OBJECTIVEThis study aimed to investigate the effects of in vitro continuous passaging on the morphological phenotype and differentiation characteristics of mouse hyaline chondrocytes, as well as on the balance of the extracellular matrix (ECM).

METHODSEnzymatic digestion was conducted to isolate mouse hyaline chondrocytes, which expanded over five passages in vitro. Hematoxylin-eosin stain was used to show the changes in chondrocyte morphology. Semi-quantitative polymerase chain reaction was performed to analyze the mRNA changes in the marker genes, routine genes, matrix metalloproteinases (MMPs), and tissue inhibitors of MMPs (TIMPs) in chondrocytes. Zymography was carried out to elucidate changes in gelatinase activities.

RESULTSAfter continuous expansion in vitro, the morphology of round or polygonal chondrocytes changed to elongated and spindled shape. The expression of marker genes significantly decreased (P < 0.05), and it was almost negatively expressed by P5 chondrocytes. By contrast, the down regulation of routine genes was insignificant. The gene expression levels of MMPs and TIMPs both decreased (P < 0.05), but the change in MMP-1 and TIMP-1 was not statistically significant (P > 0.05). Meanwhile, the ratio of MMPs/TIMPs was altered. At the protein level, the activities of gelatinases decreased after passaging, especially for P4 and P5 chondrocytes (P < 0.05).

CONCLUSIONSerially passaged chondrocytes dedifferentiated and lost specific phenotypic characteristics during in vitro expansion culture. Simultaneously, the anabolism and catabolism of the cartilage ECM became uncontrollable and led to the imbalance of ECM homeostasis. When hyaline chondrocytes are applied in research on relevant diseases or cartilage tissue engineering, P0-P2 chondrocytes should be used.

Animals ; Cartilage ; Cell Differentiation ; Cells, Cultured ; Chondrocytes ; physiology ; Cytoskeleton ; Extracellular Matrix ; Gelatinases ; Gene Expression ; Hyalin ; physiology ; Matrix Metalloproteinase 1 ; Matrix Metalloproteinases ; Mice ; RNA, Messenger ; Tissue Engineering ; Tissue Inhibitor of Metalloproteinase-1 ; Tissue Inhibitor of Metalloproteinases

The aim of this study was to investigate the inhibitory effect of heparin/fibronectin (Hep/Fn) complexes on neointimal hyperplasia following endovascular intervention. Hep/Fn complexes were immobilized onto titanium (Ti) surfaces, with subsequent X-ray photoelectron spectroscopy (XPS), Toluidine Blue O (TBO) and immunohistochemistry methods were used to characterize surface properties. Smooth muscle cell (SMC) cultures were used to evaluate the effect of Hep/Fn complexes on SMC proliferation. Results showed that Hep/Fn complexes successfully immobilized onto Ti surfaces and resulted in an inhibition of SMC proliferation. This study suggests that Hep/Fn surface-immobilized biomaterials develop as a new generation of biomaterials to prevent neointimal hyperplasia, particularly for use in cardiovascular implants.
Biocompatible Materials ; Cell Proliferation ; drug effects ; physiology ; Cells, Cultured ; Fibronectins ; chemistry ; pharmacology ; Heparin ; chemistry ; pharmacology ; Humans ; Immobilized Proteins ; chemistry ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; drug effects ; physiology ; Surface Properties ; Titanium ; chemistry ; Umbilical Arteries

Tissue injury provokes a series of events containing inflammation, new tissue formation and tissue remodeling which are regulated by the spatially and temporally coordinated organization. It is an evolutionarily conserved, multi-cellular, multi-molecular process via complex signalling network. Tissue injury disorders present grievous clinical prob-lems and are likely to increase since they are generally associated with the prevailing diseases such as diabetes, hyper-tension and obesity. Although these dynamic responses vary not only for the different types of trauma but also for the different organs, a balancing act between the tissue degradation and tissue synthesis is the same. In this process, the degradation of old extracellular matrix (ECM) elements and new ones' synthesis and deposition play an essential role, especially collagens. Lysyl oxidase (LOX) and four lysyl oxidase-like proteins are a group of enzymes capable of catalyzing cross-linking reaction of collagen and elastin, thus initiating the formation of covalent cross-links that insolubilize ECM proteins. In this way, LOX facilitates ECM stabilization through ECM formation, development, maturation and remodeling. This ability determines its potential role in tissue repair and regeneration. In this review, based on the current in vitro, animal and human in vivo studies which have shown the significant role of the LOXs in tissue repair, e.g., tendon regeneration, ligament healing, cutaneous wound healing, and cartilage remodeling, we focused on the role of the LOXs in inflammation phase, proliferation phase, and tissue remodeling phase of the repair process. By summarizing its healing role, we hope to shed light on the understanding of its potential in tissue repair and provide up to date therapeutic strategies towards related injuries.
Animals ; Cartilage ; Collagen ; Elastin ; Extracellular Matrix ; Hope ; Humans ; In Vitro Techniques ; Inflammation ; Ligaments ; Obesity ; Oxidoreductases* ; Protein-Lysine 6-Oxidase ; Regeneration ; Tendons ; Wound Healing

Objective: To evaluate the effect of tumor shape and location on pulmonary dose-volume parameters by intensity-modulated radiation therapy (IMRT) in patients with non-small cell lung cancer (NSCLC), aiming to provide a reference basis for establishing limits of the pulmonary dose-volume parameters during IMRT. Methods: Clinical data of 208 NSCLC patients undergoing radical IMRT from June 2009 to June 2016 were retrospectively analyzed. According to the tumor shape and location, 208 cases were divided into the vertical bar group (n=127) and the horizontal bar group (n=81), the superior lung group (n=103) and the inferior lung group (n=105). Regression model curve was used to evaluate the effect of tumor shape and location upon the common pulmonary dose-volume parameters(V₅, V₂₀, MLD, AVS₅ and AVS₂₀). Results: In all groups, the fitting curves of V₅, V₂₀ and MLD were manifested in the quadratic equation pattern, and AVS₅ and AVS₂₀ in the logarithmic equation manner. In the vertical bar group, the V₅(P=0.015), V₂₀(P=0.047) and MLD (P=0.012) were significantly higher, whereas the AVS₅(P=0.044) was significantly lower compared with those in the horizontal bar group. No statistical significance was observed in AVS₂₀ between two groups (P=0.490). The tumor location exerted significant effect upon V₅ alone (P=0.009). Conclusions When the tumors presents in the vertical bar shape, the limits of the common lung dose-volume parameters are likely to exceed those of tumors in the vertical bar shape. Lung tumors located in the inferior lobe exerts a more significant effect upon the low-dose region volume compared with the tumors in the superior lobe.

The high- and the low-molecular weight hyaluronic acids (HMW-HA and LMW-HA, respectively) showed different biological activities in inflammation. However, the role of LMW-HA in inflammatory response is controversial. In this study, we aimed to investigate the effect of bioactive hyaluronan (B-HA) on lipopolysaccharide (LPS)-induced inflammatory responses in human macrophages and mice. B-HA was produced from HA treated with glycosylated recombinant human hyaluronidase PH20. Human THP-1 cells were induced to differentiate into macrophages. THP-1-derived macrophages were treated with B-HA, LPS, or B-HA + LPS. The mRNA expression and the production of inflammatory cytokines were determined using quantitative real-time PCR and enzyme-linked immunosorbent assay. The phosphorylation levels of proteins in the nuclear factor-κB (NF-κB), mitogen-activated protein kinase (MAPK), and IRF-3 signaling pathways were measured using Western blot. The in vivo efficacy of B-HA was assessed in a mouse model of LPS-induced inflammation. Results showed that B-HA inhibited the expression of TNF-α, IL-6, IL-1, and IFN-β, and enhanced the expression of the antiinflammatory cytokine IL-10 in LPS-induced inflammatory responses in THP-1-derived macrophages and in vivo. B-HA significantly suppressed the phosphorylation of the TLR4 signaling pathway proteins p65, IKKα/β, IκBα, JNK1/2, ERK1/2, p38, and IRF-3. In conclusion, our results demonstrated that the B-HA attenuated the LPS-stimulated inflammatory response by inhibiting the activation of the TLR4 signaling pathway. B-HA could be a potential anti-inflammatory drug in the treatment of inflammatory disease.
Animals ; Cytokines ; Hyaluronic Acid ; Lipopolysaccharides ; Mice ; NF-kappa B/metabolism* ; Signal Transduction ; Toll-Like Receptor 4

Objective:To explore the coverage of influenza and pneumonia vaccination and factors influencing the vaccination in children.Methods:A cross-sectional questionnaire survey was conducted in children's parents in Beijing and Gansu by using two-stage cluster-sampling to investigate the influenza and pneumonia vaccination rates and influencing factors in children.Results:A total of 2 377 parents were included in the study, and the results indicated that the influenza vaccination coverage was 35.93% and the pneumonia vaccination coverage was 16.58% in children in survey areas, the vaccination rate of both vaccines was 11.65%. The top three reasons for vaccination for both vaccines were being aware of severity of the diseases (influenza vaccine: 36.02%; pneumonia vaccine: 49.61%), being required by school or organization (influenza vaccine: 28.76%; pneumonia vaccine: 25.45%) and being aware of the susceptibility of the diseases (influenza vaccine: 26.41%; pneumonia vaccine: 13.88%). The top three reasons for having no vaccinations were personal unwillingness, concern about vaccine and vaccine accessibility. Families with multi children, living in rural areas and lower family income were the negative factors for both types of vaccinations.Conclusions:The influenza and pneumonia vaccination coverage in children need further improvement, and rural families and families with multi children are the key concern groups for expanding vaccination coverage. Health education about influenza and pneumonia vaccinations, coordinating vaccine supply and decreasing vaccine prices play an important role in improving influenza and pneumonia vaccination coverage.

OBJECTIVES: To screen the DNA methylation loci associated with the age of Han males in northern China and to construct an age estimation model. METHODS: Twenty-one candidate methylation loci were screened. The DNA methylation levels of 476 blood samples from Chinese Han males were detected for 21 amplicons using EpiTYPER technology platform, and data on 153 DNA methylation loci were obtained. RESULTS: After correlation analysis, 8 age-related DNA methylation loci were finally screened. CpG1, CpG2, CpG4, CpG7, CpG8 were located on TRIM59, RASSF5, Clorf132, CSNK1D, ELOVL2,CpG5, CpG6 on PDE4C, and CpG3 on chr17:21452808. Based on the 8 loci, 352 samples were used for model construction. A multivariate linear regression age estimation model was constructed （R²=0.93）, with mean absolute deviation （MAD） of 2.69 years old. When 109 samples were used for model validation, the MAD was 3.80 years old. The test was repeated 3 times in 15 new samples, with MADs of 4.08, 4.68 and 3.93 years old, respectively. CONCLUSIONS The age estimation model on Han males in northern China constructed in this study can be used to estimate the age of victims and suspects and to narrow the scope of investigation, and therefore has practical application value.
Adaptor Proteins, Signal Transducing ; Apoptosis Regulatory Proteins ; Asian People ; Child, Preschool ; China ; CpG Islands ; DNA Methylation ; Humans ; Intracellular Signaling Peptides and Proteins ; Linear Models ; Male ; Membrane Proteins ; Metalloproteins ; Monomeric GTP-Binding Proteins ; Tripartite Motif Proteins

Primary ureteral plasmacytoid carcinomas is a rare tumor with high grade and poor diagnosis. Pathological and immunohistochemical staining play an extremely key role in diagnosis since there is no specific clinical and radiological evidence. The surgical removement is the first line treatment. Herein, we report a case of ureteral plasmacytoid carcinoma that was well controlled with multimodal therapy.

The biophysical properties of the extracellular matrix (ECM) dictate tissue-specific cell behaviour. In the skeleton system, bone shows the potential to adapt its architecture and contexture to environmental rigidity via the bone remodelling process, which involves chondrocytes, osteoblasts, osteoclasts, osteocytes and even peripheral bone marrow-derived stem/stromal cells (BMSCs). In the current study, we generated stiff (~1 014 ± 56) kPa, Young's modulus) and soft (~46 ± 11) kPa silicon-based elastomer polydimethylsiloxane (PDMS) substrates by mixing curing agent into oligomeric base at 1:5 and 1:45 ratios, respectively, and investigated the influence of substrate stiffness on the cell behaviours by characterizing cell spreading area, cell cytoskeleton and cell adhesion capacity. The results showed that the cell spreading areas of chondrocytes, osteoblasts, osteoclasts, osteocytes and BMSCs were all reduced in the soft substrate relative to those in the stiff substrate. F-actin staining confirmed that the cytoskeleton was also changed in the soft group compared to that in the stiff group. Vinculin in focal adhesion plaques was significantly decreased in response to soft substrate compared to stiff substrate. This study establishes the potential correlation between microenvironmental mechanics and the skeletal system, and the results regarding changes in cell spreading area, cytoskeleton and cell adhesion further indicate the important role of biomechanics in the cell-matrix interaction.
Actins ; Cell Adhesion ; Elastic Modulus ; Focal Adhesions ; physiology ; Humans ; Vinculin ; analysis ; metabolism

OBJECTIVE: To analyze the current status of diagnosis and management of acute appendicitis (AA) in China. METHODS: Questionnaire survey was used to retrospectively collect data of hospitalized patients with AA from 43 medical centers nationwide in 2017 (Sort by number of cases provided: Jinling Hospital of Medical School of Nanjing University, The First Affiliated Hospital of Xinjiang Medical University, Lu'an People's Hospital, Tengzhou Central People's Hospital, Dalian Central Hospital, The Affiliated Hospital of Xuzhou Medical University, Dongying People's Hospital, Jinjiang Hospital of Traditional Chinese Medicine, Huangshan Shoukang Hospital, Xuyi People's Hospital, Nanjing Jiangbei People's Hospital, Lanzhou 940th Hospital of PLA, Heze Municipal Hospital, The First College of Clinical Medical Science of China Three Gorges University, Affiliated Jiujiang Hospital of Nanchang University, The Second People's Hospital of Hefei, Affiliated Central Hospital of Shandong Zaozhuang Mining Group, The Third People's Hospital of Kunshan City, Xuzhou First People's Hospital, The 81st Group Army Hospital of PLA, Linyi Central Hospital, The General Hospital of Huainan Eastern Hospital Group, The 908th Hospital of PLA, Liyang People's Hospital, The 901th Hospital of Joint Logistic Support Force, The Third Affiliated Hospital of Chongqing Medical University, The Fourth Hospital of Jilin University, Harbin Acheng District People's Hospital, The First Affiliated Hospital of Zhengzhou University, Nanjing Luhe People's Hospital, Taixing Municipal People's Hospital, Baotou Central Hospital, The Affiliated Hospital of Nantong University, Linyi People's Hospital, The 72st Group Army Hospital of PLA, Zaozhuang Municipal Hospital, People's Hospital of Dayu County, Taixing City Hospital of Traditional Chinese Medicine, Suzhou Municipal Hospital, Beijing Guang'anmen Hospital, Langxi County Hospital of Traditional Chinese Medicine, Nanyang Central Hospital, The Affiliated People's Hospital of Inner Mongolia Medical University).The diagnosis and management of AA were analyzed through unified summary. Different centers collected and summarized their data in 2017 and sent back the questionnaires for summary. RESULTS: A total of 8 766 AA patients were enrolled from 43 medical centers, including 4 711 males (53.7%) with median age of 39 years and 958 (10.9%) patients over 65 years old. Of 8 776 patients, 5 677 cases (64.6%) received one or more imaging examinations, and the other 3 099 (35.4%) did not receive any imaging examination. A total of 1 858 (21.2%) cases received medical treatment, mainly a combination of nitroimidazoles (1 107 cases, 59.8%) doublet regimen, followed by a single-agent regimen of non-nitroimidazoles (451 cases, 24.4%), a nitroimidazole-free doublet regimen (134 cases, 7.2%), a triple regimen of combined nitroimidazoles (116 cases, 6.3%), nitroimidazole alone (39 cases, 2.1%) and nitroimidazole-free triple regimen (3 cases, 0.2%). Of the 6 908 patients (78.8%) who underwent surgery, 4 319 (62.5%) underwent laparoscopic appendectomy and 2589 (37.5%) underwent open surgery. Ratio of laparotomy was higher in those patients under 16 years old (392 cases) or over 65 years old (258 cases) [15.1%(392/2 589) and 10.0%(258/2 589), respectively, compared with 8.5%(367/4 316) and 8.0%(347/4 316) in the same age group for laparoscopic surgery, χ²=91.415, P<0.001; χ²=15.915,P<0.001]. Patients with complicated appendicitis had higher ratio of undergoing open surgery as compared to those undergoing laparoscopic surgery [26.7%(692/2 589) vs. 15.6%(672/4 316), χ²=125.726, P<0.001].The cure rates of laparoscopic and open surgery were 100.0% and 99.8%(2 585/2 589) respectively without significant difference (P=0.206). Postoperative complication rates were 4.5%(121/2 589) and 4.7%(196/4 316) respectively, and the difference was not statistically significant (χ²=0.065, P=0.799). The incidence of surgical site infection was lower (0.6% vs. 1.7%, χ²=17.315, P<0.001), and hospital stay was shorter [6(4-7) days vs. 6(5-8) days, U=4 384 348.0, P<0.001] in the laparoscopic surgery group, while hospitalization cost was higher (median 12 527 yuan vs. 9 342 yuan, U=2 586 809.0, P<0.001). CONCLUSIONS The diagnosis of acute appendicitis is still clinically based, supplemented by imaging examination. Appendectomy is still the most effective treatment at present. Laparoscopic appendectomy has become the main treatment strategy, but anti-infective drugs are also very effective.
Acute Disease ; Adolescent ; Adult ; Aged ; Anti-Bacterial Agents ; therapeutic use ; Appendectomy ; Appendicitis ; diagnosis ; therapy ; China ; Female ; Health Care Surveys ; Humans ; Laparoscopy ; Male ; Middle Aged ; Retrospective Studies ; Treatment Outcome ; Young Adult