Additive Manufacturing of Biodegradable Molybdenum - From Powder to Vascular Stent.

Magnesium, iron and zinc based biodegradable metals are widely recognized as promising candidate materials for the next generation of bioresorbable stent (BVS). However, none of those metal BVSs are perfect at this stage. Here, we developed a brand-new BVS based on a novel biodegradable metal (Molybdenum, Mo) through additive manufacturing. Nearly full-dense and crack-free thin-wall Mo was directly manufactured through selective laser melting (SLM) with fine Mo powder. Systemic analyses considering the forming quality, wall-thickness, microstructure, mechanical properties and degradation behaviors were performed. Then, Mo-based thin-strut (≤ 100 µm) stents were successfully obtained through an optimized single-track laser melting route. The SLMed thin-wall Mo owns comparable strength to its Mg and Zn based counterparts (as-drawn), whilst, it exhibits remarkable biocompatibility in vitro. Vessel related cells are well adhered and spread on SLMed Mo, and it exhibits a low risk of hemolysis and thrombus. The SLMed stent is compatible to vessel tissues in rat abdominal aorta, and it can provide sufficient support in an animal model as an extravascular stent. This work possibly opens a new era of manufacturing Mo-based stents through additive manufacturing. This article is protected by copyright. All rights reserved.

Intracellular hydroxyproline imprinting following resolution of bleomycin-induced pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease with few treatment options. The poor success in developing anti-IPF strategies has impelled researchers to reconsider the importance of the choice of animal model and assessment methodologies. Currently, it is still not settled whether the bleomycin-induced lung fibrosis mouse model finally returns to resolution. METHODS: This study aimed to follow the dynamic fibrotic features of bleomycin-treated mouse lungs over extended durations through a combination of the latest technologies (micro-computed tomography imaging and histological detection of degraded collagens) and traditional methods. In addition, we also applied immunohistochemistry to explore the distribution of all hydroxyproline-containing molecules. RESULTS: As determined by classical biochemical methods, total lung hydroxyproline contents reached a peak at 4 weeks after bleomycin injury and maintained a steady high level thereafter until the end of the experiments (16 weeks). This result seemed to partially contradict with the changes of other fibrosis evaluation parameters, which indicated a gradual degradation of collagens and a recovery of lung aeration after the fibrosis peak. This inconsistency was well reconciled by our data from immunostaining against hydroxyproline and fluorescent peptide staining against degraded collagen, together showing large amounts of hydroxyproline-rich degraded collagen fragments detained and enriched within the intracellular regions at 10 or 16 weeks rather than at 4 weeks after bleomycin treatment. CONCLUSIONS: Our present data not only offer respiratory researchers a new perspective towards the resolution nature of mouse lung fibrosis, but also remind them to be cautious when using the hydroxyproline content assay to evaluate the severity of fibrosis.

A novel prognostic signature for idiopathic pulmonary fibrosis based on five-immune-related genes.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a highly fatal lung disease of unknown etiology with a median survival after diagnosis of only 2-3 years. Its poor prognosis is due to the limited therapy options available as well as the lack of effective prognostic indicators. This study aimed to construct a novel prognostic signature for IPF to assist in the personalized management of IPF patients during treatment. METHODS: Differentially-expressed genes (DEGs) in IPF patients versus healthy individuals were analyzed using the "limma" package of R software. Immune-related genes (IRGs) were obtained from the ImmPort database. Univariate Cox regression analysis was adopted to screen significantly prognostic IRGs for IPF patients. Multiple Cox regression analysis was used to identify optimal prognostic IRGs and construct a prognostic signature. RESULTS: Compared with healthy individuals, there were a total of 52 prognosis-related DEGs in the bronchoalveolar lavage (BAL) samples of IPF patients, of which 37 genes were identified as IRGs. Of these, five genes (CXCL14, SLC40A1, RNASE3, CCR3, and RORA) were significantly associated with overall survival (OS) in IPF patients, and were utilized for establishment of the prognostic signature. IPF patients were divided into high- and low-risk groups based on the prognostic signature. Marked differences in the OS probability were observed between high- and low-risk IPF patients. The area under curves (AUCs) of the receiver operating characteristic (ROC) curve for the prognostic signature in the training and validation cohorts were 0.858 and 0.837, respectively. The expression levels between RNASE3 and SLC40A1 (P<0.01, r=0.394), between RORA and CXCL14 (P<0.01, r=-0.355), between CCR3 and CXCL14 (P<0.01, r=0.258), as well as between RNASE3 and CCR3 (P<0.01, r=0.293) were significantly correlated. CONCLUSIONS: We developed a validated and reproducible IRG-based prognostic signature that should be helpful in the personalized management of patients with IPF, providing new insights into the relationship between the immune system and IPF.

Clinical Characteristics and Prognosis of Patients With Pulmonary Mucoepidermoid Carcinoma: A SEER-Based Analysis.

BACKGROUND: Primary pulmonary mucoepidermoid carcinoma (PMEC) is an extremely rare malignancy. Its clinical characteristics and prognosis are not fully understood. This study evaluated clinical characteristics and prognostic factors of PMEC and established a nomogram to predict its 1-, 3-, 5- and 10-year cancer-specific survival (CSS) rates. METHODS: In the Surveillance, Epidemiology, and End Results database from January 1, 2016 to December 31, 2016, patients pathologically diagnosed with PMEC were identified. Kaplan-Meier analysis and Cox regression were performed to evaluate the CSS stratified by different covariates. A predictive nomogram model was built and validated by the concordance index (C-index) and calibration curves. RESULTS: A total of 585 PMEC patients were identified. A total of 408 (70%) of patients were placed into the training cohort, and 177 (30%) patients were placed into the validation cohort. The 5- and 10-year CSS rates of stage I-II PMEC patients were 91.4 and 88.9, respectively. The 1-, 3- and 5-year CSS rates of stage III-IV PMEC were 56.5, 39.45, and 32.1%, respectively. Survival curves showed that older age, large tumor size, poor differentiation, and high TNM stage were associated with a significantly worse prognosis. CSS outcomes were significantly better in patients who received surgical treatments (surgical alone, surgery plus radiation and/or chemotherapy). Patients who received radiation and/or chemotherapy had the worst prognosis. Multivariate Cox results revealed that covariates, including age, tumor laterality, tumor sizes, pathological differentiation, lymph node metastasis, distant metastasis, TNM stage and therapy, were independent prognostic factors for PMEC. These factors were used to construct a nomogram. The C-index of the nomogram was 0.921. The calibration curve presented favorable consistency between the predicted CSS and actual observations. This nomogram was validated by the validation cohort. The C-index of the validation cohort was 0.968. CONCLUSION: Age, bilateral tumors, tumor size, pathological differentiation grade, lymph node metastasis, distant metastasis, TNM stage and therapy were independent prognostic factors of PMEC patients. The first nomogram for predicting the CSS of PMEC was built and validated, showing its potential value in practice.

Inhibition of miR-199a-5p rejuvenates aged mesenchymal stem cells derived from patients with idiopathic pulmonary fibrosis and improves their therapeutic efficacy in experimental pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is an age-related disease with no cure. Mesenchymal stem cell (MSC)-based therapy has emerged as a novel strategy for IPF treatment. Nevertheless, MSCs derived from patients with IPF (IPF-MSCs) become senescent, thereby reducing their beneficial effects in IPF. MicroRNAs (miRNAs) mediate the senescence of MSCs, but the underlying mechanisms are not fully understood. We investigated the mechanisms by which miR-199a-5p regulates IPF-MSC senescence and whether its inhibition could rejuvenate IPF-MSCs and enhance their therapeutic efficacy. METHODS: Control-MSCs and IPF-MSCs were isolated from the adipose tissue of age-matched healthy and IPF donors, respectively. Cell senescence was examined by senescence-associated ß-galactosidase (SA-ß-gal) staining. The level of miR-199a-5p was measured by RT-PCR. Autophagy was determined using a transmission electron microscope (TEM). The therapeutic efficacy of anti-miR-199a-5p-IPF-MSCs was assessed using a mouse model of bleomycin-induced lung fibrosis. RESULTS: Despite similar surface makers, IPF-MSCs exhibited increased cellular senescence and decreased proliferative capacity compared with control-MSCs. The expression of miR-199a-5p was significantly enhanced in the serum of IPF patients and IPF-MSCs compared with that of healthy donors and control-MSCs. The upregulation of miR-199a-5p induced senescence of control-MSCs, whereas the downregulation rescued IPF-MSC senescence. Mechanistically, miR-155-5p suppressed autophagy of MSCs via the AMPK signaling pathway by downregulating the expression of Sirtuin 1(Sirt1), resulting in cellular senescence. Accordingly, miR-155-5p inhibition promoted autophagy and ameliorated IPF-MSC senescence by activating the Sirt1/AMPK signaling pathway. Compared with IPF-MSCs, the transplantation of anti-miR-199a-5p-IPF-MSCs increased the ability to prevent progression of pulmonary fibrosis in bleomycin-treated mice. CONCLUSIONS: Our study shows that miR-199a-5p regulates MSC senescence in patients with IPF by regulating the Sirt1/AMPK signaling pathway and miR-199a-5p is a novel target to rejuvenate IPF-MSCs and enhance their beneficial effects.

Pulmonary fibrosis alters gut microbiota and associated metabolites in mice: An integrated 16S and metabolomics analysis.

AIMS: The "gut-lung axis" reflects intimate connection and bidirectional effect between gut and lung, involving numerous lung diseases. Pulmonary fibrosis is a progressive interstitial lung disease with high fatality rate, so far, its association with gut remains unexplored. We investigated the correlation between pulmonary fibrosis and gut microbiota. MATERIALS AND METHODS: We collected feces from two pulmonary fibrotic models respectively, and performed a combinatory study using 16S rDNA sequencing and non-targeted metabonomics. Correlation matrix was used to indicate the correlation between microbiome, metabolites and fibrotic indicators, and the possibility of gut microbiota in identifying pulmonary fibrosis was assessed by ROC analysis. KEY FINDINGS: 412 genera of microflora and 26 kinds of metabolites were synchronously altered with same trend in two models but differed observably with control. Among these, 7 microorganisms and 9 metabolites were the typical representatives, which were correlated significantly and highly correlated with fibrotic indicators shown by correlation matrix. ROC analysis indicated that it was dependable to identify pulmonary fibrosis by using gut microorganisms and metabolites in both models (AUC > 0.85, p < 0.01). SIGNIFICANCE: In summary, our findings first revealed a previously unknown correlation between gut and pulmonary fibrosis in mouse models, which creates novel insights of the interaction between pulmonary fibrosis and gut microbiota.

Serpina3n is closely associated with fibrotic procession and knockdown ameliorates bleomycin-induced pulmonary fibrosis.

OBJECTIVE: Pulmonary fibrosis is a fatal interstitial lung disease that is characterized by excessive accumulation of extracellular matrix (ECM) and remodeling of lung. The precise mechanisms underlying pulmonary fibrosis still remain unclear. In the current study, we aimed to investigate the alteration and function of serine (or cysteine) peptidase inhibitor, clade A, member 3 N (Serpina3n) in pulmonary fibrotic models and explore the potential mechanisms. METHODS: We induced pulmonary fibrosis in mice by silica and bleomycin respectively and determined Serpina3n in lung tissues, and then verified the expression of Serpina3n and its correlation with pulmonary fibrosis at seven time points in a bleomycin longstanding model. Moreover, adeno-associated virus type 9 (AAV9)-mediated Serpina3n knockdown was used to treat pulmonary fibrosis in the bleomycin model, whose possible mechanisms would be preliminarily explored by detecting chymotrypsin C as an example. RESULTS: Serpina3n was up-regulated significantly in lungs of both models at mRNA and protein levels relative to control. Notably, the expression of Serpina3n peaked during the 3rd week and then decreased until nearly normal levels during the 10th week, which was closely related to fibrotic procession in bleomycin-treated mice. AAV-mediated Serpina3n knockdown in the lung tissues alleviated bleomycin-induced fibrotic symptoms at various levels and disinhibit chymotrypsin C. CONCLUSIONS: Our study revealed that Serpina3n is a critical regulator in pulmonary fibrosis and suggested Serpina3n inhibition as a potential therapeutic strategy in chronic pulmonary injuries.

Increased Specific Labeling of INS-1 Pancreatic Beta-Cell by Using RIP-Driven Cre Mutants with Reduced Activity.

Ectopically expressed Cre recombinase in extrapancreatic tissues in RIP-Cre mice has been well documented. The objective of this study was to find a simple solution that allows for improved beta-cell specific targeting. To this end, the RIP-Cre and reporter CMV-loxP-DsRed-loxP-EGFP expression cassettes were configurated into a one-plasmid and two-plasmid systems, which labeled approximately 80% insulin-positive INS-1 cells after 48 h transfection. However, off-target labeling was robustly found in more than 15% insulin-negative Ad293 cells. When an IRES element was inserted in front of Cre to reduce the translation efficiency, the ratio of recombination between INS-1 and Ad293 cells increased 3-4-fold. Further, a series of Cre mutants were generated by site-directed mutagenesis. When one of the mutants, Cre(H289P) in both configurations, was used in the experiment, the percentage of recombination dropped to background levels in a number of insulin-negative cell lines, but decreased only slightly in INS-1 cells. Consistently, DNA substrate digestion assay showed that the enzymatic activity of Cre(H289P) was reduced by 30-fold as compared to that of wild-type. In this study, we reported the generation of constructs containing RIP and Cre mutants, which enabled enhanced beta-cell specific labeling in vitro. These tools could be invaluable for beta-cell targeting and to the study of islet development.