Inhibition of UGT1A1*1 and UGT1A1*6 catalyzed glucuronidation of SN-38 by silybins.

UGT1A1 is the main enzyme that catalyzes the metabolic elimination and detoxification of SN-38, the active form of the drug irinotecan. Milk thistle products have been used widely to protect the liver from injury associated with the use of chemotherapeutic agents. To evaluate whether SN-38 metabolism can be affected by milk thistle products, the inhibitory effects of silybins on UGT1A1*1 and UGT1A1*6 were evaluated in the present investigation. Both silybin A and silybin B potently inhibited SN-38 glucuronidation catalyzed by UGT1A1*1 or UGT1A1*6. It was noteworthy that silybin A and silybin B showed synergistic effect in UGT1A1*1 microsomes at concentration around IC50, while additive effect in UGT1A1*6. According to the predicted AUCi/AUC ratios (the ratio of the area under the plasma concentration-time curve of SN-38 in the presence and absence of silybins), the coadministration of irinotecan and several milk thistle products, including silybin-phosphatidylcholine complex, two Legalon capsules, four Silymarin tablets or four Liverman capsules, may lead to clinically significant herb-drug interactions (HDI) via UGT1A1 inhibition. Meanwhile, Rgut values were much higher than 11 in all the groups, indicating potential HDI due to intestinal UGT1A1 inhibition.

Tripartite motif protein 11 (TRIM11), an oncogene for human lung cancer via the DUSP6-mediated ERK1/2 signaling pathway.

Evidence suggests that Tripartite Motif Containing 11 (TRIM11) has pro-tumor activity in human non-small cell lung cancer (NSCLC). However, the roles and underlying mechanisms of TRIM11 in NSCLC have not yet been fully elucidated. In this work, human lung cancer cell lines (A549, H446, and H1975) were transfected with siRNA or lentiviruses to knockdown or overexpress TRIM11 and dual-specificity phosphatase 6 (DUSP6). The cell tumor response was assessed by determining the rate of proliferation, apoptosis, the uptake of 2-[N-(7-nitrobenz-2-oxa-1, 3-diaxol-4-yl) amino]-2-deoxyglucose (2-NBDG), and the secretion of lactic acid (LD). Dominant-negative (dn)-MEK1 was used to block the ERK1/2 pathway. The mechanism was investigated by assessing the protein levels of pyruvate kinase isozymes M2 (PKM2) and DUSP6, as well as the activation of ERK1/2 pathway. Our data confirmed the anti-cancer effect of siTRIM11 in human lung cancer by demonstrating inhibition of cancer cell proliferation, induction of apoptosis, prevention of 2-NBDG uptake, suppression of LD production, and prevention of lung cancer cell (A549) tumorigenicity in nude mice. The underlying mechanism involved the up-regulation of DUSP6 and the inhibition of ERK1/2 activity. Overexpression of TRIM11 induced tumorigenesis of NSCLC in vitro, and the activation of ERK1/2 was significantly reversed by DUSP6 overexpression or additional dn-MEK1 treatment. Interestingly, we confirmed TRIM11 as a deubiquitinase that regulated DUSP6 accumulation, indicating that lung cancer progression is regulated via the DUSP6-ERK1/2 pathway. In conclusion, TRIM11 is an oncogene in NSCLC, likely through the DUSP6-mediated ERK1/2 signaling pathway.

Surface heparin treatment of the decellularized porcine heart valve: Effect on tissue calcification.

Tissue calcification is a major cause of failure of bioprosthetic heart valves. Aim of this study was to examine whether surface heparin treatment of the decellularized porcine heart valve reduces tissue calcification. Fresh porcine aortic heart valves were dissected as tissue discs and divided into four groups. Group A: controls without treatment, Group B: decellularization only, Group C: decellularization and glutaraldehyde cross-linking, Group D: decellularization and glutaraldehyde cross-linking followed by surface heparin treatment. After implantation in New Zealand White rabbits for 60 days, the explanted heart valve discs from the different study groups underwent a series of histological examinations as well as determination of calcium content by the methyl thyme phenol blue colorimetric method. Results of the explanted heart valve discs for the Von Kossa staining demonstrated that in Group A the heart valve tissue was the most severely stained with black color, whereas in Group D there was hardly any area that was stained black after implantation indicating the least tissue calcification. Furthermore, the inflammatory cells identified by the Hematoxylin-eosin staining appeared to be the least in Group D. The average tissue calcium content was highest in Group A (0.197 ± 0.115 µmol mg-1 ), modest in Group B (0.113 ± 0.041 µmol mg-1 ), and Group C (0.089 ± 0.049 µmol mg-1 ), and the lowest in Group D (0.019 ± 0.019 µmol mg-1 , p < 0.05). These results suggest that surface heparin treatment tends to reduce tissue calcification of the dellellularized porcine heart valve in a rabbit intramuscular implantation model. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 400-405, 2017.

Knockdown of TRIM65 inhibits lung cancer cell proliferation, migration and invasion: A therapeutic target in human lung cancer.

Lung cancer is the most commonly diagnosed type of cancer worldwide. Although TRIM65 is an important protein involved in white matter lesion, the role of TRIM65 in human cancer remains less understood. Here, we reported that TRIM65 was significantly overexpressed in lung cancer tissues compared with adjacent normal lung tissues. Furthermore, TRIM65 expression was closely related to overall survival of patients with lung cancer. Knock down of TRIM65 in two lung cancer cell lines, SPC-A-1 and NCI-H358, resulted in a significant reduction in cell proliferation, migration, invasion and adhesion and a dramatic increase in G0-G1 phase arrest and apoptosis. In vivo tumorigenesis experiment also revealed that depletion of TRIM65 expression inhibited NCI-H358 cell growth. Moreover, based on gene set enrichment analysis (GSEA) with The Cancer Genome Atlas (TCGA) dataset, we found that TRIM65 was positive related to cell cycle, metastasis up and RHOA-REG pathways, which was further validated by RT-PCR and Western blot in TRIM65 knockdown lung cancer cells and indicated a possible mechanism underlying its effects on lung cancer. In summary, our study suggests that TRIM65 may work as an oncogene and a new effective therapeutic target for lung cancer treatment.

Diagnosis and surgical treatment of esophageal gastrointestinal stromal tumors.

AIM: To retrospectively evaluate our experience with the diagnosis and surgical resection of esophageal gastrointestinal stromal tumors (GISTs). METHODS: Between January 2003 and August 2014, five esophageal GIST cases were admitted to our hospital. In this study, the hospital records, surgery outcomes, tumor recurrence and survival of these patients were retrospectively reviewed. RESULTS: The median age of the patients was 45.6 years (range: 12-62 years). Three patients presented with dysphagia, and one patient presented with chest discomfort. The remaining patient was asymptomatic. Four patients were diagnosed with esophageal GISTs by a preoperative endoscopic biopsy. Three patients underwent esophagectomy, and two patients underwent video-assisted thoracoscopic surgery. The mean operating time was 116 min (range: 95-148 min), and the mean blood loss was 176 mL (range: 30-300 mL). All tumors were completely resected. The mean length of postoperative hospital stay was 8.4 d (range: 6-12 d). All patients recovered and were discharged successfully. The median postoperative follow-up duration was 48 mo (range: 29-72 mo). One patient was diagnosed with recurrence, one patient was lost to follow-up, and three patients were asymptomatic and are currently being managed with close radiologic and clinical follow-up. CONCLUSION: Surgery is the standard, effective and successful treatment for esophageal GISTs. Long-term follow-up is required to monitor recurrence and metastasis.

Structural integrity, immunogenicity and biomechanical evaluation of rabbit decelluarized tracheal matrix.

Decellularization techniques have been widely used as an alternative strategy to produce matrices for organ reconstruction. This study investigated the impact of a detergent-enzymatic decellularization protocol on the extracellular matrix integrity, mechanical properties, and biocompatibility of decellularized tracheal matrices from rabbits. The tracheas of New Zealand white rabbits were decellularized using a modified detergent-enzymatic method (DEM). Antigenicity, cellularity, glycosaminoglycan content, DNA content, histoarchitecture, and mechanical properties were monitored during processing. The surface ultrastructure of the matrix was examined by scanning electron microscopy (SEM). Bioengineered and control tracheas were then implanted in major histocompatibility complex-unmatched rats (xenograft) heterotopically for 7, 15, and 30 days. Structural and functional analysis was performed after transplantation. The results showed that seven cycles of decellularization removed most of the cells and eliminated antigenicity. Histological and molecular biology analysis demonstrated that most of the cellular components and nuclear material were removed. SEM analysis revealed that the decellularized matrices retained the hierarchical structure of the native trachea, and biomechanical tests showed that decellularization did not significantly influence the mechanical properties. Seven, 15 and 30 days after implantation, decreased (p < 0.01) inflammatory reactions were observed in the xenograft models for decellularized matrices compared with control tracheas. No increases in IgM or IgG content were observed in rats that received bioengineered tracheas. In conclusion, this work suggests that seven cycles of the DEM generates a bioengineered rabbit tracheal matrix that is structurally and mechanically similar to native trachea.

In vitro isolation and cultivation of rabbit tracheal epithelial cells using tissue explant technique.

Epithelial cells from tracheal mucosa offer significant potential as a cell source in development of tissue-engineered trachea. The purpose of this study was to investigate and optimize a suitable culture system for tracheal epithelial cells, including the methods of primary culture, passage, identification, and cryopreservation. Epithelial cells were isolated from rabbit tracheal mucosa using tissue explant technique and were subjected to immunohistochemistry, immunofluorescence, and cryopreservation after purification. Epithelial cells reached confluency at 14-15 d. Immunohistochemical staining for cytokeratin showed brown yellow-positive cytoplasm and blue-counterstained nuclei, while immunofluorescence staining for cytokeratin showed green-positive cytoplasm and clear cell outline, indicating that the cultured cells had properties of epithelial cells. After recovery, epithelial cells exhibited high survival and viability. The results demonstrated that in vitro isolation and cultivation model was successfully established to provide high proliferative capacity, typical morphology and characteristics of tracheal epithelial cells from trachea mucosa by the use of the tissue explant technique.

Biomechanical properties of adult-excised porcine trachea for tracheal xenotransplantation.

BACKGROUND: To investigate and evaluate the biomechanical properties of adult-excised porcine trachea, thereby providing experimental methods and evidence for biomedical engineering of artificial trachea. METHODS: The TY8000 servo-handle tension test machine was used to measure biomechanical indices, such as bending stiffness, radial pedestal, and stress-straining. The residual stress and bursting strength of adult-excised porcine trachea was evaluated. RESULTS: Residual stress was retained in the adult-excised porcine trachea. The force of radial pedestal was detected as 10 N, when the diameter of a 50-mm trachea was compressed to 50%. The bursting strength decreased from 180 mmHg of pharyngeal portion to 110 mmHg in tracheal carina. When the trachea flexed forward or either right or left by 50 degrees , tension reached 0.296 to 0.131 N and 0.254 to 0.150 N, respectively. The curve of stress-straining measured, according to computer data and results, suggested that tension was maintained at a low level at 50% strain. CONCLUSIONS: Residual stress was retained in the excised porcine trachea, and the porcine trachea membrane disrupted when pressure in the inner wall increased. The porcine trachea exhibits good radial pedestal force, bending, and elongation properties.

An experimental research on cryopreserving rabbit trachea by vitrification.

Vitrification is a promising alternative to tissue preservation, in which the tissue is permeated with cryoprotective agents (CPAs) in order to circumvent the hazardous effects associated with ice formation. In this study, we evaluate the effect of vitreous cryopreservation of rabbit trachea, by comparing vitrification procedure with conventional computer-programmed slow freezing approaches. Harvested rabbit trachea were tailored and divided into groups and cryopreserved by vitrification and programmed freezing, respectively. The morphology and ultrastructure of the thawed tracheal fragments including HE dyes, terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end-labeling (TUNEL) staining, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) were studied to assess the integrity of the tracheal fragments. Morphological studies demonstrated that both cryopreservation procedure retained the integrity of trachea, both epithelial cells, cilia and cartilage cells were in good shape. Compared with slow freezing methods, vitrification was less detrimental to cartilage cells and had a higher survival rate of chondrocytes and coverage of epithelium and cilia. Therefore, vitrification procedure can be a more satisfactory method to preserve trachea and the survival of chondrocytes in situ in cartilage tissue is adequate and respiratory epithelium is soundly present.

[Experimental study on the reconstruction of circumferential tracheal defects with novel prosthesis].

OBJECTIVE: To investigate the feasibility of using new tracheal prosthesis made of biomaterials to replace extensive circumferential tracheal defects in mongrel dogs. METHODS: Three types of tracheal prostheses were developed, whose basic skeleton of tubular mesh was knitted with polypropylene monofilament and poly (lactic-co-glycolic acid) fiber. The inner side of type-I tubular mesh was first coated with polyurethane solution and then with collagen. The exterior of type-I was then immobilized with collagen-hydroxyapatite composites. In contrast, the internal and external walls of type-II were coated with polyurethane solution, which produced a prosthesis similar to a nonporous one, while type-III was coated only with collagen solution. Surgical resection and replacement of a segment of the cervical trachea was performed in 16 adult mongrel dogs. The efficacy of the implanted prosthesis periodically evaluated postoperatively. RESULTS: In group A, only one died from prosthetic dehiscence, another from anastomotic leakage, and the others had uneventful postoperative courses. The implanted prosthesis was completely incorporated with the recipient trachea, where different length of reepithelialization occurred on the luminal surface of the reconstructed trachea. Macroscopic examination showed scattered and different sizes of neo-ossification surrounding the implanted prosthesis. The prosthesis was roentgenopaque when exposed to routine X rays. In contrast, a relatively high number of complications occurred postoperatively in group B and C. CONCLUSION: Type-I tracheal prosthesis may be used effectively for long-segment circumferential tracheal replacement, and appears very promising for clinical application, with further improvements in promoting the epithelialization.

[Experimental study on a novel esophageal prosthesis made of composite biomaterials].

OBJECTIVE: To design and develop a novel esophageal prosthesis by selecting appropriate biomaterials, developing special manufacturing techniques, and investigating the feasibility of replacement of cervical esophagus in mongrel dogs. METHODS: In accordance with the requirements of ideal esophageal substitutes, we designed a new type of esophageal prostheses. The inner stent were made with polyurethane of medical grade, and the outer surface of the prosthesis was coated with collagen-chitosan sponge. The silicone tube was used as a control. Thirteen adult mongrel dogs that were divided into two groups were used to establish the experimental models. RESULTS: In the experimental group (n = 8), the esophageal prostheses were completely incorporated with the native esophagus and adherent to the surrounding host connective tissues. Epithelial linings of varying degrees were formed on the luminal surface, and complete epithelization was seen in 1 month postoperatively. The granulation at the sites of the anastomosis in this group was less significant than that of the control group. One dog has been surviving for 12 months up to now without any complications. In the control group (n = 5), esophageal epithelial was not observed on the luminal surface, constriction of the regenerated esophagus progressed and all the dogs died within 2 months after operation. CONCLUSION: These observations suggest that this esophageal prosthesis made of composite biomaterials has high biocompatibility and potential for long-segment esophageal reconstruction, which is promising for the clinical repair of esophageal defects.