Objective To optimize the preparation process of hydroxysafflor yellow A(HSYA)nanoparticle and conduct in vitro release evaluation.Methods HSYA nanoparticles were prepared with PLGA as carrier by modified compound emulsion method.The optimal preparation process of the experiment was selected by Plackett-Burman and Box-Behnken response surface method.The nanoparticles were characterized by using particle size analyzer,TEM scanning electron microscope,Fourier transform infrared spectroscopy(FT-IR),X-ray diffraction(XRD).Frozen(4℃)storage stability,stability in physiological medium,lyophilized protective agent and in vitro release rate were investigated.Results The optimal process prescription of nanoparticle is as follow:pH value is 6.95,the dosage is 2.8 mg,and carrier dosage is 18.2 mg.The size of nanoparticles obtained at optimum condition is(176.4±1.29)nm,the polydiseperse index(PDI)is 0.152±0.014,the Zeta potential is(-17.6±0.46)mV,the encapsulation rate is(78.5±0.49)％,drug loading is(7.3±0.07)％.These nanoparticles showed round and good dispersion.Good stability in 4℃ storage environment and different physiological media of nanoparticles were observed.The best lyophilized protective agent was 1％glucose and the in vitro release rate of nanoparticles at 48 hours was 85％.Conclusion The optimization method is reasonable and reliable.The obtained nanoparticles have good stability and sustained-release effect.The in vitro release behavior conformed to first-order kinetic model.

Objective: To explore the effects of porcine acellular dermal matrix (ADM) combined with human epidermal stem cells (ESCs) on wound healing of full-thickness skin defect in nude mice. Methods: The morphology of porcine ADM was analyzed by photograph of digital camera, the cell residues in porcine ADM were observed by hematoxylin-eosin (HE) staining, the surface structure of porcine ADM was observed by scanning electron microscope, the secondary structure of porcine ADM was analyzed by infrared spectrometer, the porcine ADM particle size was analyzed by dynamic light scattering particle size analyzer, and the porcine ADM potential was analyzed by nano-particle size potentiometer. The morphology of porcine ADM was observed by inverted fluorescence microscope when it was placed in culture medium for 30 min, 1 d, and 5 d (n=2). The porcine ADM was divided into 5 min group, 10 min group, 20 min group, 30 min group, 60 min group, and 120 min group according to the random number table (the same grouping method below) in static state at normal temperature for the corresponding time to calculate the water absorption by weighing method (n=3). Swiss white mouse embryonic fibroblasts (Fbs) were divided into blank control group (culture medium only), and 50.0 g/L ADM extract group, 37.5 g/L ADM extract group, 25.0 g/L ADM extract group, 12.5 g/L ADM extract group, and 6.5 g/L ADM extract group which were added with the corresponding final concentrations of ADM extract respectively. At post culture hour (PCH) 24, 48, and 72, the cell survival rate was detected by cell counting kit 8 and the cytotoxicity was graded (n=5). The erythrocytes of a 6-week-old male Sprague-Dawley male rat were divided into normal saline group, ultra-pure water group, and 5 mg/mL ADM extract group, 10 mg/mL ADM extract group, and 15 mg/mL ADM extract group which were treated with the corresponding final concentrations of porcine ADM extract respectively. After reaction for 3 h, the absorbance value of hemoglobin was detected by microplate reader to represent the blood compatibility of porcine ADM (n=3). ESCs were isolated and cultured from the discarded prepuce of a 6-year-old healthy boy who was treated in the Department of Urology of the First Affiliated Hospital of Army Medical University (the Third Military Medical University) in July 2020, and then identified by flow cytometry. The porcine ADM particles of composite ESC (hereinafter referred to as ESC/ADM) were constructed by mixed culture. After 3 days of culture, the composite effect of ESC/ADM was observed by HE staining and laser scanning confocal microscope. Thirty-six 7-8-week-old male non-thymic nude mice were divided into phosphate buffer solution (PBS) alone group, ADM alone group, ESC alone group, and ESC/ADM group, with 9 mice in each group, and the wound model of full-thickness skin defect was established. Immediately after injury, the wounds were treated with the corresponding reagents at one time. On post injury day (PID) 1, 7, 11, and 15, the wound healing was observed and the wound healing rate was counted (n=3). On PID 7, the epithelialization of wounds was observed by HE staining and the length of un-epithelialized wound was measured (with this and the following sample numbers of 4). On PID 11, the dermal area and collagen deposition of wounds were observed by Masson staining and the dermal area of wound section was calculated, the number of cells expressing CD49f, a specific marker of ESC, was calculated with immunofluorescence staining, the mRNA expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in ESC after wound transplantation was detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction. Data were statistically analyzed with independent sample t test, one-way analysis of variance, analysis of variance for repeated measurement, and least significant difference t test. Results: The porcine ADM was white particles and composed of reticular structure, with no cells inside, disordered structure, and rough surface. The absorption peak of porcine ADM appeared at the wave numbers of 1 659, 1 549, and 1 239 cm^-1, respectively. The main particle size distribution of porcine ADM in solution was 500 to 700 nm, with negative charge on the surface. The morphology of porcine ADM in static state at 30 min and on 1 and 5 d was relatively stable. The water absorption of porcine ADM remained relatively high level in static state from 30 min to 120 min. The cytotoxicity of mouse embryonic Fbs in 6.5 g/L ADM extract group, 12.5 g/L ADM extract group, and 25.0 g/L ADM extract group was grade 1 at PCH 24, and the cytotoxicity of the other groups was 0 grade at each time point. After reaction for 3 h, the absorbance value of hemoglobin of erythrocytes in ultra-pure water group was significantly higher than the values in normal saline group and 15 mg/mL ADM extract group (with t values of 8.14 and 7.96, respectively, P<0.01). After 3 days of culture, the cells of the fourth passage showed pebble-like morphology, with low expression of CD71 and high expression of CD49f, which were identified as ESCs. There was ESC attachment and growth on porcine ADM particles. On PID 1, the wound sizes of nude mice were almost the same in PBS alone group, ADM alone group, ESC alone group, and ESC/ADM group. On PID 7, 11, and 15, the wound contraction of nude mice in each group was observed, especially in ADM alone group, ESC alone group, and ESC/ADM group. On PID 7, the wound healing rates of nude mice in ESC alone group and ESC/ADM group were significantly higher than the rate in PBS alone group (with t values of 2.83 and 4.72 respectively, P<0.05 or P<0.01). On PID 11, the wound healing rate of nude mice in ESC/ADM group was significantly higher than that in PBS alone group (t=4.86, P<0.01). On PID 15, the wound healing rates of nude mice in ADM alone group, ESC alone group, and ESC/ADM group were significantly higher than the rate in PBS alone group (with t values of 2.71, 2.90, and 3.23 respectively, P<0.05). On PID 7, the length of un-epithelialized wound of nude mice in ADM alone group, ESC alone group, and ESC/ADM group was (816±85), (635±66), and (163±32) μm, respectively, which were significantly shorter than (1 199±43) μm in PBS alone group (with t values of 5.69, 10.19, and 27.54 respectively, P<0.01). On PID 11, the dermal areas of wound section of nude mice in ADM alone group, ESC alone group, and ESC/ADM group were significantly larger than the area in PBS alone group (with t values of 27.14, 5.29, and 15.90 respectively, P<0.01); the collagen production of nude mice in ADM alone group and ESC/ADM group was more obvious than that in PBS alone group, and the collagen production of nude mice in ESC alone group and PBS alone group was similar. On PID 11, in the wounds of nude mice in ESC alone group and ESC/ADM group, the cells with positive expression of CD49f were respectively 135±7 and 185±15, and the mRNA expressions of GAPDH were positive; while there were no expressions of CD49f nor mRNA of GAPDH in the wounds of nude mice in PBS alone group and ADM alone group. Conclusions: ESC/ADM particles can promote the wound healing of full-thickness skin defects in nude mice, which may be related to the improved survival rate of ESCs after transplantation and the promotion of dermal structure rearrangement and angiogenesis by ADM.
Acellular Dermis ; Animals ; Fibroblasts ; Humans ; Male ; Mice ; Mice, Nude ; Rats ; Rats, Sprague-Dawley ; Stem Cells ; Swine ; Wound Healing

Objective: The survival rate of gastric cancer cases in the whole population of Qidong City of Jiangsu Province from 1972 to 2016 was analyzed to provide a data basis for prevention and control of gastric cancer. Methods: The observed survival rate (OSR) was calculated by the life table method, the relative survival rate (RSR) was calculated by SURV 3.01 software, and the trend test was performed by Hakulinen's likelihood ratio test. Age-standardized relative survival rate (ARSR) was calculated using the International Cancer Survival Standard (ICSS). The Joinpoint regression model was used to calculate the average annual percentage change (AAPC) of survival rate, and the trend analysis of the overall survival rate of gastric cancer and the survival rates of different sexes and ages was conducted. The ARIMA model was used to predict the survival rate. Results: From 1972 to 2016, there were 18 067 new cases of gastric cancer in Qidong City. The OSRs at 1, 5, and 10 years were 37.12%, 17.16%, and 12.86%, respectively, and the RSRs at 1, 5, and 10 years were 38.83%, 21.73%, and 21.20%. Dividing 1972-2016 into 9 periods at 5-year intervals, compared with 1972-1976, the 5-year RSR in 2012-2016 increased from 12.53% to 36.26%, and the RSR in the 9 periods increased with statistical significance (χ(2)=747.19, P<0.001). Both OSR and RSR were slightly higher in men than in women, and decreased with age. The AAPC results showed that the AAPC of the 5-year RSR of gastric cancer in Qidong was 2.88%, 2.75% in males and 3.05% in females, and the upward trend was statistically significant (P<0.001). From 1972 to 2016, the 5-year RSR of gastric cancer in all age groups showed an upward trend. Except for the 35-44-year-old group (AAPC=1.07%, t=2.16, P=0.067), the age groups had statistical significance ( AAPC=2.59% in the 45-54-year-old group, AAPC=3.19% in the AAPCs of other 55-64-year-old group, AAPC=3.99% in the 65-74-year-old group, and AAPC=2.97% in the 75-year-old group, all P<0.05). The prediction results of the ARIMA model showed that in 2026, the 5-year OSR of gastric cancer in Qidong will increase to 39.82%, and the 5-year RSR will increase to 49.23%. Conclusion: The overall survival rate of registered gastric cancer cases in Qidong has been greatly improved, but there is still large room for improvement. More attention should be paid to the prevention and treatment of gastric cancer.
Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; China/epidemiology* ; Incidence ; Software ; Stomach Neoplasms ; Survival Rate

Objective:To evaluate the immunogenicity of recombinant factor H binding protein(fHBP) by detecting serum antibody titer and serum bactericidal antibody test (SBA).Methods:fHBP sequence was selected and synthesized, connected to plasmid pET43.1a, transformed to Escherichia coli BL21(DE3), and expressed two recombinant fHBP proteins, included two subfamilies, fHBPA and fHBPB. After purification, the recombinant fHBP proteins were immunized to rabbits and mice. The immune antiserum titer and the bactericidal titer to epidemic strains of meningococcal bacteria group B were measured by ELISA and SBA respectively. Results:The antiserum titer of fHBP immunized rabbits was greater than 2.0×10 6, and that of immunized mice was not less than 1.0×10 6. fHBP immunized rabbit serum had bactericidal titer more than 1∶128 to 41 strains A subfamily and 20 strains B subfamily in the SBA against 69 endemic strains, and there was no cross-protection between the subfamily bacteria. The bactericidal titers of mouse serum immunized fHBPA to strains A subfamily such as Nm210902 Nm211009、Nm450522 were 1∶1 024, 1∶608、1∶861, to Nm510703、Nm311304、Nm431002 were 1∶234、1∶861、1∶430 respectively, and mouse serum immunized fHBP B to strains B subfamily Nm311302、Nm311304、Nm431002 were 1∶876、1∶274、1∶1858, all of three strains were positive in bactericidal titers. Conclusions:the titer of fHBP antiserum was higher than 1.0×10 6, the bactericidal titer was no less than 1∶128 to 61 epidemic strains, and it has a 94.2% protective effect on 69 meningococcal epidemic strains group B.

Objective To evaluate the renal oxygenation in patients with primary nephrotic syndrome (PNS) using BOLD MRI.Methods Twenty patients with untreated first-onset PNS and 18 healthy control subjects underwent BOLD MRI.The R2* of renal cortex and medulla were measured.Blood and urine samples were obtained on the day of MRI,and the patients underwent renal biopsy after MRI.The renal tubulointerstitial damage scores (TIDS) were determined using Katafuchi criteria.All patients received corticosteroids within 7 days after MRI and were followed up for 12 months.The difference of R2* levels between the PNS patients and controls were compared,and the correlations between R2* values and TIDS,laboratory parameters (eGFR,etc.) were tested.Results R2* values of renal medulla in PNS patients significantly decreased compared that of the controls (t =-9.270,P＜0.001).R2* values of renal medulla in PNS patients were negatively correlated with eGFR (r=-0.462,P=0.040) and positively correlated with TIDS (r=0.809,P＜0.001).There was a slight tendency for higher R2* values of renal medulla in individuals with poor prognosis.Conclusion BOLD MRI is a noninvasive method for the detection of renal oxygenation changes,which can evaluate the renal function and tubulointerstitial impairment,as well as prediction of the prognosis for PNS patients.

Objective To investigate the CT and MRI features and morphology classification of intraductal papillary mucinous neoplasm of the bile duct (IPMN-B).Methods A total of 18 patients with IPMN-B proved by pathology were retrospectively analyzed.Out of 18 patients,16 patients underwent enhanced and non-enhanced CT,13 underwent contrast enhanced MR,and 11 out of 13 underwent both CT and MRI.IPMN-B was classified into 4 types:typical IPMN-B,cystic-forming IPMN-B,non-tumor IPMN-B and invasive IPMN-B,according to imaging findings and gross pathological findings.Results Typical IPMN-B (9 cases):tumors were distributed along the bile ducts,both upstream and downstream bile ducts were obviously dilated.Cystic-forming IPMN-B (5 cases):single or multiple tumors were found in aneurysmal dilatation of bile ducts.Non-tumor 1PMN-B (2 cases):no mass was found in the widely dilated bile ducts with smooth bile duct wall.Invasive IPMN-B (2 cases):tumors protruded into the dilated bile ducts causing jagged wall of bile duct,with accompanied abnormal density or signal intensity outside the bile ducts.Bile duct dilatations were shown in all 18 cases,and tumors were shown in 16 cases.In 2 cases no mass was displayed in widely dilated bile ducts.CT density of the tumor was lower than that of liver parenchyma,and higher than that of the bile and intraductal mucin.Signal intensity of the tumor was higher than that of stones,and lower than that of bile and intraductal mucin at MR T2WI.All tumors showed high intensity on DWI.Tumors showed mild to moderate enhancement after injection of contrast agent,CT density or signal intensity of the tumors were lower than that of the liver parenchyma during all three phases of contrast-enhanced CT or MRI.Conclusion IPMN-B has some specific CT and MR imaging features,which are helpful for the diagnosis and classification of IPMN-B.

OBJECTIVE: To explore the expression profile of Ephrin-B2 in the ischemic penumbra after transient focal cerebral ischemia in rats, and to clarify the mechanism of Ephrin-B2 triggering angiogenesis. METHODS: Sprague-Dawley rats were randomly divided into a normal group, a sham operation group and ischemic-reperfusion 1, 3, 7, 14, and 28 d groups. Suture-occluded method was used to establish the focal middle cerebral artery occlusion model and the ischemic brain was reperfused 2 h after the occlusion. Western blot and quantitative real-time reverse-transcription polymerase chain reaction were used to detect the dynamic expression profile of Ephrin-B2 in the penumbra cortex. Double immunofluorescence was used to speculate the location and the co-expression of Ephrin-B2 in blood vessels, neurons and astrocytes. Microvessel density was quantified by the number of CD31+ cells. Rats were subjected to neurologic functional tests by modified neurological severity scores (mNSS) before sacrifice. RESULTS: Compared with the sham group, Ephrin-B2 protein and mRNA level of the penumbra cortex in the ischemic group increased 3 days (P<0.05) after the reperfusion, peaked at day 7 and 14 (P<0.01), and declined at day 28. Double immunofluorescence indicated that Ehprin-b2 was expressed in the neurons, blood vessels and astrocytes; mNSS peaked at day 7, and gradually declined at day 14. The microvessel density of penumbra cortex in the ischemic group increased 3 days (P<0.05) after the reperfusion, peaked at day 14 (P<0.01), and gradually declined at 48 h. CONCLUSION Cerebral ischemia reperfusion induces the over-expression of Ephrin-B2, with a dynamic trend, suggesting that Ehprin-b2 may improve post-stroke functional recovery by enhancing angiogenesis and neurogenesis.
Animals ; Astrocytes ; metabolism ; Brain ; pathology ; Brain Ischemia ; metabolism ; Cerebral Cortex ; metabolism ; Ephrin-B2 ; metabolism ; Infarction, Middle Cerebral Artery ; Ischemic Attack, Transient ; Neurons ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; metabolism

Objective To explore the relationship between lecithin cholesterol acy ltransferase (LCAT) gene 608C/T and 511C/T polymorphisms and stroke in Chinese Han population in Hunan province. Methods One hundred fifty patients with cerebral infarction, 150patients with cerebral hemorrhage, and 122 age- and sex-matched healthy controls were selected.LCAT gene 608C/T and 511C/T polymorphisms were detected by using polyrnerase chain reaction, single strand conformation polymorphism, and restriction fragment length polymorphisms. Results The CT genotype frequency (14. 0% ) and T allele frequency (7. 0% )of the LCAT gene 608C/T in the cerebral infarction group were significantly higher than those in the control group (all P ＜0. 05), while there were no significant differences in the CT genotype frequency (7. 3% ) and T allele frequency (3.7%) between the cerebral hemorrhage group and the control group (P ＞ 0. 05). The CT genotype frequency (10. 0% ) and T allele frequency (5. 0% ) of the LCAT gene 511C/T in the cerebral infarction group were significantly higher than those in the control group (all P ＜0. 01), while there were no significant differences in the CT genotype frequency (3.3%) and T allele frequency (1.7%) between the cerebral hemorrhage group and the control group (P ＞0. 05). Conclusions The 608C/T and 511C/T polymorphisms may be associated with the occurrence of atherosclerotic cerebral infarction in Chinese Han population in Hunan province. They may be the predisposing factors for atherosclerotic cerebral infarction in this population; however, they are not associated with cerebral hemorrhage.

A series of 2-methoxyestradiol (2-MeO-E2) RGD peptide conjugates with coupling RGD peptides to 3- position or 17-position of 2-MeO-E2 through space linker were synthesized. Their antiangiogenic properties were preliminarily evaluated by cell migration scratch assays against HUVECs. Compound 26c binding RGDV peptide showed the best inhibitory effect. In addition, all 2-MeO-E2 RGD peptide conjugates exhibited obvious activity. These results demonstrate that conjugates with RGD peptides represent a promising means for targeting angiogenesis in cancer therapy.

BACKGROUND: Basic fibroblast growth factor (bFGF) can accelerate the bone marrow mesenchymai stem cells (BMSCs) proliferation and differentiation into nerve cells, which is considered as a mitogen of glial cells.OBJECTIVE: To investigate the survival and differentiation of bFGF gene modified BMSCs transplanted on rat models of cerebral ischemia by double immunofluorescence staining, and to study the differentiation trend of BMSCs into neuron-like cells and glial cells.DESIGN, TIME AND SETTING: The randomized control animal experiment was completed in the central laboratory of Experimental Animal Center of Central South University from July 2005 to March 2006.MATERIAL: Fifty Sprague-Dawley rats were divided into 4 groups at random: sham operation group (n=10), cerebral ischemia-reperfusion injury group (n=10), BMSCs group (n=i5) and bFGF modified BMSCs group (n=15). METHODS: Except sham operation group, rats in the other three groups were prepared for local cerebral ischemia-reperfusion models. Then BMSCs or bFGF modified BMSCs were intravenously transplanted into cerebral ischemic rats, and the same volume of DMEM were injected in the cerebral ischemia-reperfusion injury group. MAIN OUTCOME MEASURES: Survival rate and differentiation of grafted cells were observed by 5-bromo-2-deoxyuridine (BrdU)-NeuN, and BrdU-glial fibrillary acidic protein (GFAP) double immunofluorescence staining; the neurological scores and infarction volumes in each group. RESULTS: At 7 days after implantation, the number of BrdU/NeuN-positive cells and BrdU-GFAP-positive cells in the bFGF modified BMSCs group was higher than those on the BMSCs group (P < 0.05), but there were no significant differences in the co-expression cells by double immunofluorescence staining between the two groups (P > 0.05). At 7 days following reperfusion, neural function of cerebral ischemia rats were improved and infarction volume was reduced in both BMSCs group and the bFGF modified BMSCs group, and bFGF modified BMSCs group was superior to BMSCs group. CONCLUSION: BMSCs modified by bFGF can survive in cerebral ischemic region and differentiate into neuron and glial cells, which are more proper for repairing nerves.