Objective To determine the efficacy of D980-nm laser in dissolving fat and renewing skin, and to explore the clinical application of D980-nm laser in reconstruction of photodamaged skin. Methods Eighteen 12-14 month-old male Sprague-Dawley rats, weighing 400-450 g, were randomly divided into 3 groups (n=6). The rat skin at the left side was exposed to D980-nm laser irradiation at a density of 20 J/cm2, a power of 8 W, a pulse width of 20 ms, and a pulse frequency of 40 Hz for 1 time (group A), 2 times of 5-minute interval (group B), and 3 times of 5-minute interval (group C) as a treatment course, for 4 treatment courses with an interval of 1 week; the other side of the skin was not treated as the control groups (groups A1, B1, and C1, respectively). After 8 weeks, the skin was harvested for HE staining and immunohistochemical staining to observe the structure changes of skin, to measure the dermal thickness, to count the number of fibroblasts, and detect the expressions of transforming growth factor β1 (TGF-β1) and basic fibroblast growth factor (bFGF). Results Compared with groups A1, B1, and C1, the skin structure was significantly improved in groups A, B, and C. After D980-nm laser irradiation, the number of fat cells decreased; local angiogenesis was observed; the total number of fibroblasts and fibers increased; the collagen fiber had large diameter, and arranged closely and regularly; the dermal thickness and the number of the fibroblasts increased; and the expressions of TGF-β1 and bFGF were significantly enhanced, showing significant differences (P<0.05). With increased D980-nm laser irradiation times, the above indexes increased, showing significant differences between group C and groups A, B (P<0.05). Conclusion D980-nm laser treatment has lipolytic and tender effect on the skin, and the frequency of the treatment is an important factor in skin renewal.
OBJECTIVE: To study the expression of type I collagen and its receptor system-integrin alpha 2 beta 1 in different passages of osteoblasts. METHODS: The expression of type I collagen and integrin alpha 2 beta 1 in the primary, sixth and fifteenth passage of osteoblasts were detected by S-P immunohistological staining technique, and their mRNA expression by quantity RT-PCR technique. RESULTS: Type I collagen and integrin alpha 2 beta 1 were expressed in different passages of osteoblasts and there was no significant difference among three passages by immunohistological technique. Their mRNA expression was gradually decreased with subculture. CONCLUSION: Type I collagen promotes the adhesion and phenotype expression of osteoblasts through its receptor-integrin alpha 2 beta 1. The reductive expression of type I collagen-receptor system will decline the phenotype of osteoblasts.
Objective To investigate the curative effects of homograft of the mesenchymal stem cells(MSCs) compbined with the medical collagen membrane of the guided tissue regeneration(MCMG) on the full thickness defects of the articular cartilage. Methods MSCs derived from New Zealand rabbits aged 3-4 months weighing 2.1-3.4 kg were cultured in vitro with a density of 5.5×108/ml and seeded onto MCMG. The MSC/MCMG complex was cultured for 48 h and transplanted into the fullthickness defects on the inboardcondyle and trochlea. Twenty-seven healthy New Zealand rabbits were randomly divided into 3 groups of 9rabbits in each. The cartilage defects in the inboard condyle and trochlea werefilled with the auto bone marrow MSCs and MCMG complex (MSCs/ MCMG) in Group A (Management A), with only MCMG in Group B (Management B)and with nothing in Group C (Management C). Three rabbits were killed at 4, 8 and 12 weeks after operation in each group, and the reparative tissue samples evaluated grossly,histologically and immunohistochemically were graded according tothe gross and histological scale. Results Four weeks after transplantation, the cartilage and subchondralbone were regenerated in Group A;for 12 weeks, the regenerated cartilage gradually thicked; 12 week after transplantation, the defect was repaired and the structures of the carticular surface and subchondral bone was in integrity.The defects in Group A were repaired by the hylinelike tissue and the defects in Groups B and C were repaired by the fibrous tissues. Glycosaminoglycan and type Ⅱcollagen in Groups A,B and C were reduced gradually.The statistical analysis on the gross at 12 weeks and the histologicalgradings at 4 weeks,8 weeks and 12 weeks showed that the inboardcondylar repairhad no significant difference compared with the rochlearepair(Pgt;0.05).Management A was significantly better than Managements B and C (Plt;0.05), and Management B was better than Management C(Plt;0.05). Conclusion Transplantation of the MSCs combined with MCMG on the full thickness defects of the articular cartilage is a promising approach to the the treatment of cartilage defects. MCMG can satisfy the demands of the scaffold for the tissue-engineered cartilage.
The aim of this paper is to explore the prevention of rabbit postoperative abdominal cavity adhesion with poly (lactic-co-glycotic acid) (PLGA) membrane and the mechanism of this prevention function. Sixty-six Japanese white rabbits were randomly divided into normal control group, model control group and PLGA membrane group. The rabbits were treated with multifactor methods to establish the postoperative abdominal cavity adhesion models except for those in the normal control group. PLGA membrane was used to cover the wounds of rabbits in the PLGA membrane group and nothing covered the wounds of rabbits in the model control group. The hematologic parameters, liver and kidney functions and fibrinogen contents were detected at different time. The rabbit were sacrificed 1, 2, 4, 6, 12 weeks after the operations, respectively. The adhesions were graded blindly, and Masson staining and immunohistochemistry methods were used to observe the proliferation of collagen fiber and the expression of transforming growth factorβ1 (TGF-β1) on the cecal tissues, respectively. The grade of abdominal cavity adhesion showed that the PLGA membrane-treated group was significant lower than that in the model control group, and it has no influence on liver and kidney function and hematologic parameters. But the fibrinogen content and the number of white blood cell in the PLGA membrane group were significant lower than those of model control group1 week and 2 weeks after operation, respectively. The density of collagen fiber and optical density of TGF-β1 in the PLGA membrane group were significant lower than those of model control group. The results demonstrated that PLGA membrane could be effective in preventing the abdominal adhesions in rabbits, and it was mostly involved in the reducing of fibrinogen exudation, and inhibited the proliferation of collagen fiber and over-expression of TGF-β1.
Objective To retrospectively analyze the cl inical effect of l ightbulb operation with nano-hydroxyapatite/ collagen in a consecutive series of patients with osteonecrosis of the femoral head (ONFH). Methods From January 2001to July 2005, 26 patients (35 hips) were treated, 16 males and 10 females, aged 19-54 years old (33.5 on average). The course of disease was 12-36 months (18 months on average). Based on the etiology, 15 cases (22 hips) were steroid induced type, 10 (12 hips) were alcohol induced type and the other one (1 hip ) was idiopathic type. According to the system of Association Research Circulation Osseous (ARCO), there were 6 hi ps of stage IIB, 16 hi ps of stage IIC, 9 hi ps of stage IIIA, 3 hi ps of stage IIIB and 1 hip of stage IIIC. The Harris score was 62.2 ± 7.5. All the patients who had undergone l ightbulb operation with nano-hydroxyapatite/collagen were evaluated both cl inically and radiographically. The bone graft mixture rate of nanohydroxyapatite/ collagen and autogenous bone was 1 ∶ 1, and the mixed bone graft was 6 times of the scraped osteonecrosis volume (30-48 mL). Results The incisions of all 26 patients (35 hi ps) obtained heal ing by first intention. The 2 cases, which got lateral femoral cutaneous nerve injury during the operation, recovered 3-6 months after the operation without any treatment. Another 2 cases got heterotopic ossification 3 months after operation, with no special treatment. All the 26 patients (35 hips) were followed up for 2-7 years (3.5 on average). The patients’ bone heal ing began from the 3rd month after operation. The postoperative Harris score was 85.1 ± 16.2, and there was significant difference compared with the preoperative one (P lt; 0.001). There were 15 hips of excellent, 11 of good, 5 of fair, and 4 of poor which received total hip arthroplasty at the end of the follow-up. According to imaging, 5 hips were progressed from preoperative IIC to IIIA, while the other hips were radiologically stable, with no progress of ONFH. Conclusion Lightbulb operation with nano-hydroxyapatite/collagen provides a surgical treatment to treat early ONFH with satisfactory cl inical outcomes. Nano-hydroxyapatite/collagen is beneficial for the repair and reconstruction of ONFH and suitable for femoral-head-preserving operation for the patients with ONFH of stage II.
The aim of this article is to study how andrographolide-releasing collagen scaffolds influence rabbit articular chondrocytes in maintaining their specific phenotype under inflammatory environment. Physical blending combined with vacuum freeze-drying method was utilized to prepare the andrographolide-releasing collagen scaffold. The characteristics of scaffold including its surface morphology and porosity were detected with environmental scanning electron microscope (ESEM) and a density instrument. Then, the release of andrographolide from prepared scaffolds was measured by UV-visible spectroscopy. Rabbit chondrocytes were isolated and cultured in vitro and seeded on andrographolide-releasing collagen scaffolds. Following culture with normal medium for 3 d, seeded chondrocytes were cultured with medium containing interleukin-1 beta (IL-1β) to stimulate inflammation in vitro for 7 d. The proliferation, morphology and gene transcription of tested chondrocytes were detected with Alamar Blue assay, fluorescein diacetate (FDA) staining and reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR) test respectively. The results showed that the collagen scaffolds prepared by vacuum freeze-dry possess a high porosity close to 96%, and well-interconnected chambers around (120.7±17.8) μm. The andrographolide-releasing collagen scaffold continuously released andrographolide to the PBS solution within 15 d, and collagen scaffolds containing 2.22% andrographolide significantly inhibit the proliferation of chondrocytes. Compared with collagen scaffolds, 0.44% andrographolide-containing collagen scaffolds facilitate chondrocytes to keep specific normal morphologies following 7 d IL-1β induction. The results obtained by RT-qPCR confirmed this effect by enhancing the transcription of tissue inhibitor of metalloproteinase-1 (TIMP-1), collagen II (COL II), aggrecan (Aggrecan) and the ratio of COL II/ collagen I(COL I), meanwhile, reversing the promoted transcription of matrix metalloproteinase-1 (MMP-1) and matrix metalloproteinase-13 (MMP-13). In conclusion, our research reveals that andrographolide-releasing (0.44%) collagen scaffolds enhance the ability of chondrocytes to maintain their specific morphologies by up-regulating the transcription of genes like COL II, Aggrecan and TIMP-1, while down-regulating the transcription of genes like MMP-1 and MMP-13 which are bad for phenotypic maintenance under IL-1β simulated inflammatory environment. These results implied the potential use of andrographolide-releasing collagen scaffold in osteoarthritic cartilage repair.
Osteoporosis is a degenerative disease characterized by decreased bone mass and destruction of bone microstructure. At present, previous studies have found that the structure and content of type Ⅰ collagen fibers are closely related to osteoporosis. However, there have been few studies on the prevention and treatment of osteoporosis using type Ⅰ collagen fibers as therapeutic targets. In this paper, the relationships between type Ⅰ collagen fibers and osteoporosis, biomechanics, bone matrix and bone strength are discussed. At the same time, the regulation of type Ⅰ collagen-related signaling pathways in osteoporosis is summarized, such as the signaling pathways of cathepsin K, transforming growth factor-β/Sma- and Mad-related protein, transforming growth factor-β/bone morphogenetic protein, c-jun N-terminal protein kinase and Wnt/β-catenin, in order to provide a new therapeutic direction for the prevention and treatment of osteoporosis.
Objective To review the lately new progress of fish collagen as biomedical materials, and then analyze feasibility and risk management of its application as a substitute of collagen originated from mammals in clinical practice. Methods Based on extensive research on new application and investigation of fish collagen, the paper was prepared to bring comprehensive analysis of its research and application status, and then several key points were focused on. Results Fish collagen has been proved to be a novel collagen of rich source, low risk of virus transmission, low biological risk, less religious barrier, and high biocompatibility. Fish collagen has promising prospect when applied in clinical practice as novel collagen especially as a substitute of collagen derived from mammals. However, very few related translational medicine research of fish collagen has been reported up to now in China. Conclusion As a novel potential substitute of collagen source derived from mammals, fish collagen is concerned to be clinical feasible and necessary in translational medicine. However, massive applied basic researches should be focused on in the further investigations.
Objective To develop a novel porous three-dimensional scaffold and to investigate its physico-chemical properties for tissue engineering cartilage.Methods Refined 88% deacetylation degree chitosan was prepared and dissolved in 0.2 mol/L acetate acid and fully mixed with highly purified porcine type Ⅱcollagen in 0.5 mol/L acetate acid solution in a ratio of 4 to 1 (wt/wt). Freeze-drying process was employed to fabricate the composite scaffold. The construct wascross-linked by use of 1-ethyl-3(3-dimethyl aminopropyl) carbodiimide (EDC) and Nhydroxysuccinimide (NHS). A mechanical tester was utilized to determine the tensilestrength change before and after cross-linking. The microstructure was observed via scanning electron microscopy (SEM). The lysozyme degradation was performedto evaluate the degradability of the scaffold in vitro. Results A bulk scaffold with desired configuration was obtained. The mechanical test showed that the crosslinking treatment could enhance the mechanical strength of the scaffold. The SEM results revealed that the two constituents evenly distributed in the scaffold and that the matrix was porous, sponge-like with interconnected pore sizing 100250 μm. In vitro lysozyme degradation indicated that crosslinked or uncross-linked composite scaffolds had faster degradation rate than the chitosan matrix. Conclusion Chitosan and typeⅡcollagen can be developed into a porous three-dimensional scaffold. The related physico-chemical tests suggest that the composite socaffold meets requirements for tissue engineered scaffold and may serve as an alternative cellcarrier for tissue engineering cartilage.
ObjectiveTo construction the telmisartan/collagen/polycaprolactone (PCL) nerve conduit and assess its effect on repairing sciatic nerve defect in rats. Methods The 60% collagen/hexafluoroisopropanol (HFIP) solution and 40% PCL/HFIP solution were prepared and mixed (collagen/PCL solution). Then the 0, 5, 10, and 20 mg of telmisartan were mixed with the 10 mL collagen/PCL solution, respectively. Telmisartan/collagen/PCL nerve conduits were fabricated via high voltage electrospinning technology. The structure of nerve conduit before and after crosslinking was observed by using scanning electron microscope (SEM). The drug release efficiency was detected by in vitro sustained release method. RAW264.7 cells were cultured with lipopolysaccharide to induce inflammation, and then co-cultured with nerve conduits loaded with different concentrations of telmisartan for 24 hours. The mRNA expressions of inducible nitric oxide synthase (iNOS) and Arginase 1 (Arg-1) were detected by using real-time fluorescence quantitative PCR. Forty adult Wistar rats were randomly divided into 4 groups (n=10). After preparing 15-mm-long sciatic nerve defect, the defect was repaired by cross-linked nerve conduits loaded with 0, 5, 10, and 20 mg telmisartan in groups A, B, C, and D, respectively. After operation, the general condition of rats was observed after operation; the sciatic function index (SFI) was tested; the bridging between the nerve conduit and sciatic nerve, and the integrity of nerve conduit were observed; the tissue growth in nerve conduit and material degradation were observed by HE staining; the expressions of CD86 (M1 macrophage marker), CD206 (M2 macrophage marker), myelin basic protein (MBP), and myelin protein 0 (P0) in new tissues were also observed by immunohistochemical staining; the expressions of neurofilament 200 (NF-200) and S-100β in new tissues were assessed by immunofluorescence staining. Results The general observation showed that the inner diameter of the nerve conduit was 1.8 mm and the outer diameter was 2.0 mm. After cross-linking by genipin, the nanofiber became thicker and denser. The drug release test showed that the telmisartan loaded nerve conduit could be released gradually. With the increase of telmisartan content in nerve conduit, the iNOS mRNA expression decreased and the Arg-1 mRNA expression increased; and the differences between 20 mg group and other groups were significant (P<0.05). In vivo experiment showed that all animals in each group survived until the completion of the experiment. The SFI was significantly higher in groups C and D than in groups A and B at different time points (P<0.05) and in group D than in group C at 6 months after operation (P<0.05). HE staining showed that there were significantly more new tissues in the middle of the nerve conduit in group D after operation than in other groups. Immunohistochemical staining showed that CD86 and CD206 stainings were positive in each group at 1 month after operation, among which group D had the lowest positive rate of CD86 and the highest positive rate of CD206, and there were significant differences in the positive rate of CD206 between group D and groups A, B, and C (P<0.05); the MBP and P0 stainings were positive in groups C and D at 6 months, and the positive rate in group D was significantly higher than that in group C (P<0.05). Immunofluorescence staining showed that the NF-200 and S-100β expressions in group D were significantly higher than those in other groups. ConclusionTelmisartan/collagen/PLC nerve conduit can promote the sciatic nerve defect repair in rats through promoting the polarization of M1 macrophages to M2 macrophages, and the nerve conduit loaded with20 mg telmisartan has the most significant effect.