氯化锂通过TGFBI促进角膜基质成纤维细胞增殖和自噬

目的：研究TGFBI和微管相关蛋白轻链3(LC3)在角膜营养不良患者中的表达,及氯化锂(LiCl)通过TGFBI对角膜基质成纤维细胞增殖能力的影响。

方法：用免疫组化和Western-blot方法检测角膜营养不良及正常角膜组织中TGFBI和LC3的表达。实验构建了TGFBI过表达载体并转染角膜基质成纤维细胞,分别以5、10、20、40mmol/L LiCl作用于突变型TGFBI转染的角膜基质成纤维细胞,检测不同时间(0、1、6、12h)后,TGFBI与LC3蛋白表达变化,并用CCK-8法检测细胞增殖活性。

结果：TGFBI和LC3在角膜营养不良患者角膜组织中显著高表达。TGFBI过表达抑制角膜基质成纤维细胞增殖活性(P<0.05)。LiCl抑制突变型TGFBI转染的角膜基质成纤维细胞中TGFBI和LC3蛋白表达,并增强其细胞增殖活性(P<0.05)。

结论：LiCl可以促进角膜基质成纤维细胞增殖活性和自噬,其作用机制与下调TGFBI和LC3的表达有关。     

Constructing Large Scale Cohort for Clinical Study on Heart Failure with Electronic Health Record in Regional Healthcare Platform: Challenges and Strategies in Data Reuse

Regional healthcare platforms collect clinical data from hospitals in specific areas for the purpose of healthcare management. It is a common requirement to reuse the data for clinical research. However, we have to face challenges like the inconsistence of terminology in electronic health records (EHR) and the complexities in data quality and data formats in regional healthcare platform. In this paper, we propose methodology and process on constructing large scale cohorts which forms the basis of causality and comparative effectiveness relationship in epidemiology. We firstly constructed a Chinese terminology knowledge graph to deal with the diversity of vocabularies on regional platform. Secondly, we built special disease case repositories (i.e., heart failure repository) that utilize the graph to search the related patients and to normalize the data. Based on the requirements of the clinical research which aimed to explore the effectiveness of taking statin on 180-days readmission in patients with heart failure, we built a large-scale retrospective cohort with 29647 cases of heart failure patients from the heart failure repository. After the propensity score matching, the study group (n=6346) and the control group (n=6346) with parallel clinical characteristics were acquired. Logistic regression analysis showed that taking statins had a negative correlation with 180-days readmission in heart failure patients. This paper presents the workflow and application example of big data mining based on regional EHR data.     

Overexpression of NIMA-related kinase 6 (NEK6) contributes to malignant growth and dismal prognosis in Human Breast Cancer

NIMA Related Kinase 6 (Nek6) is a protein kinase involved in various cellular processes, including cell cycle regulation, apopotosis, senescence, telomere maintainance and chemoresistance. In the present study, we investigated the role of Nek6 in breast tumorigenesis and the prognostic merit of Nek6 expression in breast cancer. Immunohistochemistry and Western blot analyses was conducted to determine the expression profile of Nek6 in 133 breast cancer specimens. Nek6 was overexpressed in a majority of breast cancer specimens, compared with the adjacent non-tumorous tissues. Furthermore, we revealed that high expression of Nek6 was associated with histologic grade, tumor size and TNM stage in breast cancer. Liner regression analysis showed a significant association between the levels of Nek6 and Ki-67. Cox regression analysis indicated that Nek6 expression was an independent prognostic predictor for breast cancer. Moreover, intracellular level of Nek6 was remarkably increased following the release from serum starvation in MCF-7 cells. EdU incorporation assay indicated that depletion of Nek6 remarkably impaired the proliferation of MCF-7 cells. Finally, spheroid formation assay revealed that interference of Nek6 led to diminished oncospheroid-forming capacity of breast cancer cells. In conclusion, our results imply that Nek6 plays a facilitating role in breast cancer cell proliferation and may serve as a promising therapeutic target for breast cancer.     

Osteogenic Differentiation of Marrow Stromal Cells on Random and Aligned Electrospun Poly(l-lactide) Nanofibers

The fibrillar structure and sub-micron diameter of electrospun nanofibers can be used to reproduce the morphology and structure of the natural extracellular matrix (ECM). The objective of this work was to investigate the effect of fiber alignment on osteogenic differentiation of bone marrow stromal (BMS) cells. Random and aligned poly(l-lactide) (PLLA) nanofibers were produced by collecting the spun fibers on a stationary plate and a rotating wheel, respectively, as the ground electrode. Morphology and alignment of the BMS cells seeded on the fibers were characterized by SEM. The effect of fiber orientation on osteogenic differentiation of BMS cells was determined by measuring alkaline phosphatase (ALPase) activity, calcium content, and mRNA expression levels of osteogenic markers. There was a strong correlation between the fiber and cell distributions for the random (p = 0.16) and aligned (p = 0.81) fibers. Percent deviation from ideal randomness (PDIR) values indicated that cells seeded on the random fibers (PDIR = 6.5%) were likely to be distributed randomly in all directions while cells seeded on the aligned fibers (PDIR = 86%) were highly likely to be aligned with the direction of fibers. BMS cell seeded on random and aligned fibers had similar cell count and ALPase activity with incubation time, but the calcium content on aligned fibers was significantly higher after 21 days compared to that of random fibers (p = 0.003). Osteopontin (OP) and osteocalcin (OC) expression levels of BMS cells on fibers increased with incubation time. However, there was no difference between the expression levels of OP and OC on aligned vs. random fibers. The results indicate that BMS cells aligned in the direction of PLLA fibers to form long cell extensions, and fiber orientation affected the extent of mineralization, but it had no effect on cell proliferation or mRNA expression of osteogenic markers.     

Continuous Renal Replacement Therapy (CRRT) Attenuates Myocardial Inflammation and Mitochondrial Injury Induced by Venovenous Extracorporeal Membrane Oxygenation (VV ECMO) in a Healthy Piglet Model

In this study, we investigated the myocardial inflammation and mitochondrial function during venovenous extracorporeal membrane oxygenation (VV ECMO) and further evaluated the effects of continuous renal replacement therapy (CRRT) on them. Eighteen piglets were assigned to the control group, ECMO group, and ECMO+CRRT group. Myocardial inflammation was assessed by the activity of myeloperoxidase (MPO), myocardial concentrations, and mRNA expression of TNF-α, IL-1β, and IL-6; mitochondrial function was assessed by activities of mitochondrial complexes I–V. VV ECMO elicited a general activation of serum and myocardial inflammation and significantly decreased the activities of mitochondrial complexes I and IV. After being combined with CRRT, serum and myocardial concentrations of IL-1β and IL-6, myocardial mRNA expression of IL-6, and the activity of MPO were decreased significantly; the activities of mitochondrial complexes were increased. We conclude that myocardial inflammation was activated during ECMO therapy, inducing mitochondrial injury; moreover, CRRT reduced myocardial inflammation and partially ameliorated mitochondrial function.     

MEASUREMENT OF AMPLITUDE AND DELAY OF STIMULUS FREQUENCY OTOACOUSTIC EMISSIONS

Although stimulus frequency otoacoustic emissions (SFOAEs) have been used as a non-invasive measure of cochlear mechanics, clinical and experimental application of SFOAEs has been limited by difficulties in accurately deriving quantitative information from sound pressure measured in the ear canal. In this study, a novel signal processing method for multicomponent analysis (MCA) was used to measure the amplitude and delay of the SFOAE. This report shows the delay-frequency distribution of the SFOAE measured from the human ear. A low level acoustical suppressor near the probe tone significantly suppressed the SFOAE, strongly indicating that the SFOAE was generated at characteristic frequency locations. Information derived from this method may reveal more details of cochlear mechanics in the human ear.     

Quantitative analysis of hyperpolarized 129Xe ventilation imaging in healthy volunteers and subjects with chronic obstructive pulmonary disease

In this study, hyperpolarized ¹²⁹Xe MR ventilation and ¹H anatomical images were obtained from three subject groups: young healthy volunteers (HVs), subjects with chronic obstructive pulmonary disease (COPD) and age‐matched controls (AMCs). Ventilation images were quantified by two methods: an expert reader‐based ventilation defect score percentage (VDS%) and a semi‐automated segmentation‐based ventilation defect percentage (VDP). Reader‐based values were assigned by two experienced radiologists and resolved by consensus. In the semi‐automated analysis, ¹H anatomical images and ¹²⁹Xe ventilation images were both segmented following registration to obtain the thoracic cavity volume and ventilated volume, respectively, which were then expressed as a ratio to obtain the VDP. Ventilation images were also characterized by generating signal intensity histograms from voxels within the thoracic cavity volume, and heterogeneity was analyzed using the coefficient of variation (CV). The reader‐based VDS% correlated strongly with the semi‐automatically generated VDP (r = 0.97, p < 0.0001) and with CV (r = 0.82, p < 0.0001). Both ¹²⁹Xe ventilation defect scoring metrics readily separated the three groups from one another and correlated significantly with the forced expiratory volume in 1 s (FEV₁) (VDS%: r = –0.78, p = 0.0002; VDP: r = –0.79, p = 0.0003; CV: r = –0.66, p = 0.0059) and other pulmonary function tests. In the healthy subject groups (HVs and AMCs), the prevalence of ventilation defects also increased with age (VDS%: r = 0.61, p = 0.0002; VDP: r = 0.63, p = 0.0002). Moreover, ventilation histograms and their associated CVs distinguished between subjects with COPD with similar ventilation defect scores, but visibly different ventilation patterns. Copyright © 2012 John Wiley & Sons, Ltd.