Objective To investigate the effect of mRNA expression of gelatinase A on the invasion and metastasis of human gastric carcinoma (HGC). MethodsThirtysix cases of HGC were examined by in situ hybridization technique. ResultsPositive expression rates of gelatinase A in the normal gastric tissue, peritumor tissue and HGC were 8.3%,35.7% and 83.3% respectively （P＜0.01）. The positive rates of gelatinase A in the group with serosal invasion and lymph node metastasis were 93.1% and 90.6%, much higher than those in the group with negative ones (42.9% and 25.0%).By in situ hybridization, gelatinase A mRNA was showed to be expressed in the extracellular matrix of tumor tissues,which surrounded the invasive margin of cancer tissues. The positive cells at these sites were mainly tumorinfiltrating macrophages. Conclusion There is good correlation between gelatinase A mRNA expression and the invasion, metastasis of HGC. So it can be used as a useful marker for invasion and metastasis of HGC.
Objective To compare the characteristics of gelatin microspheres crossl inked by glutaraldehyde (GA) or geni pin (GP). Methods Gelatin microspheres, prepared by the improved emulsified cold-condensation method, were crossl inked by GP and GA, respectively. After being dispersed in PBS, two kinds of microspheres with 60% degree of cross l inking were compared in terms of morphology, swell ing and degrading properties. rhBMP-2 were loaded into the GP and GAmicrospheres, and the encapsulation rate, drug loading and releasing capacity were measured; 100%, 50% and 25% leaching l iquid of GP and GA microspheres were respectively cultured with rat osteoblast (DMEM group as the control), and cell prol iferation was measured by MTT method to grade the cell cytotoxicity. Results GP and GA microspheres were both spherical with the diameters of (78 ± 18) μm and (65 ± 10) μm, and there were no difference between both microspheres in drug loading and encapsulation rate. But, GP microspheres, with long degrading period (28 days) compared to GA microspheres (21 days), had better dispersibil ity, and swell ing rate (89.0% ± 4.8%), the percentage of cumulative drug releasing at 10 days (78.80% ± 4.96%) were both lower than GA microsphere (118.0% ± 7.6%, 90.50% ± 5.12%). The percentages of drug loading of GP and GA were (921 ± 73) and (965 ± 62) ng/g, and the encapsulation rates were 88.5% ± 2.1% and 89.7% ± 1.8%; showing no significant difference (P gt; 0.05). The cell cytotoxicity of 100%, 50% and 25% leaching l iquid of GP microspheres was all at the level I, but leaching l iquid of GA microspheres with corresponding concentration were at the levels of III, III and II. Conclusion GP crossl inked gelatin microspheres are superior to GA crossl inked gelatin microspheres and can be widely used in tissue engineering field.
Objective To investigate the feasibil ity of prefabricating urethra in the expander capsule with gelatin sponge and micro-mucosa compound transplantation. Methods Eight 8-week-old Guizhou miniature pigs (male and/or female) weighing 20-25 kg were used. Six expanders (15 mL) were placed subcutaneously on the dorsal thorax of each miniaturepig. Autologous oral mucosa of every pig was harvested 2 weeks later to prepare micro-mucosa with a diameter less than 1 mm. Gelatin sponge 3 cm × 2 cm in size was transplanted to the expander capsule after being coated by the autologous micromucosa at the area expansion ratio of 4 ∶ 1 (group A), 8 ∶ 1 (group B), and 16 ∶ 1 (group C), respectively (n=2 per group). The implantation of gelatin sponge served as the blank control (group D, n=2). Physiological sal ine was injected into the expander immediately after operation, and the pressure in the expander was 40 mm Hg (1 mm Hg=0.133 kPa). The postoperative general condition of the animals was observed. At 1, 2, and 3 weeks after operation, the animals were killed to receive general, HE staining, and immunohistochemistry staining observations. Results All animals survived till the end of the experiment. The wounds healed well. General observation: in groups A, B, and C at 1 week after operation, there was no obvious degeneration of gelatin, the mucous was survived partially, and there were significant differences among three groups in terms of mucosa healing rate (P lt; 0.05), groups A and B were better than group C, and group A was better than group B; at 2 weeks, the gelatin sponge was partly absorbed, most of the mucosa survived, and the mucosa healing rate of groups A and B was better than that of group C (P lt; 0.05); at 3 weeks, the gelatin sponge was still not absorbed completely, the wound reached epithel ial ization approximately,and there were no significant differences among three groups in terms of mucosa heal ing rate (P gt; 0.05). No neo-mucosa was evident in group D at each time point. Histology and immunohistochemistry staining observation: at each time point, the mucosa epithel ium survival, inflammatory cell infiltration, and pan-cytokeratin were evident in groups A, B, and C; at 3 weeks after operation, the stratified squamous epithel ium presented obvious polarity and the submucous neovascularization was abundant in groups A, B, and C. There was no mucosa epithelium and positive stained pan-cytokeratin in group D. For the percentage of positive pan-cytokeratin stained area, there were significant differences among groups A, B, and C 1 week after operation (P lt; 0.05); at 2 and 3 weeks after operation, there was significant difference between group A and group C, and between group B and group C (P lt; 0.05); but no significant difference was evident between group A and group B (P gt; 0.05). Conclusion Micro-mucosa and gelatin spongy compound transplantation on the expander capsule can form mucosal l ining, achieve complete epithel ial ization in 2 weeks, and contribute to maintain the normal function of prefabricatied urethra.
Objective To investigate the histological and keratinous variation of prefabricated urethra in the capsule with micro-mucosa and gelatin sponge compound graft. Methods Five 8-week-old Guizhou miniature pigs (2 females and3 males) weighing 20-25 kg were used. Eight tissue expanders were bilaterally inserted into subcutaneous position on the dorsal thorax of each pig. Forty inserted expanders were randomized into two groups (n=20 per group). For the experimental group, the free buccal mucosa was cut into particles less than 1 mm in diameter, spread onto the gelatin sponge (3 cm × 2 cm) and then transplanted to the capsule; the area expansion ratio of autogenous micro-mucosa was 8 ∶ 1. For the control group, soft tissue expander without mucosa graft was implanted. The pressure in inserted expander was about 40 mm Hg (1 mm Hg=0.133 kPa). Inflation should be stopped when the injected sal ine volume reached 15 mL. The animals were killed 1 and 2 weeks and 1, 2, and 4 months after the implant to receive examination. Macroscope, histology, and immunohistochemistry changes were observed. Results All the animals survived to the end of the experiment and the wounds healed by first intention. There was no obvious degeneration of gelatin sponge, and some of the mucosa survived 1 week after implant. The gelatin sponge was partly absorbed, most of the mucosa survived 2 weeks after implant. Visual examination showed complete epithel ial ization of the entire cavity 1 month after implant. The experimental group at 2 and 4 months were similar to that of at 1 month in gross observations.The neo-mucosa was not found in the control group at different time points after implant. Histology examination revealed that compound implant was mainly infiltrated by inflammatory cells and the micro-mucosa survived well 1 week after implant in the experimental group. The stratified squamous epithel ium presented obvious polarity and the submucous neovascularization was abundant 2 weeks after implant. The compound implant achieved complete epithel ial ization 1 month after implant. The epithel ium degeneration occurred 2 months after implant. The stratified squamous epithel ium presented no abovious polarity 4 months after implant. No neo-mucosa was evident in control group at different time points. The experimental group was positive for the pan-cytokeratin staining at 1, 2 weeks, and 1, 2 months after implant, but negative at 4 months after implant The pan-cytokeratin staining was negative in the control group at different time points. Conclusion The buccal micromucosa and gelatin sponge compound graft can grow well on the expanded capsule 1 month after implant and the epithel ium degeneration is evident 2 months after implant. Environment of implanted mucosa has great influence on epithel ium mucosa.
ObjectiveTo study the growth of adipose-derived stem cells (ADSCs) planted in three-dimensional (3D) materials, a 3D cultured ADSCs system based on microbial transglutaminase (mTG) enzyme crosslinked gelatin hydrogel was constructed. MethodsADSCs were isolated from the subcutaneous adipose tissue of a Sprague Dawley rat by collagenase digestion and centrifugation, and were cultured for passage. The mTG enzyme crosslinked gelatin hydrogel was firstly synthesized by mixing gelatin and mTG, and then the ADSCs were encapsulated in situ (2D environment) and cultured in the 3D materials (3D environment). The morphology and adhesion of cells were observed by inverted phase contrast microscope. In addition, HE staining and Masson staining were carried out to observe the distribution of cells in the material. Living and death situation of ADSCs in the materials was observed by fluorescence microscope and laser scanning confocal microscopy. Scanning electron microscopy was used to observe the adhesion of ADSCs on hydrogel surface. Alamar-Blue method was used to detect the proliferation of ADSCs in the hydrogel. Moreover, the results were compared between the cells cultured in 2D environment and those in 3D environment. ResultsThe result of 2D culture showed that ADSCs grew well on the hydrogel surface with normal functioning and had good adhesion. The results of 3D culture showed that ADSCs grew well in 3D cultured mTG enzyme crosslinked gelatin hydrogel, and presented 3D shape. Cells obviously extended in all directions. The number of apoptotic cells was very small. The cells of 3D culture at each time point was significantly less than that of the conventional culture cells, difference was statistically significant (P < 0.05). But after 8 days culture, the proliferation of the cells cultured in the mTG enzyme crosslinked gelatin hydrogel increased more quickly. ConclusionADSCs can grow well with good adhesion and show high viability in 3D culture system constructed by mTG enzyme crosslinked gelatin hydrogel.
ObjectiveThe antifriction and antiwear effects of gelatin nanoparticles (GLN-NP) on artificial joint materials in bionic joint lubricant were investigated to provide a theoretical basis for the development of new bionic joint lubricant. MethodsGLN-NP was prepared by cross-linking collagen acid (type A) gelatin with glutaraldehyde by acetone method, and the particle size and stability of GLN-NP were characterized. The biomimetic joint lubricants with different concentrations were prepared by mixing 5, 15, and 30 mg/mL GLN-NP with 15 and 30 mg/mL hyaluronic acid (HA), respectively. The friction reduction and antiwear effects of the biomimetic joint lubricants on zirconia ceramics were investigated on a tribometer. The cytotoxicity of each component of bionic joint lubricant on RAW264.7 mouse macrophages was evaluated by MTT assay. ResultsThe particle size of GLN-NP was about 139 nm, and the particle size distribution index was 0.17, showing a single peak, indicating that the particle size of GLN-NP was uniform. In complete culture medium, pH7.4 PBS, and deionized water at simulated body temperature, the particle size of GLN-NP did not change more than 10 nm with time, indicating that GLN-NP had good dispersion stability and did not aggregate. Compared with 15 mg/mL HA, 30 mg/mL HA, and normal saline, the friction coefficient, wear scar depth, width, and wear volume were significantly reduced by adding different concentrations of GLN-NP (P<0.05); there was no significant difference between different concentrations of GLN-NP (P>0.05). Biocompatibility test showed that the cell survival rate of GLN-NP, HA, and HA+GLN-NP solution decreased slightly with the increase of concentration, but the cell survival rate was more than 90%, and there was no significant difference between groups (P>0.05). ConclusionThe bionic joint fluid containing GLN-NP has good antifriction and antiwear effect. Among them, GLN-NP saline solution without HA has the best antifriction and antiwear effect.
The study aimed to evaluate the therapeutic effect of nilotinib-loaded biocompatible gelatin methacryloyl (GelMA) microneedles patch on cardiac dysfunction after myocardial infarction(MI), and provide a new clinical perspective of myocardial fibrosis therapies. The GelMA microneedles patches were attached to the epicardial surface of the infarct and peri-infarct zone in order to deliver the anti-fibrosis drug nilotinib on the 10th day after MI, when the scar had matured. Cardiac function and left ventricular remodeling were assessed by such as echocardiography, BNP (brain natriuretic peptide) and the heart weight/body weight ratio (HW/BW). Myocardial hypertrophy and fibrosis were examined by WGA (wheat germ agglutinin) staining, HE (hematoxylin-eosin staining) staining and Sirius Red staining. The results showed that the nilotinib-loaded microneedles patch could effectively attenuate fibrosis expansion in the peri-infarct zone and myocardial hypertrophy, prevent adverse ventricular remodeling and finally improve cardiac function. This treatment strategy is a beneficial attempt to correct the cardiac dysfunction after myocardial infarction, which is expected to become a new strategy to correct the cardiac dysfunction after MI. This is of great clinical significance for improving the long-term prognosis of MI patients.