Electropolishing influence on biocompatibility of additively manufactured Ti-Nb-Ta-Zr: in vivo and in vitro.

Balling defect of the additively manufactured titanium lattice implants easily leads to muscle tissue rejection, which might cause failure of implantation. Electropolishing is widely used in surface polishing of complex components and has potential to deal with the balling defect. However, a clad layer could be formed on the surface of titanium alloy after electropolishing, which may affect the biocompatibility of the metal implants. To manufacture lattice structured ß-type Ti-Ni-Ta-Zr (TNTZ) for bio-medical applications, it is necessary to investigate the impact of electropolishing on material biocompatibility. In this study, animal experiments were conducted to investigate the in vivo biocompatibility of the as-printed TNTZ alloy with or without electropolishing; and proteomics technology was used to elaborate the results. The following conclusions were drawn: (a) a 30% oxalic acid electropolishing treatment was effective in solving balling defects, and ~21 nm amorphous clad layer would be formed on the surface of the material after polishing; (b) the electropolished TNTZ suggested decreased cell cytotoxicity and improved blood biocompatibility as compared to as-printed TNTZ; (c) the amorphous clad layer could make a barrier to prevent Ta and Zr ions from penetrating into the muscle tissue, and could form a good tissue regeneration at the implantation site during 4 weeks, indicating that the electropolished TNTZ has the potential as implants; and (d) the cells attached to the electropolished TNTZ showed higher antioxidant capacity but less proliferation than attached to as-printed TNTZ.

Research on function and mechanisms of a novel small molecule WG449E for hypertrophic scar.

BACKGROUND: Hypertrophic scars are complications of severe wound healing characterized by excessive fibrosis associated with aberrant function of fibroblasts. However, no available drugs can be utilized to effectively treat these scars. The transforming growth factor ß (TGFß) signalling pathway regulates collagen synthesis and plays an important role in scar formation. OBJECTIVES: To evaluate the anti-scar effects of TGFß inhibitors in vitro and in vivo. METHODS: Col1α2-luciferase reporter assay was used to screen the compounds suppress type I collagen gene transcription. Sulforhodamine B colorimetric assay and colony formation assay were used to test the compound's effect on cell proliferation. Wound healing and transwell assay were performed to test the cell migration and invasion. Western blotting, immunofluorescence, immunohistochemistry and Q-PCR assay were used to determine the protein and mRNA levels. 3D cell contraction assay was used to examine the cell contraction. Flow cytometry was performed to analyse cell apoptosis. Masson stain, H&E stain and immunochemistry were used to analyse the scar formation in vivo. RESULTS: WG449E, as one of the most potent inhibitors, was identified to significantly downregulate the mRNA and protein levels of collagen in hypertrophic scar-derived fibroblasts through inhibiting Smad2/3 phosphorylation. WG449E inhibited the proliferation, migration and contraction of fibroblasts in vitro and in vivo. In addition, WG449E induced cell apoptosis through the activation of cleaved-caspase3. Moreover, WG449E significantly attenuated hypertrophic scar formation and collagen deposition in a mechanical load-induced mouse model. CONCLUSIONS: WG449E is a potential candidate for the treatment of hypertrophic scars.WG449E downregulates the mRNA and protein levels of collagen in hypertrophic scar-derived fibroblasts through inhibiting Smad2/3 phosphorylation and nucleic localization. WG449E blocks HSF migration and invasion by regulating F-actin assignment. In addition, WG449E induces HSF apoptosis through the activation of cleaved-caspase3.

Effects of habitat fragmentation on the genetic diversity and differentiation of Dendrolimus punctatus (Lepidoptera: Lasiocampidae) in Thousand Island Lake, China, based on mitochondrial COI gene sequences.

Thousand Island Lake (TIL) is a typical fragmented landscape and an ideal model to study ecological effects of fragmentation. Partial fragments of the mitochondrial cytochrome oxidase subunit I gene of 23 island populations of Dendrolimus punctatus in TIL were sequenced, 141 haplotypes being identified. The number of haplotypes increased significantly with the increase in island area and shape index, whereas no significant correlation was detected between three island attributes (area, shape and isolation) and haplotype diversity. However, the correlation with number of haplotypes was no longer significant when the 'outlier' island JSD (the largest island) was not included. Additionally, we found no significant relationship between geographic distance and genetic distance. Geographic isolation did not obstruct the gene flow among D. punctatus populations, which might be because of the high dispersal capacity of this pine moth. Fragmentation resulted in the conversion of large and continuous habitats into isolated, small and insular patches, which was the primary effect on the genetic diversity of D. punctatus in TIL. The conclusion to emphasize from our research is that habitat fragmentation reduced the biological genetic diversity to some extent, further demonstrating the importance of habitat continuity in biodiversity protection.

Extracellular vesicles promote esophageal cancer progression by delivering lncZEB1-AS1 between cells.

OBJECTIVE: To explore the expression of extracellular vesicle-derived lncZEB1-AS1 in esophageal cancer and its role in esophageal cancer progression. PATIENTS AND METHODS: The extracellular vesicles (EVs) from esophageal cancer patients (n = 26) and normal subjects (n = 26) were isolated by differential centrifugation. The expression of lncZEB1-AS1 in EVs was detected by Real-time PCR (polymerase chain reaction). The clinical data of normal subjects and patients were analyzed. In addition, the concentration of EVs and lncZEB1-AS1 in blood samples from normal subjects and esophageal cancer patients were assessed. After co-culture of esophageal cancer cell line EC109 and EVs with or without lncZEB1-AS1 knockdown, cell proliferation was detected by CCK-8 assay. The possible target microRNAs of lncZEB1-AS1 in cytoplasm were predicted with miRcode, followed by correlation analysis of lncZEB1-AS1 and miR-214. Through literature review, lncZEB1-AS1 positively regulates ZEB1 expression, which was consistent with our result. RESULTS: Quantitative Real-time PCR showed that the serum levels of EVs and the content of lncZEB1-AS1 in EVs from esophageal cancer patients were significantly higher than those in normal controls. LncZEB1-AS1 was overexpressed in esophageal cancer cells co-cultured with EVs of esophageal cancer patients. CCK-8 results indicated that EC109 cells co-cultured with EVs of esophageal cancer patients had stronger proliferative capacity. miRcode showed that miR-214 ranked the first of microRNAs that lncZEB1-AS1 might target, and miR-214 expression was significantly increased after lncZEB1-AS1 knockdown in EC109. After overexpressing lncZEB1-AS1 in EC109 or co-culturing EVs of esophageal cancer patients with EC109 cells, we found that lncZEB1-AS1 positively regulates ZEB1. In contrast, interfering with the expression of lncZEB1-AS1 in esophageal cancer cell lines can effectively reduce the expression of ZEB1. CONCLUSIONS: EVs in the peripheral blood from esophageal cancer patients promote esophageal cancer progression by delivering lncZEB1-AS1 to esophageal cancer cells and targeting miR-214.

USP4 inhibits p53 and NF-κB through deubiquitinating and stabilizing HDAC2.

Histone deacetylases (HDACs) are major epigenetic modulators involved in a broad spectrum of human diseases including cancers. As HDACs are promising targets of cancer therapy, it is important to understand the mechanisms of HDAC regulation. In this study, we show that ubiquitin-specific peptidase 4 (USP4) interacts directly with and deubiquitinates HDAC2, leading to the stabilization of HDAC2. Accumulation of HDAC2 in USP4-overexpression cells leads to compromised p53 acetylation as well as crippled p53 transcriptional activation, accumulation and apoptotic response upon DNA damage. Moreover, USP4 targets HDAC2 to downregulate tumor necrosis factor TNFα-induced nuclear factor (NF)-κB activation. Taken together, our study provides a novel insight into the ubiquitination and stability of HDAC2 and uncovers a previously unknown function of USP4 in cancers.

Varicocele anatomy during subinguinal microsurgical varicocelectomy in Chinese men.

Knowledge of subinguinal microsurgical varicocelectomy is of fundamental importance to ensure that varicocele is resolved and testicular function is preserved. Our study aimed to describe the number of veins, arteries and lymphatics in the subinguinal spermatic cord and to clarify their differences between two sides, between patients with different complaints and between varicoceles with different clinical grades. A total of 102 consecutive patients underwent 162 primary subinguinal microsurgical varicocelectomies, during which the number of vessels with different diameters was recorded. A mean number of 12.9 internal spermatic veins, 0.9 external spermatic veins, 1.8 internal spermatic arteries and 2.9 lymphatics were identified per cord. 88.2% of the internal spermatic arteries were surrounded by a dense complex of adherent veins. The external spermatic vein or veins were found in 49.4% of the cases. The mean number of medium (1-3 mm in diameter) internal spermatic veins on the left was larger than that on the right (P < 0.001). The mean number of medium internal spermatic veins in grade III varicocele was larger than that in grade I or grade II (P < 0.015). There was no significant anatomical difference between the men presenting for infertility, chronic testicular pain and both the two complaints.

Lentivirus-mediated knockdown of tumor protein D52-like 2 inhibits glioma cell proliferation.

TPD52L2 (tumor protein D52-like 2) is a member of TPD52 family which has been implicated in multiple human cancers. Recently, TPD52 protein was shown to be associated with several malignancies, but very little is known about the function of TPD52L2 in cancers, especially in glioma to date, and its roles in glioma occurrence and progression remain to be elucidated. In the present study, we employed lentivirus-mediated RNA interference (RNAi) to knock down TPD52L2 expression in human glioma cell line U251. We found that knockdown of TPD52L2 significantly not only inhibited cell proliferation and colony formation, but also induced G0/G1 cell cycle arrest in vitro. Taken together, these findings suggest that TPD52L2 might play an important role in glioma tumorigenesis.

The role of LAIR-1 (CD305) in T cells and monocytes/macrophages in patients with rheumatoid arthritis.

The LAIR-1 receptor is expressed on a majority of mononuclear leukocytes. It is used as a biomarker when testing synovial fluid for evidence of rheumatoid arthritis (RA). The primary objective of this study was to measure T cell- and monocyte/macrophage-specific LAIR-1 expression in RA patients and compare this to LAIR-1 expression in osteoarthritis (OA) patients and healthy individuals. LAIR-1 expression was significantly decreased in circulating CD4(+) T cells in RA patients compared to both OA patients and healthy individuals. In contrast, LAIR-1 is high in CD14(+) monocytes and local CD68(+) macrophages in synovial tissues from RA patients. Upon stimulation with TNF-α, LAIR-1 expression decreased in T-helper (Th)1 and Th2 CD4(+) T cells from healthy donors. These results indicate that LAIR-1 may exert different functions on T cells and monocytes/macrophages and suggest that LAIR-1 may be a novel therapeutic target for the treatment of RA.

Sonographic appearance of cervical lymphadenopathy due to infectious mononucleosis in children and young adults.

AIM: To depict the grey-scale and Doppler features of cervical lymphadenopathy due to infectious mononucleosis (IM) and to compare the findings with other benign conditions and lymphoma. MATERIALS AND METHODS: One hundred and four patients <30 years old with 138 enlarged lymph nodes (LNs) were enrolled for sonographic analysis. These LNs were grouped as: IM LNs (59 LNs in 30 patients), lymphoma (30 LNs in 30 patients), bacterial lymphadenitis (24 LNs in 20 patients), tuberculosis (TB; 14 LNs in 13 patients), and reactive hyperplasia (11 LNs in 11 patients). Sonographic assessments included shape, echotexture, hilum, border, matting, cystic necrosis, calcification, and vascular pattern. For each sonographic feature, Fisher's exact test was performed to determine whether the difference between IM LNs and any another aetiology were statistically significant. RESULTS: IM LNs tended to be round in shape (69%), heterogeneous in echotexture (61%), absent of echogenic hilum (66%), indistinct margins (80%), bilateral distribution (91%), and matting (83%) [even bilateral matting (66%)], and central hilar vascularity (89.8%). On analysis, bilateral matting had the highest specificity to IM LNs; however, its sensitivity was relatively low. In contrast to IM LNs, TB LNs were more likely to have unilateral matting, cystic necrosis, and calcification. Indistinct margins and decreased echogenicity of the hilum were more frequently seen in IM LNs than in bacterial LNs. Furthermore, central hilar vascularity was a common feature of IM LNs and other benignity, which can distinguish these from lymphoma and TB LNs. CONCLUSION: Although an individual sonographic feature had considerable overlaps between IM LNs and other aetiologies, the combination of several features may be helpful in the diagnosis of IM.

JNK phosphorylation promotes natural degeneration of cervical endplate chondrocytes by down-regulating expression of ANK.

BACKGROUND: Endplate degeneration leads to accelerated degeneration of the intervertebral disc. The importance of endplate chondrocytes in this process is unclear. Many cellular processes in chondrocytes are controlled by activated c-Jun N-terminal kinases (JNK) and protein kinase B (AKT). However, the involvement of their pathways in the degeneration process needs to be elucidated. AIM: To study activation of JNK and AKT signaling pathways and their significance for degeneration of endplate chondrocytes, as well as involvement of progressive ankylosis protein (ANK) in this process. MATERIALS AND METHODS: Rat primary chondrocytes were grown to confluence and subcultured until passage 4. Morphological appearances (microscope, hematoxylin & eosin staining, toluidine blue staining) and proliferation rates of cells (MTT test) were observed. Further, levels of type II collagen, aggrecan, phosphorylated JNK and AKT, total JNK, AKT and ANK were evaluated by qPCR, flow cytometry and Western blot assays. Furthermore, inhibition experiments with SP600125, the JNK inhibitor, were carried out in the passage 4 cells to assess the effects of the JNK pathway on natural degeneration of endplate chondrocytes. RESULTS: The proliferative speed of endplate chondrocytes progressively decreased during passaging. Expressions of type II collagen and aggrecan were significantly decreased with cells at higher passages. Furthermore, phosphorylation of JNK, but not AKT, was significantly up-regulated and accompanied by reduced ANK expression. Inhibition of the JNK pathway increased expression of type II collagen, aggrecan and ANK and facilitated proliferation rates. CONCLUSIONS: Phosphorylation of JNK promotes natural degeneration of cervical endplate chondrocytes, likely by down-regulating ANK expression.

Expression and clinical significance of SHP2 in gastric cancer.

OBJECTIVES: To investigate the expression and clinical significance of the protein tyrosine phosphatase, nonreceptor type 11 (PTPN11 or SHP2) gene, which encodes Src homology 2 domain-containing phosphatase (SHP-2) in gastric cancer. METHODS: SHP2 expression was detected by immunohistochemical staining and real-time quantitative reverse transcription-polymerase chain reaction in tissue samples of normal gastric mucosa and different grades of gastric cancer. Correlation between SHP2 expression and standard clinico pathological parameters was analysed. RESULTS: Immunohistochemical staining revealed significantly higher rates of SHP2 expression in gastric cancer tissues (72.5%) versus normal gastric mucosa (21.9%). SHP-2 mRNA levels were also significantly higher in gastric cancer tissues versus normal gastric mucosa. SHP2 expression correlated significantly with tumour differentiation, clinical classification and lymph node metastases, but was independent of sex and age. CONCLUSIONS: SHP-2 is upregulated in gastric cancer and may be related to the development of gastric cancer. SHP-2 may be a potential prognostic marker of, or a therapeutic target for, gastric cancer.

Contrast-enhanced ultrasound of hepatocellular carcinoma: Correlation of washout time and angiogenesis.

BACKGROUND: The use of the pure blood pool ultrasound contrast agent SonoVue (Bracco, Italy) with specific ultrasound imaging software has enabled the dynamic visualization of tumor microcirculation. AIM: The present study was designed to investigate the washout time of hepatocellular carcinoma (HCC) and correlate it with angiogenesis parameters. METHODS: Thirty-one surgically confirmed HCC cases were prospectively evaluated with contrast-enhanced ultrasound (CEUS), and parameters such as wash-in time, peak enhancement time and washout time were determined offline. We also calculated microvessel density and the percentage of microvessel area (MVA) and compared CEUS parameters between a well differentiated group and a poorly to moderately differentiated group. The Spearman rank order correlation method was used to analyze the relationship between washout time and angiogenesis parameters. RESULTS: The washout time was longer in well differentiated HCC patients compared to those with poorly to moderately differentiated HCC (p < 0.05). In addition, the washout time of HCC was positively correlated with the percentage of MVA (r = 0.510). CONCLUSIONS: Given that the percentage of MVA was positively correlated with tumor blood volume, washout time may be associated with HCC blood volume.

PADI4 gene polymorphism is not associated with ankylosing spondylitis in Chinese Han population.

The cause of ankylosing spondylitis (AS) remains unelucidated. Both genetic and environmental factors are suspected playing an important role in AS development. Peptidyl arginine deiminase type IV (PADI4) is a member of gene family that encodes enzymes responsible for the conversion of arginine to citrulline residues. A strong linkage between PADI4 polymorphism and rheumatoid arthritis has been found in Japanese and Korean patients, but there is no association study about PADI4 in AS. We speculated that PADI4 may be a pivotal gene for AS development. So we investigated the PADI4 polymorphisms in AS in Chinese Han population. A total of 316 Chinese AS patients of Han nationality and 439 healthy controls were recruited. Five single-nucleotide polymorphisms (SNP), PADI4-89 (rs11203366), PADI4-90 (rs11203367), PADI4-92 (rs874881) PADI4-94 (rs2240340) and PADI4-104 (rs1748033), of PADI4 gene were selected, and the major allele frequencies between cases and controls were assessed as 0.571 versus 0.597, 0.565 versus 0.585, 0.565 versus 0.574, 0.446 versus 0.421 and 0.614 versus 620, respectively. No significant differences in the frequency of PADI4 alleles and genotypes between the cases and controls were observed. Two haplotypes ACGGC and GTCGC were significant with AS even after Bonferroni's correction but were with a tiny frequency in AS cases as 1.0% and 1.2%, respectively. These results indicate that PADI4 polymorphisms may not play an important role in the development of AS in Chinese Han population.

Synthesis of carboxylated chitosan and its adsorption properties for cadmium (II), lead (II) and copper (II) from aqueous solutions.

Carboxylated chitosan (CKCTS) was prepared for the removal of Cd(II), Pb(II), and Cu(II) from aqueous solutions. The effects of experimental parameters such as pH value, initial concentration, contact time and temperature on the adsorption were studied. From the results we can see that the adsorption capacities of Cd(II), Pb(II), and Cu(II) increase with increasing pH of the solution. The kinetic rates were best fitted to the pseudo-second-order model. The adsorption equilibrium data were fitted well with the Langmuir isotherm, which revealed that the maximum adsorption capacities for monolayer saturation of Cd(II), Pb(II), and Cu(II) were 0.555, 0.733 and 0.827 mmol/g, respectively. The adsorption was an exothermic process.

Association of HLA genes with ankylosing spondylitis in Han population of eastern China.

Ankylosing spondylitis (AS) is a chronic inflammatory disorder with a multifactorial genetic basis. HLA-B27 was reported with the greatest susceptibility to AS but did not act alone. The aim of this study was to search for other gene(s) associated with AS independently of HLA-B27 using 13 microsatellite markers spanning 1.5 Mb from locus TAP1 to HLA-Cw and a single-nucleotide polymorphism marker within NFkappaBIL1 gene promoter. Genotyping for microsatellites was performed in 175 AS patients of eastern Chinese and 219 ethnically matched healthy controls using polymerase chain reaction with fluorescence-labelled primers, whereas the SNP marker was genotyped by direct DNA sequencing. Allele as well as haplotype frequencies were compared between cases and controls, and a linkage disequilibrium analysis was performed to estimate the LD relationship between the candidate regions. The frequencies of alleles D6S2811*128, STR_MICA*A5.1 and D6S2672*109, as well as haplotypes D6S2811*128-D6S2927*213-D6S2810*340, D6S2927* 221-D6S2810*350-MICA*A5.1, and D6S2810*350-MICA*A5.1-D6S2800* 136 were significantly increased in B27-positive AS patients when compared with B27-positive controls. The results indicated that there may be other gene(s) within the HLA region, especially around locus HLA-B or HLA-Cw, with susceptibility to AS independently of HLA-B27.

Effects of a single nucleotide polymorphism on the expression of human tumor necrosis factor-alpha.

Tumor necrosis factor (TNF)-alpha plays a prominent role in inflammations and is a proinflammatory cytokine that has been implicated in the pathogenesis of autoimmune and infectious diseases. Recent association studies have found that the TNF-alpha-857T allele was associated with several disorders. Here we demonstrate, with reporter genes under the control of the two allelic TNF-alpha promoters, that the minor allele -857T is a much stronger transcriptional activator than the major allele -857C in RAW264.7 cell line in response to lipopolysaccharide stimulation. However, the result was not consistent in HeLa cell line. Furthermore, for the quantitative analysis of TNF-alpha synthesis between the -857C/C genotype from healthy subjects and the -857C/T genotype from AS patients, the quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay were performed separately. There was no significant difference between the two groups at the level of mRNA and protein. These results show that this polymorphism may have a direct effect on TNF-alpha regulation in a tissue-specific manner, and apart from the polymorphism at -857 in the TNF-alpha promoter, there may be other factors affecting the expression of TNF-alpha.

Separation and determination of ephedrine alkaloids and tetramethylpyrazine in Ephedra sinica Stapf by gas chromatography-mass spectrometry.

A simple, sensitive, and reliable method using gas chromatography (GC)-mass spectrometry (MS) is developed for the simultaneous determination of ephedrine alkaloids and 2,3,5,6-tetramethylpyrazine (TMP) in Ephedra sinica Stapf. The sample is extracted with ethyl ether and submitted to GC-MS for identification and quantitation without derivatization. The column used for GC is an HP-5 (30.0 m x 250 microm x 0.25 microm, 5% phenyl methyl siloxane), and the carrier gas is helium. The detection limits for ephedrine, pseudoephedrine, and TMP are 0.4 ng 0.7 ng, and 0.02 ng (signal-to-noise ratio of 3), respectively. The reproducibility of the total procedure is proved to be acceptable (RSD < 2%), and the recoveries are above 93%.