BACKGROUND:The repair of temporomandibular joint disc is stil a great clinical chal enge. Pluripotent adipose-derived mesenchymal stem cells bring a hope for fibrocartilage tissue repair. Up to now, there are few studies concerning transforming growth factor-β3 to induce differentiation of adipose-derived mesenchymal stem cells into fibrochondrocytes.
OBJECTIVE:To observe the growth and morphology of adipose-derived mesenchymal stem cells under the induction of transforming growth factor-β3.
METHODS:We observed the morphology of fibrochondrocytes differentiated from adipose-derived mesenchymal stem cells under the induction of transforming growth factor-β3. Histological and immunofluorescence staining were employed to detect the expression of extracellular matrix Ⅰ, Ⅱ col agens and proteoglycansthe produced by adipose-derived mesenchymal stem cells. We also evaluated the feasibility of adipose-derived mesenchymal stem cells as seed cells for fibrocartilage tissue engineering.
RESULTS AND CONCLUSION:Under the inverted fluorescence microscope, induced cells showed obvious aggregation growth, polygonal shape, and increased secretion of extracellular matrix. Alcian blue staining results showed that under the induction of transforming growth factor-β3, adipose-derived mesenchymal stem cells exhibited the distinctly deep blue, indicating a large number of glycosaminoglycan was synthesized. Immunostaining results showed that under the induction of transforming growth factorβ3, adipose-derived mesenchymal stem cells synthesized extracellular matrix Ⅰ, Ⅱ col agen. These findings suggest that transforming growth factorβ3 can induce adipose-derived mesenchymal stem cells to differentiate into fibrouscartilage-like cells, and that adipose-derived mesenchymal stem cells can serve as seed cells for fibrocartilage tissue engineering.

The aim of this study was to obtain the alginate gels which could have proper compressive strength and excellent permeability for cell proliferation and could have more promising potentials in the application of tissue engineering. Through the reaction of the carboxyl of the alginate and the amino of methacrylic acid, methylacrylic was generated into alginate long chain which could be enhanced by the polymerization of double bond under thermal reaction condition. And then alginate gel beads were prepared using the mixture of calcium chloride and barium chloride solution as cationic crosslinker, and the compressive modulus and permeability of the prepared alginate gel beads were investigated. When the ratio of barium ions to calcium ions was 5:5, the compression modulus was 189.7 kPa, and it showed the best permeability for trypsin with molecular weight of 24 kDa and entrapment effect for bovine serum albumin with molecular weight of 67 kDa. Compared to compositions of other ratios, the alginate gel beads made in 5:5 mixture indicated excellent compressive modulus and permeability. These results indicated that the alginate hydrogel beads with the ratio of barium ions to calcium ions being 5:5 have a potential application in tissue engineering as a support material and encapsulating materials in cell culture.
Alginates ; chemistry ; Barium Compounds ; chemistry ; Calcium Chloride ; chemistry ; Cations ; Cell Culture Techniques ; Cell Proliferation ; Chlorides ; chemistry ; Cross-Linking Reagents ; chemistry ; Gels ; chemistry ; Metals ; Polymethacrylic Acids ; chemistry ; Tissue Engineering

Background and purpose:Tumor budding is a poor prognostic factor in colorectal cancer. In this study, we studied the tumor budding by counting the actual number in 10 high power fields and evaluated itsclinical application in predicting lymph node metastasis of T1 colorectal cancer.Methods:Tissue specimens from 307 patients with histologically conifrmed T1 colorectal cancer were enrolled. The clinicopathological characteristics including tumor budding were evaluated for their predictive value in lymph node metastasis. A formula was created to calculate the risk score for prediction of lymph node metastasis which was validated by 14 new cases.Results:In the multivariate analysis, it showed that tumor grade, lymphovascular invasion and the number of tumor budding were signiifcantly associated with lymph node metastasis. The probability of lymph node metastasis was calculated using the following equations:Z=1.571×(lymphovascular state: invasion, 1; no invasion, 0)+2.661×(tumor grade: high grade, 1; low grade, 0)+0.024×(budding counts)-3.885; Probability=1/1+e-Z. The high scores were correlated with the lymph node metastasis in the validations.Conclusion:We can accurately assess the risk of lymph node metastasis by counting the number of tumor budding in 10 high power fields. Therefore tumor budding could potentially assist treatment decision making in T1 colorectal cancer patients with high-risk lymph node metastasis.

Objective:To investigate the effects of cyclic tensile stress on the function and degeneration of nucleus pulposus cells.Methods:The human primary nucleus pulposus cells were isolated and cultured. The cyclic tensile stress (100 000 μ?, 10% tensile strain, 0.1 Hz, 8 640 cycles) was loaded on the cells for 24 h. The proliferation of the cells was examined by MTT method. The cell cycle and apoptosis were detected through flow cytometry. Gene expression profile chip was used to detect the differentially expressed genes between the tensile stress group and control group. The function of these gene was analyzed by bioinformatics. The expression of inflammatory related factors, TGF-β, matrix degrading enzymes and extracellular matrix molecules were examined by qRT-PCR.Results:The cyclic tensile stress significantly promoted proliferation and cell cycle of nucleus pulposus cells. The cell percentage of S phase ( t=5.336, P<0.05) and G2/M phase ( t=7.288, P<0.01) was significantly different between the tensile stress group and control group. The cyclic tensile stress inhibited apoptosis of nucleus pulposus cells (8.56%±0.48% vs 10.63%±0.32%, t=4.474, P<0.05). A total of 866 differentially expressed genes were detected. Gene ontology analysis showed the roles of these genes in cells including focal adhesion, extractable matrix, membrane raft, condensed chrome kinetochore, cytoskeleton, etc. The cyclic tensile stress significantly affected the mRNA expression of inflammatory related factors, TGF-β genes, matrix proteinase and extracellular matrix molecules. Compared with the control group, the mRNA expression of inflammatory related factors IL15 ( t=5.379, P<0.05), IGF1 ( t=5.454, P<0.05) and IGFBP7 ( t=13.57, P<0.01) were significantly decreased in the tensile stress group; The mRNA expression of TGF-β genes TGFB1 ( t=6.931, P<0.05), TGFB2 ( t= 15.56, P<0.01) and TGFB3 ( t=7.744, P<0.05) were significantly increased in the tensile stress group; The mRNA expression of matrix proteinase ADAMTS3 ( t=5.241, P<0.05) and MMP19 ( t=24.72, P<0.01) were significantly decreased, and TIMP3 ( t=8.472, P<0.01) increased in the tensile stress group; The mRNA expression of extracellular matrix molecules COL2A1 ( t=5.871, P<0.05), FLRT2 ( t=5.216, P<0.05) and FN1 ( t=4.289, P<0.05) were significantly increased. Conclusion:The cyclic tensile stress promoted cell cycle and proliferation and inhibited apoptosis of nucleus pulposus cells. The cyclic tensile stress may affect the function and degeneration of nucleus pulposus cells by regulating the expression of inflammatory related factors, TGF-β, matrix degradation enzymes and ECM molecules.

Aim To investigate the protective effects of beraprost sodium on cerebral cortical neuron injury in chronic aluminum-overload rats and its effects on PGIS-IP signaling pathway. Methods 75 SD rats were randomized into five groups: normal control group, chronic aluminum-overload group ( model group) and beraprost sodium groups-low dose (6 μg· kg-1 ), medium dose ( 12 μg · kg-1 ) and high dose (24 μg·kg-1). Aluminum gluconate (Al3+ 200 mg ·kg-1 d-1, once a day, 5d a week, for 20 weeks, p. o. ) was administered to rats of cerebral damage model. The rats of experimental groups were concomi-tantly treated with beraprost sodium ( p. o. ) daily for 20 weeks. After the model was built successfully, the spatial learning and memory( SLM) function was done by Morris water maze. The cortical neurons damage was detected by HE staining, SOD activities and MDA contents. The 6-k-PGF1α levels in cortex were meas-ured by ELISA. The expressions of PGIS, IP mRNA and IP protein were also studied. Results Compared with the rats of normal control group, the SLM function was significantly impaired ( P<0. 01 ) and considera-ble karyopycnosis was observed in model group rats. The SOD activities were weakened ( P <0. 01 ), the MDA contents increased ( P<0. 05 ) and the levels of 6-k-PGF1α raised significantly ( P <0. 01). The ex-pressions of PGIS and IP mRNA in the rats cortex obvi-ously increased ( P<0. 01 ), so did the expression of IP protein(P<0. 05). Compared with the rats of mod-el group, the SLM function of rats in experimental groups decreased significantly ( P<0. 01 ) and damage of cortical neurons reduced remarkably. The SOD ac-tivities increased ( P <0. 01 ) and the MDA contents decreased ( P <0. 01). Besides, the content of 6-k-PGF1α, the expressions of PGIS mRNA and IP protein in the rats cortex decreased significantly ( P<0. 05 ) as well as IP mRNA ( P<0. 01). Conclusion Our re-sults demonstrate that in cerebral cortical neuron of chronic aluminum-overload rats, beraprost sodium has notably protective effects and the mechanism might be related to PGIS-IP signaling pathway.

Molecular mechanisms underlying the effects of Fyn on cell morphology, pseudopodium movement, and cell migration were investigated. The Fyn gene was subcloned into pEGFP-N1 to produce pEGFP-N1-Fyn. Chinese hamster ovary (CHO) cells were transfected with pEGFP-N1-Fyn. The expression of Fyn mRNA and proteins was monitored by reverse transcription-PCR and Western blotting. Additionally, transfected cells were stained with 4',6-diamidino-2-phenylindole and a series of time-lapse images was taken. Sequences of the recombinant plasmids pMD18-T-Fyn and pEGFP-N1-Fyn were confirmed by sequence identification using National Center for Biotechnology Information in USA, and Fyn expression was detected by RT-PCR and Western blotting. The morphology of CHO cells transfected with the recombinant vector was significantly altered. Fyn expression induced filopodia and lamellipodia formation. Based on these results, we concluded that overexpression of mouse Fyn induces the formation of filopodia and lamellipodia in CHO cells, and promotes cell movement.
Animals ; Blotting, Western ; CHO Cells ; Cricetinae ; Cricetulus ; Genetic Vectors ; Green Fluorescent Proteins/genetics ; Mice ; Proto-Oncogene Proteins c-fyn/genetics/*metabolism ; Pseudopodia/*metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Time-Lapse Imaging ; Transfection

Objective:To explore the correlation between fibular head height and varus knee osteoarthritis occurrence and severity.Methods:A retrospective analysis was performed on 618 participants (618 knees, 184 males and 434 females, mean age 61.12±10.98 years) who underwent standard weight-bearing full-leg radiographs and were diagnosed as non-knee osteoarthritis or varus knee osteoarthritis from January 2019 to June 2019. Knee osteoarthritis was diagnosed according to Kellgren-Lawrence grading: 0-I grades were diagnosed as non-osteoarthritis, II-IV grades were diagnosed as osteoarthritis. Joint line convergence angle (JLCA), medial proximal tibial angle (MPTA) and hip-knee-ankle angle were measured on X-rays to reflect varus deformity. The fibular head height was defined as the vertical distance from upper edge of fibular head to lateral tibial plateau. Patients were divided into 5 groups according to Kellgren-Lawrence grading. Differences of age, gender, height, weight, body mass index, varus deformity (JLCA, MPTA and hip-knee-ankle angle) between Kellgren-Lawrence 0-IV grades were compared. Ordinal logistic regression was performed to analyze the correlation between fibular head height and Kellgren-Lawrence grades. Pearson's correlation analysis was used for the correlation among fibular head height, JLCA, MPTA and hip-knee-ankle angle, and the main factor of JLCA, MPTA and hip-knee-ankle angle was extracted by factor analysis. Multiple linear regressions were used to analyze the correlation between fibular head height and varus deformity score.Results:There were 68, 66, 97, 98, 289 participants in Kellgren-Lawrence grades 0-IV respectively that was 134 participants were diagnosed as non-osteoarthritis and 484 participants were diagnosed as osteoarthritis. Fibular head height and MPTA showed a decreasing trend ( F=129.076, 24.875; P<0.001) while JLCA and hip-knee-ankle angle showed an increasing trend ( F=414.346, 105.996; P<0.001) with the increase in Kellgren-Lawrence grading. Age, body mass index and fibular head height are influencing factors of Kellgren-Lawrence grading with OR(95%CI) were 1.116(1.093, 1.141), 1.363(1.060, 1.754), 0.617(0.575, 0.662) . Fibular head height was negatively correlated with JLCA and hip-knee-ankle angle ( r=-0.641, -0.478; P<0.001) , respectively, and positively correlated with MPTA ( r=0.320, P<0.001). There were significant correlations between age, fibular head height and the varus deformity score ( β=0.274, -0.457; P<0.001). Conclusion:Fibular head height of patients with varus knee osteoarthritis is lower than that of non-osteoarthritis. In addition to age and body mass index, fibular head height is a risk factor for varus knee osteoarthritis occurrence. The smaller the fibular head height is, the more serious the osteoarthritis severity and varus deformity are.

Objective To investigate the diagnostic value of superb microvascular imaging ( SM I ) , advanced dynamic flow ( ADF ) and color Doppler flow imaging ( CDFI) in breast microcarcinoma ,and to compare the correlation coefficients between these three indicators and postoperative pathological tumor microvascular density ( M VD ) . Methods A total of 85 patients ( 87 lesions ) with BI‐RADS 4 and the maximum diameter≤1 cm were selected ,of which ,46 lesions were benign ( benign group) and 41 lesions were malignant( malignant group) . All the patients and the corresponding lesions were examined by SM I , ADF and CDFI . Blood flow grading was performed on the images using Adler grade of blood flow ,and the difference of blood flow among the 3 methods was compared . T he expression level of M VD in pathological tumor tissues was detected and analyzed for its correlation with Adler classification by three detection techniques . Results T he areas under the ROC curve( AUR) of CDFI ,ADF and SM I were 0 .694 ,0 .705 and 0 .776 respectively based on the gold standard with pathological diagnosis . T he sensitivity ,specificity , positive predictive value ,negative predictive value ,and accuracy of CDFI were 78 .0% ,54 .3% ,60 .4% , 73 .5% ,and 65 .5% , respectively ; those of ADF were 75 .6% , 60 .9% , 63 .3% , 73 .7% and 67 .8% , respectively ; and those of SM I were 78 .0% ,69 .6% ,69 .6% ,78 .0% and 73 .6% ,respectively . T he Adler grades of CDFI , ADF and SM I were positively correlated with M VD ( P ＜ 0 .05 ) , w hich the highest correlation coefficient between SM I and M VD ( r ＝0 .430 , P ＜0 .001 ) . Conclusions SM I is superior to ADF and CDFI in detecting the abundance of breast microcarcinoma , and has the highest correlation coefficient among those 3 detection techniques with tumor pathological M VD ,which indicates that SM I may be used for differential diagnosis of breast microcarcinoma and indirectly evaluate the prognosis of patients .

Objective:To investigate the distribution of potential suitable habitats for Ixodes persulcatus in Heilongjiang Province during different climatic periods. Methods:The geographical location data of Ixodes persulcatus in Heilongjiang Province from 1980 to 2023 were collected through literature search and field ecological investigation. The environmental factor variables were downloaded from the WorldClim website and the Resource and Environmental Science and Data Platform. A combination of SDMtune, ENMTools packages, and the maximum entropy (MaxEnt) model were used to predict the potential habitats of Ixodes persulcatus in Heilongjiang Province during different climatic periods and determine its dominant environmental factors. Among them, the time range of historical climatic period data was 1970 - 2000. The future climatic period data (2021 - 2040, 2041 - 2060) were selected as the low forcing scenario (SSP126 scenario) of the shared socioeconomic pathways (SSP) in the Beijing Climate Center - Climate System Model version 2-Medium Resolution, to simulate the impact of global warming on the distribution of potential habitats for Ixodes persulcatus in Heilongjiang Province. Results:A total of 83 geographical distribution sites of Ixodes persulcatus in Heilongjiang Province were obtained. After screening, 10 environmental factors were included in the MaxEnt model. Among them, the cumulative contribution percentage of annual precipitation, average annual temperature, and vegetation type was 67.3%. Compared with historical climatic period, the total potential habitats of Ixodes persulcatus in Heilongjiang Province increased from 29.81 × 10 4 km 2 to 32.24 × 10 4 km 2 in 2041 - 2060. The center of potential habitats moved to 47.75° N, and 128.66° E, with a northwest offset of 126.07° and a migration distance of 45.83 km. Conclusion:With global climate warming, the total potential habitats of Ixodes persulcatus in Heilongjiang Province has increased, showing an overall trend of migration to the northwest.

Objective:To investigate the effect of CHI3L1 on the biological function of chondrocytes and its role in lumbar facet joint degeneration.Methods:The human lumbar facet joint articular cartilage were collected, and the relative mRNA expression of CHI3L1 gene detected by quantitative fluorescence PCR. Then explored the correlation between joint degeneration and gender, age and relative mRNA expression of CHI3L1. Human chondrocytes were cultured in vitro. The effects of CHI3L1 on chondrocyte proliferation, cycling, and apoptosis, as well as expression of related inflammatory factors, were investigated. The mechanism by which CHI3L1 regulates the degeneration of articular cartilage was investigated using the signal transduction pathway protein chip.Results:There was a positive correlation between the grade of degeneration in lumbar facet joint and the relative expression of CHI3L1 gene mRNA ( r=0.76, P<0.001). There was no correlation with the patient's gender ( r=-0.12, P=0.500). A positive correlation between the age of patients and the relative expression of CHI3L1 gene mRNA was found ( r=0.47, P=0.005). Compared with the non-degenerative group, the expression of CHI3L1 gene mRNA significantly increased in the degenerative group, and the expression of CHI3L1 gradually increased with the aggravation in the grade of degeneration ( F=18.90, P<0.001). Compared with the non-degenerative group, the chondrocytes in the CHI3L1 group had significantly lower proliferation at 48 h (OD 490/fold=7.132), 72 h (OD 490/fold=4.803), 96 h (OD 490/fold=2.431) and 120 h (OD 490/fold=0.009). The ratio of chondrocytes in G1 phase, S phase and G2/M phase were 85.03%±3.05%, 12.78%±2.29% and 0.90%±0.76% in the CHI3L1 group, and 73.93%±2.73%, 22.81%±1.93% and 0.99%±0.87% in control group, respectively. There were significant differences in the percentage of chondrocytes in G1 phase ( t=4.70, P<0.001) and S phase ( t=5.80, P<0.001) between the two groups. The percentages of apoptosis in chondrocyte in CHI3L1 group and control group were 8.64%±0.76% and 5.68%±1.13%, which has a statistically difference ( t=4.47, P<0.001). The expression of IL-6 in chondrocytes of CHI3L1 group was 49.60±0.01 pg/ml, which was higher than that of 47.88±0.01 pg/ml in the control group ( t=132.70, P<0.001). The expression of TNF-α was 95.93±0.02 pg/ml, which was higher than 90.69±0.02 pg/ml in the control group ( t=376.10, P<0.001). There was significant difference in expression of IL-6 in chondrocytes between the CHI3L1 group and the control group ( t=132.72, P<0.001). The expression of TNF-α ( t=376.10, P<0.001) was statistically difference. Protein chip detected 53 proteins with significant differences in expression and 43 proteins with significant differences in protein phosphorylation levels. Bioinformatics analysis was used to identify 16 signaling pathways in which the above different proteins might be involved, including ErbB, PI3K, Akt, Ras, JAK, STAT3, MAPK pathway. In the MAPK pathway, the expression of MAPK1 and RAF1 proteins was higher in the chondrocytes of the CHI3L1 group than in the control group (1.094±0.00 vs. 0.814±0.00, 0.988±0.00 vs. 0.786±0.00; t=103.16, P<0.001; t=54.32, P<0.001). Compared with the control group, the expression of MAPK1 and RAF1 proteins was significantly increased in the chondrocytes of the CHI3L1 group. Conclusion:The expression of CHI3L1 is corrected to articular cartilage degeneration. CHI3L1 is able to inhibit the proliferation of articular chondrocytes, which regulated the cycling of chondrocytes from G1 phase to S phase, promote the apoptosis of chondrocytes, and promote the expression of IL-6 and TNF-α in chondrocytes. Regulation of chondrocytes biological function through the MAPK pathway, which is a potential biomarker for the clinical diagnosis and treatment of lumbar joint degeneration.