObjectiveTo investigate the effect of serum on the differentiation of neural stem cells.MethodsThe neural stem cells were isolated from the embryonic hippocampus tissues of Sprague Dawley rats at 14 day of pregnancy. After culturing and passaging, the 3rd generation cells were identified by immunocytochemical staining. Then, the cells were divided into 3 groups according to the concentrations of fetal bovine serum (FBS) used in the differentiation cell culture medium: 5% (group A), 1% (group B), 0 (group C), respectively. The other components of the culture media in 3 groups were the same. Cell viability was determined by using the Live/Dead cell staining at 8 days; the expressions of glial cell marker [glial fibrillary acidic protein (GFAP)] and neuronal marker (β-Ⅲ Tubulin) were determined and analyzed by immunocytochemical staining and real-time fluorescent PCR at 4 and 8 days of culture.ResultsBased on cell morphology and immunocytochemical staining, neural stem cells were identified. Cells were growing well with no death in all groups. With decreasing FBS concentration, the expression of GFAP was significantly decreased on both protein and mRNA level, whereas the expression of β-Ⅲ Tubulin was evidently increased. The staining of each group at 8 days was more obvious than that at 4 days. There were significant differences in mRNA expressions of GFAP and β-Ⅲ Tubulin at 4 and 8 days between groups (P<0.05).ConclusionSerum can promote the differentiation of neural stem cells into glial cells. At the same time, it inhibits the differentiation of neural stem cells into neurons, the lower the serum concentration, the smaller the effect.
Objective To investigate the possibility of culturing human oral keratinocyte using autologous serum in order to provide theoretical and technical foundation for clinical application of tissue engineering oral mucosa epithelium.Methods The human oral keratinocytes were cultured by the medium containing different concentrations of autologous serum(10%,20%,30%)and fetalbovine serum (10%), respectively. The growth conditions for the cell and the mucosa epithelium in the groups were observed, the cell growth curves were drawn, and the population doubling time (PDT) was counted. Results The results showed that the human oral keratinocyte could proliferate well in the medium containing autologous serum or fetal bovine serum. The differences in the 24hour clone rate and PDT were not significant. Both the area and the thickness of the cultured oral epithelium increased with the increase of the autologous serum concentration, and the difference between autologous serum and fetal bovine serum was significant, especially with the medium containing 20% autologous serum( P＜0.05) . The human nature of the cultured epithelium was demonstrated by the immunofluorescent mouse anti-HLA antigen. Conclusion The autologous serum can replace the fetal bovine serum to culture the oral keratinocyte well, and the cultured oral mucosa epithelium can be better differentiated in the autologous serum than in the fetal bovine serum.
Objective To prepare the immunonanospheres［SC3Ab-HSA(5-Fu)-NS］ against human colorectal cancer and evaluate its immunoreactivity and effects on cancer. Methods SC3Ab-HSA(5-Fu)-NS was prepared by intermolecular cross-linking the monoclonal antibody SC3Ab with human serum albumin nanospheres containing 5-Fu ［HAS(5-Fu)-NS］ via new hetero-bifunctional crosslinker SPDP. Condensation test and immunoflurecence were used to evaluate the immunoreactivity, the specific binding of SC3Ab-HSA(5-Fu)-NS with colorectal cancer cell line SW1116 was observed by microscope and electron microscope. The specific cytotoxic effects on target cells were evaluated in vitro by MTT assay. SC3AbHSA(5-Fu)-NS, HSA(5-Fu)-NS and 5-Fu were injected into nude mice bearing human colorectal carcinoma, to study the inhibitory activity of SC3Ab-HSA(5-Fu)-NS in vivo. Results The immunoreactivity of SC3Ab-HSA(5-Fu)-NS was well preserved. SC3Ab-HSA(5-Fu)-NS can bind the SW1116 cells specifically. The IC50 value for SC3Ab-HSA(5-Fu)-NS on SW1116 cells was 24.6 μg/ml,which was lower than that of HSA(5-Fu)-NS(345.3 μg/ml) and 5-Fu(325.6 μg/ml). The inhibitory rate of SC3Ab-HSA(5-Fu)-NS on the growth of colorectal cancer xenografts was significantly higher than that of HSA(5-Fu)-NS or 5-Fu(P＜0.001).Conclusion SC3Ab-HSA(5-Fu)-NS has immunoreactivity and specific active targeting to the colorectal cancer cells. The anticancer ability of SC3Ab-HSA(5-Fu)-NS is significantly higher than that of HSA(5-Fu)-NS and 5-Fu.
摘要:目的: 探讨64排多层螺旋CT(MSCT)和血清淀粉样蛋白A(serum amyloid A protein, SAA)联合术前评估直肠癌在肿瘤分期诊断中的作用。 方法 :纳入经根治术治疗的直肠癌患者通过MSCT扫描进行评估,同时取患者静脉血测量术前SAA水平,行MSCT分期与MSCT和SAA联合分期以比较二者的诊断价值。 结果 :本研究纳入患者121例。MSCT检测T分期的准确度为851%。在评估淋巴结转移方面,MSCT和SAA联合分期的准确度为760%,明显高于MSCT分期(595%, 〖WTBX〗P lt;0001)。MSCT正确判断所有远处转移。同单一的MSCT检测相比,MSCT和SAA联合评估能显著的提高术前TNM分期的准确率(785% vs. 636%,〖WTBX〗P =0011)。 结论 :MSCT联合SAA检测比单一的MSCT检测显著提高了直肠癌术前肿瘤分期和淋巴结转移方面的准确度。这种新的术前评估方法的为肿瘤进展评估和术前治疗决策提供了更加可靠的信息。Abstract: Objective: To determine the role of combinative assessment of 64 multislice spiral computer tomography (MSCT) and serum amyloid A protein (SAA) in preoperative rectal cancer staging. Methods : Enrolled consecutive rectal cancer patients undergoing curative surgery were evaluated by MSCT scan. Meanwhile venous blood specimens were taken to measure preoperative SAA concentration. Both MSCT staging and MSCT plus SAA staging were performed to compare with each other. Results : The study population consisted of 121 patients. The accuracy of T staging was 851% for MSCT. The accuracy in evaluating lymph nodes metastases was 760% for MSCT plus SAA compared with 595% for MSCT alone (〖WTBX〗P lt;0001). All the distant metastases were correctly detected by MSCT. The method combining MSCT with SAA led to significant improvement on preoperative TNM staging compared with MSCT alone (785% vs. 636%, 〖WTBX〗P =0011). Conclusion : MSCT plus SAA showed greater accuracy than MSCT alone in rectal cancer staging and lymph node metastases. This novel strategy of preoperative evaluation appears to provide more accurate information on tumor progression and preoperative therapy decisionmaking.
ObjectiveTo explore the effects of glycemia and serum calcium on occurrence and development of aortic root dilation disease. MethodsThe clinical data of patients with aortic root dilation who underwent surgical treatment in the Department of Cardiac Surgery of the First Affiliated Hospital of Xinjiang Medical University from January 2011 to October 2021 were retrospectively collected. They were divided into two groups according to whether they were accompanied by acute aortic dissection (Stanford type A), and were matched with the propensity scoring method. Logistic univariate and multivariate regression analyses were used to analyze the glycemia and the serum calcium of the patients in 24 hours at admission, and their receiver operating characteristic (ROC) curves were plotted. Results Finally 184 pairs of patients were matched, including 297 males with an average age of 48.76±9.62 years and 71 females with an average age of 49.97±10.97 years. There were statistical differences in ethnicity, history of hypertension, aortic root diameter, serum calcium and glycemia between the two groups (P<0.05). Logistic multivariate regression analyses results showed that age＜40 years (OR=4.106, P=0.010), Han nationality (OR=2.863, P<0.001), aortic root diameter＜45 mm (OR=5.063, P<0.001), hypertension (OR=2.736, P=0.001), hyperglycemia (OR=4.426, P<0.001) and hypocalcemia (OR=5.375, P<0.001) were independent risk factors for aortic root dilation disease with dissection. ROC curve analysis suggested that the area under the curve (AUC) of glycemia was 0.742 and the AUC of serum calcium was 0.737, all of which had some predictive value. Conclusion Hyperglycemia and hypocalcemia are risk factors for the development of aortic root dilation disease, and to some extent, they can be used as indicators for screening high-risk patients with aortic root dilation disease.
Objective To investigate whether recombinant human serum albumin (rHSA) can replace traditional B27 as a basic medium for differentiation of human pluripotent stem cells (hPSCs) into cardiomyocytes. Methods hPSCs were seeded at a cell density of 1.2×104/cm2; until up to 75% confluency hPSCs were induced by differentiation medium containing various concentration of rHSA (0, 50, 100, 200 g/L). Light microscope and fluorescence microscope recorded the whole process of stem cells differentiating into myocardium. Flow cytometry was used to detect the cardiac differentiation efficiency at different concentrations of rHSA. Immunofluorescence staining was used to detect the cardiac specific protein α-actinin and troponin T (cTnT) and electron microscope to observe the ultrastructure of human pluripotent stem cell-derived cardiomyocytes (hPSC-CM) and beating rates of hPSC-CMs response to drugs. Results A large number of spontaneous beating cardiomyocytes were observed 9 days after induction and differentition. The percentage of colonies showing beating cardiomyocytes was 60.4% at the concentration of 200 g/L of rice derived-rHSA. Beating cardiomyocytes were α-actinin and cTnT positive. Ultrastructural analysis showed scattered sarcomeres and mitochondrial. hPSC-CMs were dose-dependent on isopropyl adrenaline and verapamil. Conclusion Using such simple media to differentiate hPSCs into functional cardiomyocytes is cost-effective and highly efficient, and can be used in the clinical research.
Hypernatremia is one of the commonly syndromes in critically ill patients. Severe hypernatremia has a low incidence (0.6%–1.0%) but with a very high mortality (58%–87%). Conventional treatments include the limitation of sodium intake and the supplement of sodium free liquid according to the assessed water lost. The reduction rates of conventional treatments are commonly not effective enough to decrease the serum sodium concentration in severe euvolemic or hypervolemic hypernatremia patients. Continuous renal replacement therapy (CRRT) has been reported to be effective on the reduction of sodium level in severe hypernatremia patients. However, the evidences on the use of CRRT for hypernatremia are limited. Our present review summarizes the current evidences on the prevalence of hypernatremia, the outcome of hypernatremia patients, the conventional treatment of hypernatremia, and the advantages and indications of CRRT for the management of hypernatremia. Additionally, we introduce our experiences on the management of hypernatremia using CRRT as well.
Objective To investigate the effects of a mixed bacterial lysate (OM-85 BV) on lung function and serum IgE in asthmatic mice under acute attack, and to explore the therapeutic effect of OM-85 BV on acute attack and the application value of OM-85 BV in non-acute attack. Methods A total of 30 SPF Kunming mice aged 4 to 6 weeks were randomly divided into 3 groups, namely a blank control group (Group A), an asthma model group (Group B), and an OM-85 BV intervention group (Group C). The mice in groups B and C were sensitized by intraperitoneal injection of ovalbumin on day 1, 8 and 15, respectively. From day 22, the asthma model was stimulated by inhalation of 5% ovalbumin every day for 30 min for 5 consecutive days. The mice in group C were treated with OM-85 BV dissolved in normal saline from day 1, and each mouse was gavaged continuously for 10 days. The intraperitoneal injection, intragastric administration and aerosol inhalation reagent of mice in group A were all replaced by normal saline, while the intragastric administration of mice in group B was replaced by normal saline. One hour after the last stimulation, the mice were anesthetized, their lung function was measured, blood was collected from the eyeballs and then they were sacrificed, and the blood was centrifuged and the serum was separated and stored. Hematoxylin and eosin staining was used for pathological examination. Serum IgE was determined by enzyme-linked immunosorbent assay. Results Compared with group A, forced vital capacity in 0.15 second (FEV0.15), FEV0.15/forced vital capacity (FVC) and peak expiratory flow (PEF) of mice in group B were significantly decreased. The lung function of group C was improved compared with group B. In group B, the pathological manifestations were dysplasia and collapse of bronchial epithelium, infiltration of inflammatory cells, mainly lymphocytes and eosinophils, a small amount of mucus and shed epithelial cells in the tracheal lumen, and significant thickening of airway wall. The asthma mouse model was well established. The pathological manifestations of airway in group C were less severe than those in group B, the thickness of airway wall was reduced, and the inflammatory cells were also significantly reduced. The serum IgE concentration in groups B and C increased, and the IgE level in group C decreased significantly compared with group B. The differences were statistically significant (all P<0.05). Conclusions Exogenous administration of OM-85 BV in asthmatic mice can effectively reduce the concentration of serum IgE, alleviate airway inflammation, reduce eosinophil infiltration, and improve the pulmonary function performance of asthmatic mice during acute attack, showing that FEV0.15/FVC, FEV0.15 and PEF indicators are significantly improved. OM-85 BV can alleviate the symptoms of bronchial asthma in the acute attack of mice, improve the physiological function of the lung during the acute attack, inhibit airway inflammation, and have certain application value in the stable asthma control.
Objective The method of metabonomics based on nuclear magnetic resonance (NMR) imaging was used to explore the difference in metabolites of serum and bile, and to analyze the metabolic variation related to the pathogenesis of gallbladder stones between normal people/liver transplantation donors and patients with gallbladder stones. Methods Prospectively collected the serum samples (17 cases) and bile samples (19 cases) in 19 patients with gallbladder stones who underwent surgery in West China Hospital form March 2016 to December 2016, as well as the serum samples of 10 healthy persons and the bile samples of 15 liver transplantation donors at the same time period. The differences of metabolites in the blood and bile in these 3 groups were compared by using 1H-NMR metabonomics technology and chemometric methods. Results The concentrations of valine, alanine, lysine, glutamine, glutamate, pyruvate, creatinine, choline, alpha-glucose, beta-glucose, tyrosine, histidine, and hypoxanthine in serum of patients with gallbladder stones decreased significantly, comparing with those of healthy people without gallbladder stones (P<0.05), while 1, 2-propanediol, acetoacetate, and lactate increased significantly in the serum of patients with gallbladder stones (P<0.05). The concentrations of taurine conjugated bile acids, glycine conjugated bile acids, choline, and phosphatidylcholine decreased significantly in the bile of patients with gallbladder stones when compared with those of liver transplantation donors (P<0.05), while cholesterol increased significantly in the bile of patients with gallbladder stones (P<0.05). Conclusions There are significant differences of the serum and bile metabolites between patients with gallbladder stones and healthy men without gallbladder stones/liver transplantation donors. 1H-NMR metabonomics is helpful to investigate the pathogenesis of gallbladder stones.