Objective To compare the effect of ibuprofen and glucosamine on synoviocyte proliferation and cartilage oligomeric matrix protein (COMP) expression in human knee osteoarthritis. Methods Human synoviocytes were isolated from synovium (earlier stage and late stage of OA) by tissue culture and were cocultured with ibuprofen and glucosamine. The concentration of COMP was determined by MTS/PMS method and hCOMP kit. Two-tailed t-test was used for statistical analysis. Results The observation time of tissue culture was determined at 5～7 day by the MTS/PMS method. The A values of glucosamine [ late stage group (0.054±0.021), early stage group (0.777±0.034)] were less than the normal serum control group (P＜0.05).Both ibuprofen [late stage group (35.4±1.9), early stage group (46.0±2.2)] and glucosamine [late stagegroup (36.6±1.3), early stage group (48.8±1.3) ] could decrease the concentration of COMP in synoviocyte secretion in vitro (P＜0.05). Conclusion Glucosamine can inhibit the synoviocyte proliferation of human knee osteoarthritis (both early stage and late stage) in vitro. Both ibuprofen and glucosamine can inhibit the COMP secretion of synoviocyte in vitro.

Objective To compare the effect of glucosamine (Virtral-s) on the cartilage oligomeric matrix protein (COMP) secretion of chondrocytes and synoviocytes in vitro.Methods Chondrocytes and synoviocytes isolated from knee cartilage of osteoarthritic patients were cultured by phased enzymatic digestion.Sera containing Virtral-s of the experimental animals were obtained after orally administrated Virtral-s at the dosages that equal to human.Cells were cultured in the medium with Virtral-s containing sera.Super-natant COMP level was tested by enzyme-linked immunoabsorbent assays (ELISA).Results COMP conceu-tration of synoviocytes cultured in vitro was significantly higher than that of chondrocytes (P＜0.05).Virtral-s could significantly increase COMP secretion in cultured chondrocytes in vitro (P＜0.05),however,it had a weaker role on synoviocytes,ie,it could only mildly reduce COMP secretion of synoviocytes.Conclusion Glucosamine (Virtral-s)-containing serum can promote COMP secretion of chondrocytes in vitro,and it has no significant effect on synoviocytes in vitro.

Objective To study the levels of Matrix Metalloproteinase-3 (MMP-3) and interleukin (IL-1) in the synovial fluid and plasma of C57 black mice with osteoarthritis (OA) and their relationships with the severity of pathological changes so as to investigate their effects and correlation with OA. Methods The C57 black mice with OA were enrolled for this study. Different levels of exercise were appicated based on their age. Knee joint pathological changes were examined for pathological severity of OA. ELISA sandwich method was used to measure the levels of MMP-3 and IL-1 in serum and synovial fluid. Correlation analysis was performed to demonstrate the relationship between the levels of MMP-3 and IL-1 in the serum and synovial fluid, and the pathological severity of OA. Results ①Morphological observations: C57 black mice were characterized by spontaneously developing OA and the incidence and the severity of osteoarthritis gradually increased with age and exercising burdens. ② The level of MMP-3 and IL-1 in the synovial fluid of exercising mice MMP-3 (84±6) ng/ml, IL-1 (48±3) ng/ml was higher than that in the aged ones [MMP-3 (84±6) ng/ml, IL-1 (71±5) ng/ml J, the difference was significant (P<0.01). The level of MMP-3 and IL-1 level in the serum had a linear correlation with that of the synovial fluid. At the same time, they also had linear correlation with the pathological severity of OA (All r>0.67, and all P<0.01). Conclusion The levels of MMP-3 and IL-1 in serum and synovial fluid can help to make early diagnosis of OA, especially elevated MMP-3 level.

OBJECTIVETo investigate the role of human Wnt7b gene in rat chondrocyte degeneration.

METHODSWnt7b gene obtained by PCR was cloned to PCDH-GFP. 293ft cell line was transfected with PCDH-GFP and PCDH-Wnt7b, and the supernatant and transfected cells were collected. The expression level of Wnt7b in 293ft cells was detected by Western blotting. The first passage of chondrocytes were isolated from articular cartilages of newborn born (within 24 h) SD rats were cultured in the supernatants from the transfected cells (at 10- and 50-fold dilutions). The cell morphology of the rat chondrocytes was observed under inverted microscope, and the protein expressions of MMP13, MMP3 and type II collagen and mRNA expressions of A-can, ADAMTS5, Col X and Sox9 were examined by Western blotting or real-time PCR.

RESULTSHuman Wnt7b gene cloned to PCDH-GFP was expressed efficiently in 293ft cell line. Rat chndrocytes cultured for 24 h in the supernatants from PCDH-Wnt7b-transfected 293ft cells underwent changes from a polygonal to a spindle-shaped morphology. The protein expression levels of MMP13 and MMP3 increased while type II collagen decreased significantly, and the mRNA levels of A-can and Sox9 were down-regulated while Col X and ADMATS5 up-regulated in ratchondrocytes after incubation in supernatants from PCDH-Wnt7b-transfected 293ft cells.

CONCLUSIONHuman Wnt7b gene can be expressed efficiently in 293ft cell line and can induce rat chondrocyte degeneration in vitro.

Animals ; Cartilage, Articular ; cytology ; Cell Line ; Chondrocytes ; pathology ; Humans ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley ; Real-Time Polymerase Chain Reaction ; Transfection ; Wnt Proteins ; genetics

Objective To observe the effects of percutaneous endoscopic gastrostomy (PEG)on mortality and complications in patients with persistent dysphagia after stroke using a points scoring system for selecting PEG indication.Methods A total of 75 patients were divided into low score group without PEG,high score group without PEG and low score group with PEG (n=25 each).The follow-up period was 18 months,and the differences in complications,mortalities and survival periods among groups were compared.Results The number of times of aspiration pneumonia was (1.36± 1.44) in low score group,(1.96±2.28) in high score group,(0.36±0.64) in low score group with PEG,with statistically significant differences among three groups (H=7.148,P=0.028).No difference in the morbidity of aspiration pneumonia was found between low score group and high score group (P=0.189).The number of times of aspiration pneumonia was decreased in low score groups after PEG versus in low score group without PEG (P=0.030) and in high score group (P＜0.01).The numberof times of gastrointestinal hemorrhage was (0.48± 0.77)in low score group,(0.64± 0.91) in high score group,(0.12±0.33) in low score group with PEG,with statistically significant differences among three groups (H=5.532,P =0.063).No statistically significant difference in gastrointestinal hemorrhage was found between low score groups and low score group after PEG (P=0.430),as well as between low score group and low score group with PEG (P=0.079).The morbidity of gastrointestinal hemorrhage was lower in low score group than in high score group (P=0.012).The survival rate at the observation end was 88.0％ (22/25),52.0％ (13/25) and 92.0％ (23/25) in low score group,high score group and low score group with PEG,respectively,with statistically significant difference among the three groups (x2 =7.906,P =0.001).Kaplan-Meier survival curve showed that the survival period were longer in the low score group with or without PEG than in high score group (P＜0.01),but no statistically significant difference was found between low score groups with or without PEG (P=0.626).Conclusions The reasonable evaluation using a points-scoring system before PEG might predict the prognosis of such patients:the higher score would indicate higher mortality.PEG operation for low score group with better condition could decrease the aspiration pneumonia and decrease gastrointestinal hemorrhage significantly,but could not prolong general survival time and decrease general mortality.

Objective To investigate the role of human Wnt7b gene in rat chondrocyte degeneration. Methods Wnt7b gene obtained by PCR was cloned to PCDH-GFP. 293ft cell line was transfected with PCDH-GFP and PCDH-Wnt7b, and the supernatant and transfected cells were collected. The expression level of Wnt7b in 293ft cells was detected by Western blotting. The first passage of chondrocytes were isolated from articular cartilages of newborn born (within 24 h) SD rats were cultured in the supernatants from the transfected cells (at 10-and 50-fold dilutions). The cell morphology of the rat chondrocytes was observed under inverted microscope, and the protein expressions of MMP13, MMP3 and type II collagen and mRNA expressions of A-can, ADAMTS5, Col X and Sox9 were examined by Western blotting or real-time PCR. Results Human Wnt7b gene cloned to PCDH-GFP was expressed efficiently in 293ft cell line. Rat chndrocytes cultured for 24 h in the supernatants from PCDH-Wnt7b-transfected 293ft cells underwent changes from a polygonal to a spindle-shaped morphology. The protein expression levels of MMP13 and MMP3 increased while type II collagen decreased significantly, and the mRNA levels of A-can and Sox9 were down-regulated while Col X and ADMATS5 up-regulated in ratchondrocytes after incubation in supernatants from PCDH-Wnt7b-transfected 293ft cells. Conclusion Human Wnt7b gene can be expressed efficiently in 293ft cell line and can induce rat chondrocyte degeneration in vitro.

OBJECTIVETo investigate the effect of inhibition and activation of MAPK-ERK1/2 pathway on the expression of osteogenic genes and proliferation of rat osteoblasts in vitro.

METHODSPrimarily cultured rat osteoblasts, identified by cell morphology studies and ALP staining, were exposed to 1% or 5% rat serum for 24 h or to the specific MAPK-ERK1/2 inhibitor PD0325901. The downstream molecules of MAPK-ERK1/2 pathway including p-ERK1/2 and ERK1/2, osteogenic genes such as Runx2 and Type I collagen, and proliferating cell nuclear antigen (PCNA) were detected by Western Blotting, and alkaline phosphatase activities were analyzed quantitatively.

RESULTSCompared with 1% rat serum-treated cells, exposure of the cells to a higher concentration (5%) of rat serum caused a significantly increased phosphorylation level of p-ERK1/2 (P<0.05) and obviously enhanced expressions of the osteogenic genes (Runx2, type I collagen and ALP) and PCNA (P<0.05). Inhibition of the MAPK-ERK1/2 pathway with PD0325901 resulted in suppressed expressions of the osteogenic genes and PCNA.

CONCLUSIONThe activation of MAPK-ERK1/2 pathway promotes the expression of osteogenic genes such as Runx2, type I collagen and ALP and enhances the proliferative activity of the osteoblasts, while inhibition of this pathway suppresses the expressions of these genes and the cell proliferation, suggesting that this pathway may potentially serve as a therapeutic target for osteoporosis.

Alkaline Phosphatase ; metabolism ; Animals ; Antineoplastic Agents ; pharmacology ; Benzamides ; pharmacology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen Type I ; metabolism ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Diphenylamine ; analogs & derivatives ; pharmacology ; Female ; Gene Expression Regulation ; drug effects ; MAP Kinase Signaling System ; drug effects ; Male ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Osteoblasts ; cytology ; metabolism ; Phosphorylation ; drug effects ; Proliferating Cell Nuclear Antigen ; metabolism ; Rats ; Rats, Sprague-Dawley

Objective To investigate the role of human Wnt7b gene in rat chondrocyte degeneration. Methods Wnt7b gene obtained by PCR was cloned to PCDH-GFP. 293ft cell line was transfected with PCDH-GFP and PCDH-Wnt7b, and the supernatant and transfected cells were collected. The expression level of Wnt7b in 293ft cells was detected by Western blotting. The first passage of chondrocytes were isolated from articular cartilages of newborn born (within 24 h) SD rats were cultured in the supernatants from the transfected cells (at 10-and 50-fold dilutions). The cell morphology of the rat chondrocytes was observed under inverted microscope, and the protein expressions of MMP13, MMP3 and type II collagen and mRNA expressions of A-can, ADAMTS5, Col X and Sox9 were examined by Western blotting or real-time PCR. Results Human Wnt7b gene cloned to PCDH-GFP was expressed efficiently in 293ft cell line. Rat chndrocytes cultured for 24 h in the supernatants from PCDH-Wnt7b-transfected 293ft cells underwent changes from a polygonal to a spindle-shaped morphology. The protein expression levels of MMP13 and MMP3 increased while type II collagen decreased significantly, and the mRNA levels of A-can and Sox9 were down-regulated while Col X and ADMATS5 up-regulated in ratchondrocytes after incubation in supernatants from PCDH-Wnt7b-transfected 293ft cells. Conclusion Human Wnt7b gene can be expressed efficiently in 293ft cell line and can induce rat chondrocyte degeneration in vitro.

Bone tissue engineering (BTE) is a rapidly developing strategy for repairing critical-sized bone defects to address the unmet need for bone augmentation and skeletal repair. Effective therapies for bone regeneration primarily require the coordinated combination of innovative scaffolds, seed cells, and biological factors. However, current techniques in bone tissue engineering have not yet reached valid translation into clinical applications because of several limitations, such as weaker osteogenic differentiation, inadequate vascularization of scaffolds, and inefficient growth factor delivery. Therefore, further standardized protocols and innovative measures are required to overcome these shortcomings and facilitate the clinical application of these techniques to enhance bone regeneration. Given the deficiency of comprehensive studies in the development in BTE, our review systematically introduces the new types of biomimetic and bifunctional scaffolds. We describe the cell sources, biology of seed cells, growth factors, vascular development, and the interactions of relevant molecules. Furthermore, we discuss the challenges and perspectives that may propel the direction of future clinical delivery in bone regeneration.
Animals ; Bone Regeneration ; Cell Differentiation ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stem Cells ; cytology ; Osteogenesis ; Tissue Engineering ; methods ; Tissue Scaffolds

Objective To observe changes in genetic material in the peripheral blood of pediatric patients with vascular malformations after interventional procedures. Methods A total of 108 children with vascular malformations who underwent interventional procedures at Shandong University Affiliated Children’s Hospital between February 2021 and January 2024 were selected as the research subjects. Clinical data and peripheral venous blood samples before and after the interventional procedures were collected from the children. Two biological indicators, γ-H2AX and peripheral blood lymphocyte chromosomal aberration (CA), were used to determine the levels of genetic material damage in children with vascular malformations before and after interventional procedures. Results The median age of the children was 7 years and the median body weight was 27 kg. The median dose-area product (DAP) was 24.20 Gy·cm² and the median DAP/kg was 1.04 Gy·cm²/kg. The incidence rates of both γ-H2AX foci and CA in children with vascular malformations significantly increased after the interventional procedures (Z = 5.924, P < 0.001; Z = 8.515, P < 0.001). The incidence of postoperative CA in 7 children were significantly higher than that in others, approaching or exceeding 4%. The incidence rates of postoperative γ-H2AX foci and CA in children with DAP/kg ≥ 1 Gy·cm²/kg were significantly higher than those in children with DAP/kg < 1 Gy·cm²/kg (U = 7.586, P = 0.031; U = 6.835, P = 0.009). No significant differences were observed in the incidence rates of postoperative γ-H2AX foci and CA among subgroups based on age, body weight, or surgical site. A positive correlation was observed between the difference in the incidence rates of γ-H2AX foci before and after the procedure and DAP/kg (R = 0.493, P = 0.027). Conclusion Ionizing radiation exposure during interventional procedures can increase peripheral blood genetic material damage levels in children with vascular malformations, and the damage levels show a correlation with the radiation dose, with some children being abnormally sensitive. Further research is needed to explore the influencing factors for genetic material damage in children with vascular malformations after interventional procedures, which is of great significance for reducing long-term cancer risks and achieving personalized treatment strategies.