Objective To investigate the feasibility of fetal liver cells for liver tissue engineering, the supporting function of poly L lactic acid (PLLA) scaffold for fetal liver cells and the effects of oncostatin M (OSM), nicotinamide (NA) and dimethyl sulfoxide(DMSO) on growth and hepatic differentiation. Methods After three dimensional PLLA scaffolds having a porous structure were prepared by using NH 4HCO 3 particle, fetal liver cells obtained from E14.5 C57BL/6CrSlc murine embryos were inoculated in the scaffolds. Cells were cultured in Williams’E medium with or without OSM, NA and DMSO for 30 days. Changes in cell number, liver-specific function, and cellular morphology were observed. Results When compared with in monolayer culture, cell number and albumin secretion increased obviously in three-dimensional PLLA. Alburmin secretion increased slightly in OSM group of monolayer culture, but increased obviously in OSM groupo of PLLA culture and in OSM/NA/DMSO group of both monlayer and PLLA cultures. Conclusion The three-dimensional PLLA scaffold is a good supporting material for the cultivation of tetal liver cells. OSM, NA and DMSO remarkaly stimulated maturation of hepatic parenchymal cells in vitro in terms of morphology and liver-specific function.
Objective To observe the degradation of the polyactic glycolate acid (PLGA) microparticles with releasing-slowly vascular endothelial growth factor(VEGF) synthesized by the method of emulsification-diffusion. Methods The method of emulsification-diffusion is to incorporate VEGF into microparticles composed of biodegradable PLGA. The controlled release of microparticles are acquired. The content of the VEGF released slowly from PLGA microparticles in vitro was detected with ELISA at different time. Results We synthesized 100 releasing-slowly VEGF PLGA microparticles with the size of 0.20-0.33 μm by 5 times. The contents were 62±11 ng/L, 89±14 ng/L, and 127±19 ng/L in the 1st, the 2nd and the 3rd months after degradation, respectively. Conclusion The PLGAmicroparticles with releasing-slowly VEGF can be synthesized by the method of emulsification-diffusion.
ObjectiveTo evaluate the biocompatibility of poly lactic acid/bone matrix gelatin (PLA/BMG) composite biomaterial so as to lay a foundation for bone defect repair. MethodsRats'MC3T3-E1 cells were cultured with leaching solution of PLA/BMG and PLA material respectively for 7 days. The cell proliferation rate was tested by MTT and cell toxicity grading was carried out everyday. The PLA/BMG and MC3T3-E1 cells were co-cultured, the cell shape and proliferation were observed by inverted phase contrast microscope at 1, 3, and 5 days and cell adhesion by scanning electron microscope at 5 days. The PLA and PLA/BMG were implanted subcutaneously in 15 Wistar rats. The histological observation was done, and the thickness of fibrous membrane, the number of inflammatory cells, and the vascularization area were measured at postoperative 2nd, 4th, and 8th week. ResultsThe tests for cytotoxicity in vitro showed that the cell proliferation rates were over 100% and the cell cytotoxic grades were grade 0 at 1-7 days in PLA/BMG group. While in PLA group, the cell proliferation rates were less than 100% and the cell cytotoxic grades were grade 1 at 2, 4, and 7 days. After co-culture of PLA/BMG and MC3T3-E1 cells, cells grew on the surface and in the pores of PLA/BMG, and the cellular morphology was triangle or polygon with abundant microvillus on the surface. After subcutaneous implantation, the rats survived to the end of experiment, and incision healed well. PLA was wrapped by connective tissue where there were a lot of lymphocytes and neutrophilic granulocytes. The cells and tissue grew slowly in PLA. The PLA/BMG materials were wrapped by little connective tissue where there were a few inflammatory cells. The connective tissue ingrowth was observed in the center of PLA/BMG. There was no significant difference in the thickness of fibrous membrane between 2 groups at each time point (P>0.05). The number of inflammatory cells of PLA/BMG group were significantly less than those in PLA group at 2, 4, and 8 weeks (P<0.05); the vascularization area was significantly larger than that in PLA group (P<0.05). ConclusionPLA/BMG composite biomaterials prepared by super critical-CO2 technique are good in cell and tissue biocompatibilty.
Objective To study the influence of in vitro force-vascularization on in vivo vascularization of porous polylactic glycolic acid copolymer(PLGA) scaffolds with internal network channels (PPSINC). Methods After the in vitro forcevascula ization of PPSINCs covered with microvessel endothelial cells (MVEC) of mice, they were divided into two groups: the force-vascularization group (group A) and the control group with only PSINCs (group B). All the PPSINCs were planted in the mesentery of 12 mice for 2 and 4 weeks, the PPSINCs were cut out, the vascular ization of PPSINCs was investigated by histology and immunohistochemistry, and the vascularization area of the histologic section of the PPSINCswas measured with the computer-assistant image analysis system. Result After the in vitro forcevascularization of PPSINCs, the MVEC of the mice sticking on the channel wall could be seen. After the scaffold was im planted into the mice for 2 weeks, the vascularization area of the histologic section of PPSINCs (VA) in group A (2 260.91±242.35 μm2) was compared with that in group B (823.64±81.29 μm2)，and the difference was sig nificant in statistics（P＜0.01).The VA for 4 weeks in group A (17 284.36 ±72.67 μm2) was compared with that in group B (17 041.14±81.51 μm2), and the difference was not significant in statistics（P＞0.05）.The area of the actin positivestaining (AA) in the histologi c section of PPSINCs for 2 weeks’ implantation in group A (565.22±60.58 μm2) was compared with that in group B (205.91±16.25 μm2), and the difference was signi ficant in statistics（P＜0.01). After the implantation for 4 weeks, the VA in group A (4 321.09±19.82 μm2) was compared with group B (4 260.28±27.17 μm2), and the difference was not significant in statistics（P＞0.05）. Conclusion The PPSINC is a good simple scaffold model of vasculariazation. The in vitro force-vascularization can increase the in vivo vascularization of PPSINCs in the early stage.
Objective To investigate the correlation between the initial arterial blood lactic acid and Acute Physiology and Chronic Health Evaluation (APACHE) Ⅱ score in trauma patients and its value in prognosis. Methods From August 1st 2015 to July 31st 2016, the clinical data of trauma patients treated in Department of Emergency were analyzed retrospectively. All patients were divided into survival group and death group by observing 28-day prognosis. We compared the relationship between the initial blood lactate level and APACHEⅡ score, and analyzed the relationship between the above indexes and the prognosis of the patients. Results A total of 743 patients were enrolled, with692 in survival group and 51 in death group.The APACHEⅡ score and initial blood lactate level in the survival group [(9.93±4.62) points, (2.02±1.44) mmol/L] were significantly lower than those in the death group [(22.84±7.26) points, (4.60±3.69) mmol/L] with significant differences (t=18.20, 9.77; P<0.01). The APACHEⅡ score and the mortality rate of patients with lactic acid level >4 mmol/L were significantly higher than those of patients with lactic acid of 2-4 mmol/L and <2 mmol/L; the differences were significant (P<0.05). The blood lactate and mortality in patients with APACHEⅡ score >20 were significantly higher than those in the patients with ≤10 and 11-20; the differences were significant (P<0.05). There was a significant positive correlation between initial blood lactate level and APACHEⅡ score (r=0.426, P<0.01). Conclusions The initial blood lactate level and APACHEⅡ score of trauma patients are correlated with the severity of injury and mortality. Both of the increase of initial blood lactic acid level and APACHEⅡ score suggest the risk of death in trauma patients.
Objective To evaluate the predictive value of lactate dehydrogenase (LDH) to albumin (Alb) ratio (LAR) in the prognosis of severe pneumonia patients complicated with DIC. Methods A total of 312 patients with severe pneumonia hospitalized in the intensive care unit (ICU) of the Affiliated Changzhou No.2 People's Hospital with Nanjing Medical University from January 1, 2018 to March 1, 2023 were retrospectively collected. The clinical parameters, such as gender, age, underlying diseases, and lactate dehydrogenase, albumin etc. l of the first test on admission were collected. LAR, sequential organ failure assessment (SOFA) and acute physiology and chronic health evaluation Ⅱ(APACHEⅡ) within 24 hours were calculated. The firstly endpoint of the study was the incidence of disseminated intravascular coagulation (DIC), the secondary endpoint was the 30-day in-hospital mortality in severe pneumonia patients with DIC. Univariate and multivariate logistic regression were used to analyze the risk factors of severe pneumonia with DIC. The receiver operating characteristic curve (ROC curve) was drawn and the area under the ROC curve (AUC) was calculated to evaluate the predictive value of LAR for the incidence of DIC in patients with severe pneumonia. Results The level of LAR was higher in the severe pneumonia patients with DIC than the severe pneumonia patients without DIC [LAR median ratio 12.72 (8.72, 21.89) vs. 7.23 (5.63, 10.90), P<0.001]. Multiple logistic regression analysis showed that LAR [OR=1.071, 95%CI 1.038 - 1.106, P<0.001] was the independent risk factor of the incidence of DIC in the patients with severe pneumonia. ROC curve analysis showed that the AUC for LAR to predict the incidence of DIC was 0.723, 95%CI 0.650 - 0.796, P<0.001. When the LAR cut-off value was 8.08, the sensitivity was 79.7% and the specificity was 56.1%. Kaplan-Meier survival analysis curve showed that the patients in the above LAR cut-off value group had a significantly lower 30-day survival rate than those in the below LAR cut-off value group (P<0.001). In the subgroup analysis and numerical variable transformed analysis, LAR was still the risk factor of DIC. Conclusion The increased LAR is a high risk factor of the incidence of DIC and mortality in patients with severe pneumonia, which is useful for predicting prognosis of patients with severe pneumonia.
ObjectiveTo investigate the effect of silk fibroin-poly-L-lactic acid (SF-PLLA) microcarriers on the expansion and differentiation of adipose-derived stem cells (ADSCs).MethodsADSCs were extracted from adipose tissue donated voluntarily by patients undergoing liposuction by enzymatic digestion. The 3rd generation ADSCs were inoculated on CultiSpher G and SF-PLLA microcarriers (set up as groups A and B, respectively), and cultured in the rotary cell culture system. ADSCs cultured in normal two-dimensional plane were used as the control group (group C). Scanning electron microscope was used to observe the microcarriers structure and cell growth. Live/Dead staining and confocal fluorescence microscope was used to observe the distribution and survival condition of cells on two microcarriers. DNA quantification was used to assess cell proliferation on two microcarriers. Real-time fluorescence quantitative PCR (qRT-PCR) was used to detect chondrogenesis, osteogenesis, and adipogenesis related gene expression of ADSCs in 3 groups cultured for 18 days. Flow cytometry was used to identify the MSCs surface markers of ADSCs in 3 groups cultured for 18 days, and differential experiments were made to identify differentiation ability of the harvested cells.ResultsADSCs could be adhered to and efficiently amplified on the two microcarriers. After 18 days of cultivation, the total increment of ADSCs of the two microcarriers were similar (P>0.05). qRT-PCR results showed that chondrogenesis related genes (aggrecan, cartilage oligomeric matrix protein, SOX9) were significantly up-regulated for ADSCs on SF-PLLA microcarriers and adipogenesis related genes (peroxisome proliferator-activated receptor γ, lipoprotein lipase, ADIPOQ) were significantly up-regulated for ADSCs on CultiSpher G microcarriers, all showing significant differences (P<0.05). Flow cytometry and differentiation identification proved that the harvested cells of the two groups were still ADSCs.ConclusionThe ADSCs can be amplified by SF-PLLA microcarriers, and the chondrogenic differential ability of harvested cells was up-regulated while the adipogenic differential was down-regulated.
Objective To explore the effect of NaOH on the surface morphology of three-dimensional (3D) printed poly-L-lactic acid (PLLA) mesh scaffolds. Methods The 3D printed PLLA mesh scaffolds were prepared by fused deposition molding technology, then the scaffold surfaces were etched with the NaOH solution. The concentrations of NaOH solution were 0.01, 0.1, 0.5, 1.0, and 3.0 mol/L, and the treatment time was 1, 3, 6, 9, and 12 hours, respectively. There were a total of 25 concentration and time combinations. After treatment, the microstructure, energy spectrum, roughness, hydrophilicity, compressive strength, as well as cell adhesion and proliferation of the scaffolds were observed. The untreated scaffolds were used as a normal control. Results 3D printed PLLA mesh scaffolds were successfully prepared by using fused deposition molding technology. After NaOH etching treatment, a rough or micro porous structure was constructed on the surface of the scaffold, and with the increase of NaOH concentration and treatment time, the size and density of the pores increased. The characterization of the scaffolds by energy dispersive spectroscopy showed that the crystal contains two elements, Na and O. The surface roughness of NaOH treated scaffolds significantly increased (P<0.05) and the contact angle significantly decreased (P<0.05) compared to untreated scaffolds. There was no significant difference in compressive strength between the untreated scaffolds and treated scaffolds under conditions of 0.1 mol/L/12 h and 1.0 mol/L/3 h (P>0.05), while the compression strength of the other treated scaffolds were significantly lower than that of the untreated scaffolds (P<0.05). After co-culturing the cells with the scaffold, NaOH treatment resulted in an increase in the number of cells on the surface of the scaffold and the spreading area of individual cells, and more synapses extending from adherent cells. Conclusion NaOH treatment is beneficial for increasing the surface hydrophilicity and cell adhesion of 3D printed PLLA mesh scaffolds.