Sulodexide protects against hyperglycemia-induced retinal vascular injury via suppressing NOX4/NLRP3 pathway

AIM To investigate the effect of sulodexide (SDX) on high glucose-induced damage in retinal microvascular endothelial cells. METHODS (1) High-fat diet combined with intraperitoneal injection of streptozocin were used to induce type 2 diabetes mellitus (DM) followed by injection of saline or SDX in C57BL/6J male mice. Retinal microvascular leakage and density, and the protein levels of NLRP3 inflammasome-related proteins, zonula occludens-1 (ZO-1) and NADPH oxidase 4 (NOX4) were measured. (2) Human retinal microvascular endothelial cells (HRMECs) were treated with normal glucose or high glucose with or without SDX, and were further transfected with siRNA to knock down NOX4, or infected by adenovirus to over-express NOX4. The protein levels of ZO-1, VE-cadherin (VE-Cad), NOX4 and NLRP3 inflammasome-related proteins as well as the level of reactive oxygen species (ROS) were detected. RESULTS Treatment with SDX increased the protein level of ZO-1, attenuated retinal leakage and NLRP3 inflammasome activation, and enhanced the density of microvasculature and the number of ganglion cells in diabetic retinas. The protein levels of ZO-1 and VE-Cad were decreased, while the levels of NOX4, NLRP3 inflammasome-related proteins and ROS generation were increased in high glucose-treated HRMECs. Silencing of NOX4 inhibited high glucose-induced increases in NLRP3 inflammasome and ROS generation, and decreases in the protein levels of ZO-1 and VE-Cad. Over-expression of NOX4 significantly increased the levels of NLRP3 inflammasome-related proteins and ROS generation in HRMECs, and reduced the protein levels of ZO-1 and VE-Cad. Treatment with SDX partly reversed NOX4 over-expression-induced changes. CONCLUSION SDX alleviates hyperglycemia-induced retinal microvascular endothelial injury via inhibiting NOX4/ROS/NLRP3 pathways.     

Mouse melanoma cell exosomes promote expression of Rac1 protein in fibroblasts

AIM To investigate the effect of exosomes secreted by mouse melanoma cells on the expression of Ras-related C3 botulinum toxin substrate 1 (Rac1) protein in fibroblasts. METHODS Ultracentrifugation was adopted to separete exosomes secreted by mouse melanoma B16-F10 cells. The morphological structure of exosomes was observed by negative-staining electron microscopy. The size distribution of exosomes was determined by nanoparticle tracking analysis (NTA). The exosomal markers, tumor susceptibility gene 101 (Tsg101) and tyrosinase-related protein 2 (Tyrp2), were identified by Western blot. Laser confocal microscopy was used to observe the process that mouse embryonic fibroblasts (MEF) took in exosomes during co-culture. Immunocytochemical staining and Western blot were used to detect the expression of Rac1 protein in MEF. RESULTS B16-F10 cell exosomes showed a typical tea tray-like structure, with a size range of 141~255 nm, and expressed protein markers Tsg101 and Tyrp2. The results of laser confocal microscopy showed that compared with co-culture at 0 h, a small number of exosomes appeared in the MEF at 12 h, and a large number of exosomes accumulated in the MEF after co-cultured for 24 and 36 h. Western blot analysis showed that compared with co-culture at 0 h, the expression of Rac1 protein in the MEF was significantly increased at 24 h and 36 h of co-culture (P<0.01). The results of immunocytochemical staining showed that compared with co-culture at 0 h, the positive expression level of Rac1 in the MEF cells was significantly increased at 12 h, 24 h and 36 h of co-culture (P<0.05 or P<0.01). CONCLUSION Intake of exosomes secreted by mouse melanoma cells promotes the expression of Rac1 protein in fibroblasts.     

ROCK promotes high glucose-induced cardiomyocyte apoptosis by inhi-biting PI3K/Akt signaling pathway

AIMTo investigate whether Rho-associated coiled-coil kinase (ROCK) is involved in high glucose-induced apoptosis of primary cardiomyocytes by regulating PI3K/Akt signaling pathway. METHODSPrimary Wistar rat cardiomyocytes were cultured and identified by α-sarcomeric actin (α-SCA) immunohistochemistry. Cardiomyocytes were treated with 5.5, 33 and 40 mmol/L glucose for 48 h. The cell viability was measured by MTT assay, and the mRNA expression of ROCK1 and ROCK2 in the cardiomyocytes was detected by RT-qPCR. Flow cytometry was used to analyze the apoptosis of the cardiomyocytes. The protein levels of ROCK1, ROCK2, cleaved caspase-3, Bcl-2, PI3K, Akt and p-Akt were determined by Western blot. In order to confirm the regulatory effect of ROCKs on PI3K/Akt signaling pathway, the cells were divided into control group (5.5 mmol/L glucose), high glucose group (33 mmol/L glucose) and high glucose+Y27632 (ROCK inhibitor) group. Western blot was used to detect the protein levels of ROCK1, ROCK2, PI3K, Akt and p-Akt. RESULTSAfter 48 h of high glucose exposure, the values of relative cell viability in 33 and 40 mmol/L glucose groups were (79.71±2.43)% and (68.41±7.49)%, respectively, both of which were significantly decreased compared with normal control group (P<0.05). After 48 h of high glucose exposure, the relative mRNA levels of ROCK1 and ROCK2 in 33 and 40 mmol/L glucose groups were significantly increased compared with normal control group (P<0.05). Compared with normal control group, the apoptotic rate in 33 and 40 mmol/L glucose groups was increased significantly (P<0.05). Compared with normal control group, the protein expression of ROCK1, ROCK2 and cleaved caspase-3 in 33 and 40 mmol/L glucose groups was increased (P<0.05), while the protein expression of Bcl-2 was decreased (P<0.05). No significant difference in the protein levels of PI3K and Akt among the 3 groups was observed, while the protein level of p-Akt in 33 and 40 mmol/L glucose groups was decreased compared with normal control group (P<0.05). Compared with high glucose group, the expression of ROCK1 and ROCK2 was decreased in high glucose+Y27632 group. No significant difference in the protein levels of PI3K and Akt among the 3 groups was observed. Compared with normal control group, the protein level of p-Akt in high glucose group was decreased, and the protein level of p-Akt in high glucose+Y27632 group was increased significantly compared with high glucose group. CONCLUSION Under high glucose environment, ROCK may reduce the level of p-Akt by inhibiting the PI3K/Akt signaling pathway， thus promoting the apoptosis of cardiomyocytes.     

赤霉素不同处理浓度和浸种时间对托鲁巴姆发芽的影响

托鲁巴姆种子的休眠较深,发芽较困难。为了打破其种子的休眠期,加快种子萌发,提高种子发芽的整齐度,以托鲁巴姆为试材,研究了不同的赤霉素处理浓度和浸种时间对其发芽的影响。结果表明:生产中适宜的赤霉素处理浓度为750~1 250 mg/L,浸种时间为36 h,发芽率可达93.00%;可有效提高托鲁巴姆种子的发芽速率和发芽整齐度。     

Development of a linear mixed-effects individual-tree basal area increment model for masson pine in Hunan Province,South-central China

ABSTRACT

An individual-tree basal area increment model was developed for masson pine based on 26276 observations of 13,138 trees in 987 sample plots from the 7th (2004), 8th (2009), and 9th (2014) Chinese National Forest Inventory in Hunan Province, South-central China. The model was built using a linear mixed-effects approach with sample plots included as random effects since the data have a hierarchical stochastic structure and biased estimates of the standard error of parameter estimates could be a consequence of applying ordinary least square (OLS) for regression. In addition, within-plot heteroscedasticity and autocorrelation were also considered. The final mixed-effects model was determined according to the Akaike information criterion (AIC), Bayesian information criterion (BIC), log-likelihood (Loglik), and the likelihoodratio test (LRT). The results revealed that initial diameter (DBH), the sum of the basal area (m²/ha) in trees with DBHs larger than the DBH of the subject tree (BAL), number of trees per hectare (NT), and elevation (EL) had a significant impact on individual-tree basal area increment. The mixed-effects model performed much better than the basic model produced using OLS. Additionally, the variance structure of the model errors was successfully modeled using the power function. However, the autocorrelation structures were not defined because there was no autocorrelation amongst the data. It is believed that the final model will contribute to the scientific management of the masson pine.     

Site‐specific management is crucial to managing Mikania micrantha

Increasingly, weeds have been taking on global distributions. With the proliferation of invasive weeds has come the challenge of managing these species over broad geographical regions, with diverse habitats and political jurisdictions. Here, we review the management of Mikania micrantha Kunth (Asteraceae; mile‐a‐minute) throughout its invaded range, extending through most of the Pacific islands and southern and south‐east Asia. Context matters when determining the best course of action for managing M. micrantha, as it has invaded a large variety of agricultural and natural systems. In Queensland, Australia and Florida, USA, M. micrantha has been targeted in relatively successful eradication campaigns, highlighting the importance of early detection and rapid response methods, while elsewhere in its invaded range, populations are either still increasing or showing limited signs of decline. An inter‐regional approach to research and management should incorporate successful management strategies employed throughout the invaded range including, but not limited to, chemical and cultural control practices, manual and mechanical control, classical biological control using the rust fungus Puccinia spegazzinii, plant–plant competition and integrated approaches utilising two or more control methods concurrently. Additional knowledge of M. micrantha genetics is required to determine if management approaches could be fine‐tuned for particular populations. Countries bordering the Mekong River formed a network in 2011 to co‐ordinate the management of invasive species such as M. micrantha. Expanding such a collaborative approach to other regions could further reduce populations of M. micrantha and limit its spread.     

牛星状病毒EvaGreen实时荧光定量PCR检测方法的建立及应用

牛星状病毒(BAstV)是我国新发的犊牛腹泻病原,本试验的目的是建立检测BAstV的Real-time PCR方法。根据BAstV流行株的ORF1a基因序列设计引物,通过优化反应条件和体系,成功建立基于EvaGreen检测BAstV的Real-time PCR方法。结果表明,该检测方法的Ct值与标准品模板在1.36×10¹~1.36×10⁸拷贝/μL线性关系良好,相关系数R²=0.999,扩增效率为93.79%;该方法可特异性检出BAstV,对犊牛腹泻其他相关病原呈阴性;最低检测下限为13.6拷贝/μL;批间和批内的变异系数均小于2%,重复性好。对2017年9月至2019年5月采自河南省的221份犊牛腹泻样本进行检测,BAstV的检出率为18.1%(40/221),采样场阳性率为100.0%(14/14)。本试验所建方法灵敏度高、特异性强、稳定性好,为BAstV的检测和流行病学调查提供了有力手段。     

Nutritional Qualities of the Low-Value Bivalve Mollusks Paphia malabarica and Villorita cyprinoides at the Estuarine Waters of the Southwestern Coast of India

The bivalve mollusks Paphia malabarica and Villorita cyprinoides collected from the estuarine system located in the southwest coastal waters of India were evaluated for their nutritional composition. A balanced essential to nonessential amino acid ratio (> 1.0) with a greater quantity of sulfur-containing amino acids in these species demonstrated that they could provide well-balanced protein depositions. The n-3/n-6 polyunsaturated fatty acid ratio in P. malabarica was found to be greater than 2, and, therefore, can be considered as a healthy diet. The C₂₀–C₂₂ long-chain polyunsaturated n-3 fatty acids (for example, docosahexaenoic acid and eicosapentaenoic acid) accounted for more than 23% of the total fatty acid content in P. malabarica as compared to about 7% in V. cyprinoides. P. malabarica showed greater hypocholesterolemic/hypercholesterolemic ratio (1.7) and lesser atherogenicity (0.7), thrombogenicity (0.3) indices when contrasted with those in V. cyprinoides, thereby indicating the nutritional superiority of the former. A greater content of vitamin D₃ (> 150 IU), along with significant quantities of calcium and phosphorus (> 500 mg/100 g) in the clams, signified their importance in preventing osteoporosis. This study demonstrated the importance of bivalves, in particular, P. malabarica as a valued species for human consumption.     

Cyprinid herpesvirus 3 as a potential biological control agent for carp (Cyprinus carpio) in Australia: susceptibility of non‐target species

Carp (Cyprinus carpio L.) is a pest species in Australian waterways, and cyprinid herpesvirus 3 (CyHV‐3) is being considered as a potential biological control (biocontrol) agent. An important consideration for any such agent is its target specificity. In this study, the susceptibility to CyHV‐3 of a range of non‐target species (NTS) was tested. The NTS were as follows: 13 native Australian, and one introduced, fish species; a lamprey species; a crustacean; two native amphibian species (tadpole and mature stages); two native reptilian species; chickens; and laboratory mice. Animals were exposed to 100–1000 times the approximate minimum amount of CyHV‐3 required to cause disease in carp by intraperitoneal and/or bath challenge, and then examined clinically each day over the course of 28 days post‐challenge. There were no clinical signs, mortalities or histological evidence consistent with a viral infection in a wide taxonomic range of NTS. Furthermore, there was no molecular evidence of infection with CyHV‐3, and, in particular, all RT‐PCRs for viral mRNA were negative. As a consequence, the results encourage further investigation of CyHV‐3 as a potential biocontrol agent that is specific for carp.