One-Step Fast Fabrication of Electrospun Fiber Membranes for Efficient Particulate Matter Removal.

Rapid social and industrial development has resulted in an increasing demand for fossil fuel energy, which increases particulate matter (PM) pollution. In this study, we employed a simple one-step electrospinning technique to fabricate polysulfone (PSF) fiber membranes for PM filtration. A 0.3 g/mL polymer solution with an N,N-dimethylformamide:tetrahydrofuran volume ratio of 3:1 yielded uniform and bead-free PSF fibers with a diameter of approximately 1.17 µm. The PSF fiber membrane exhibited excellent hydrophobicity and mechanical properties, including a tensile strength of 1.14 MPa and an elongation at break of 116.6%. Finally, the PM filtration performance of the PSF fiber membrane was evaluated. The filtration efficiencies of the membrane for PM2.5 and PM1.0 were approximately 99.6% and 99.2%, respectively. The pressure drops were 65.0 and 65.2 Pa, which were significantly lower than those of commercial air filters. Using this technique, PSF fiber membrane filters can be easily fabricated over a large area, which is promising for numerous air filtration systems.

Oil mistparticulate matter exposure induces hyperlipidemia-related inflammation via microbiota/ SCFAs/GPR43 axis inhibition and TLR4/NF-κB activation.

Metabolites produced by the human gut microbiota play an important role in fighting and intervening in inflammatory diseases. It remains unknown whether immune homeostasis is influenced by increasing concentrations of air pollutants such as oil mist particulate matters (OMPM). Herein, we report that OMPM exposure induces a hyperlipidemia-related phenotype through microbiota dysregulation-mediated downregulation of the anti-inflammatory short-chain fatty acid (SCFA)-GPR43 axis and activation of the inflammatory pathway. A rat model showed that exposure to OMPM promoted visceral and serum lipid accumulation and inflammatory cytokine upregulation. Furthermore, our research indicated a reduction in both the "healthy" microbiome and the production of SCFAs in the intestinal contents following exposure to OMPM. The SCFA receptor GPR43 was downregulated in both the ileum and white adipose tissues (WATs). The OMPM treatment mechanism was as follows: the gut barrier was compromised, leading to increased levels of lipopolysaccharide (LPS). This increase activated the Toll-like receptor 4 Nuclear Factor-κB (TLR4-NF-κB) signaling pathway in WATs, consequently fueling hyperlipidemia-related inflammation through a positive-feedback circuit. Our findings thus imply that OMPM pollution leads to hyperlipemia-related inflammation through impairing the microbiota-SCFAs-GPR43 pathway and activating the LSP-induced TLR4-NF-κB cascade; our findings also suggest that OMPM pollution is a potential threat to humanmicrobiota dysregulation and the occurrence of inflammatory diseases.

Identification of potential glioma drug resistance target proteins based on ultra-performance liquid chromatography-mass spectrometry differential proteomics.

In this study, to screen for candidate markers of temozolomide (TMZ) resistance in glioblastoma, we artificially established TMZ drug-resistant glioblastoma (GBM) cell lines, U251-TMZ and U87-TMZ. In the U251-TMZ and U87-TMZ cell lines, we screened and analyzed differentially expressed proteins using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) differential proteomics. Compared with the U251 and U87 control cell lines, 95 differential proteins were screened in the U251-TMZ and U87-TMZ cell lines, of which 28 proteins were upregulated and 67 proteins were down-regulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of the co-upregulated proteins showed that most of the differentially expressed proteins were located in the cytoplasm and were significantly upregulated in the biological processes related to vesicular transport in the intimal system and inflammatory response mediated by myeloid leukocytes. Seven candidates were identified as potential GBM markers of TMZ resistance. Combined with existing research findings, our study supports that UAP1L1 and BCKDK are promising potential markers of TMZ resistance in GBM. This is important for further understanding the molecular mechanisms that drive the development and enhancement of TMZ resistance.

PM2.5 triggers autophagic degradation of Caveolin-1 via endoplasmic reticulum stress (ERS) to enhance the TGF-ß1/Smad3 axis promoting pulmonary fibrosis.

Air pollution is highly associated with respiratory diseases. However, the influence and mechanism of particulate matter with aerodynamic equal to or less than 2.5 µm (PM2.5) in lung homeostasis remain unclear. Herein, we demonstrated the induction of pulmonary fibrosis (PF) by PM2.5 exposure. The animal model showed that PM2.5 exposure could activate the oxidative stress and inflammation response, promoting epithelial-mesenchymal transition and accumulation of collagen, high expression of pro-fibrotic factors, and pathological characteristics of fibrosis. The proteomic analysis indicated that PM2.5 exposure decreased the expression of caveolin-1 (Cav-1), and many differential proteins were enriched in the TGF-ß1/Smad, endoplasmic reticulum stress (ERS) and autophagy pathways. Combining in vivo and in vitro experiments, it was found that PM2.5 exposure could reduce Cav-1 protein levels and activate TGF-ß1/Smad3 signaling pathways through ERS and autophagy pathways, thereby inducing cell apoptosis and promoting pulmonary fibrosis. However, inhibiting ERS could alleviate the occurrence of autophagy, and blocking the autophagy system could increase the level of Cav-1 protein and inhibit TGF- ß 1/Smad3 signaling pathway to improve pulmonary fibrosis. Therefore, we demonstrated that the exposure of PM2.5 could enhance the ERS induced-autophagy-mediated Cav-1 degradation, thus activating the TGF-ß1/Smad3 axis to promote pneumonocytes apoptosis and overproduction of extracellular matrix (ECM), finally aggravating PF. Moreover, our findings revealed that intermittent exposure to high doses of PM2.5 was more toxic than continuous exposure to low dose.

Correction: Jia et al. Exposure to Polypropylene Microplastics via Oral Ingestion Induces Colonic Apoptosis and Intestinal Barrier Damage through Oxidative Stress and Inflammation in Mice. Toxics 2023, 11, 127.

In the original publication [...].

Distribution of Micro-Nano PS, DEHP, and/or MEHP in Mice and Nerve Cell Models In Vitro after Exposure to Micro-Nano PS and DEHP.

Polystyrene (PS) and di-(2-ethylhexyl) phthalate (DEHP) exist widely in the environment. However, their distribution in organisms remains unclear. We used three sizes (50 nm, 500 nm, and 5 µm) of PS and DEHP to study the distribution and accumulation of PS, DEHP, and mono(2-ethylhexyl) phthalate (MEHP) in mice and nerve cell models (HT22 and BV2 cells) and their potential toxicity. Results showed that PS entered the blood of mice, and the distribution of different particle sizes in different tissues was different. After the combined exposure to PS and DEHP, PS carried DEHP, which significantly increased the DEHP content and MEHP content and the highest content of MEHP was in the brain. With the decrease in PS particle size, the contents of PS, DEHP, and MEHP in the body increased. The levels of inflammatory factors were increased in the serum of the PS or/and DEHP group. In addition, 50 nm polystyrene can carry MEHP into nerve cells. These results suggest for the first time that PS and DEHP combined exposure can induce systemic inflammation, and the brain is an important target organ of PS and DEHP combined exposure. This study may serve as a reference for further evaluation of the neurotoxicity induced by combined exposure to PS and DEHP.

Exposure to Polypropylene Microplastics via Oral Ingestion Induces Colonic Apoptosis and Intestinal Barrier Damage through Oxidative Stress and Inflammation in Mice.

Extensive environmental pollution by microplastics has increased the risk of human exposure to plastics. However, the biosafety of polypropylene microplastics (PP-MPs), especially of PP particles < 10 µm, in mammals has not been studied. Thus, here, we explored the mechanism of action and effect of exposure to small and large PP-MPs, via oral ingestion, on the mouse intestinal tract. Male C57BL/6 mice were administered PP suspensions (8 and 70 µm; 0.1, 1.0, and 10 mg/mL) for 28 days. PP-MP treatment resulted in inflammatory pathological damage, ultrastructural changes in intestinal epithelial cells, imbalance of the redox system, and inflammatory reactions in the colon. Additionally, we observed damage to the tight junctions of the colon and decreased intestinal mucus secretion and ion transporter expression. Further, the apoptotic rate of colonic cells significantly increased after PP-MP treatment. The expression of pro-inflammatory and pro-apoptosis proteins significantly increased in colon tissue, while the expression of anti-inflammatory and anti-apoptosis proteins significantly decreased. In summary, this study demonstrates that PP-MPs induce colonic apoptosis and intestinal barrier damage through oxidative stress and activation of the TLR4/NF-κB inflammatory signal pathway in mice, which provides new insights into the toxicity of MPs in mammals.

The impact of subchronic ozone exposure on serum metabolome and the mechanisms of abnormal bile acid and arachidonic acid metabolisms in the liver.

Ambient ozone (O3) pollution can induce respiratory and cardiovascular toxicity. However, its impact on the metabolome and the underlying mechanisms remain unclear. This study first investigated the serum metabolite changes in rats exposed to 0.5 ppm O3 for 3 months using untargeted metabolomic approach. Results showed chronic ozone exposure significantly altered the serum levels of 34 metabolites with potential increased risk of digestive, respiratory and cardiovascular disease. Moreover, bile acid synthesis and secretion, and arachidonic acid (AA) metabolism became the most prominent affected metabolic pathways after O3 exposure. Further studies on the mechanisms found that the elevated serum toxic bile acid was not due to the increased biosynthesis in the liver, but the reduced reuptake from the portal vein to hepatocytes owing to repressed Ntcp and Oatp1a1, and the decreased bile acid efflux in hepatocytes as a results of inhibited Bsep, Ostalpha and Ostbeta. Meanwhile, decreased expressions of detoxification enzyme of SULT2A1 and the important regulators of FXR, PXR and HNF4α also contributed to the abnormal bile acids. In addition, O3 promoted the conversion of AA into thromboxane A2 (TXA2) and 20-hydroxyarachidonic acid (20-HETE) in the liver by up-regulation of Fads2, Cyp4a and Tbxas1 which resulting in decreased AA and linoleic acid (LA), and increased thromboxane B2 (TXB2) and 20-HETE in the serum. Furthermore, apparent hepatic chronic inflammation, fibrosis and abnormal function were found in ozone-exposed rats. These results indicated chronic ozone exposure could alter serum metabolites by interfering their metabolism in the liver, and inducing liver injury to aggravate metabolic disorders.

Intestinal toxicity of micro- and nano-particles of foodborne titanium dioxide in juvenile mice: Disorders of gut microbiota-host co-metabolites and intestinal barrier damage.

The wide use of TiO2 particles in food and the high exposure risk to children have prompted research into the health risks of TiO2. We used the microbiome and targeted metabolomics to explore the potential mechanism of intestinal toxicity of foodborne TiO2 micro-/nanoparticles after oral exposure for 28 days in juvenile mice. Results showed that the gut microbiota-including the abundance of Bacteroides, Bifidobacterium, Lactobacillus, and Prevotella-changed dynamically during exposure. The organic inflammatory response was activated, and lipopolysaccharide levels increased. Intestinal toxicity manifested as increased mucosal permeability, impaired intestinal barrier, immune damage, and pathological changes. The expression of antimicrobial peptides, occludin, and ZO-1 significantly reduced, while that of JNK2 and Src/pSrc increased. Compared with micro-TiO2 particles, the nano-TiO2 particles had strong toxicity. Fecal microbiota transplant confirmed the key role of gut microbiota in intestinal toxicity. The levels of gut microbiota-host co-metabolites, including pyroglutamic acid, L-glutamic acid, phenylacetic acid, and 3-hydroxyphenylacetic acid, changed significantly. Significant changes were observed in the glutathione and propanoate metabolic pathways. There was a significant correlation between the changes in gut microbiota, metabolites, and intestinal cytokine levels. These, together with the intestinal barrier damage signaling pathway, constitute the network mechanism of the intestinal toxicity of TiO2 particles.

Melatonin-ROS signal module regulates plant lateral root development.

Lateral root (LR) branches from primary root. LR is vital for plants acquiring water and nutrients from soil, especially under stress conditions. LR development involves the complicated signaling network, which has not yet been fully understood. Melatonin, a novel endogenous plant regulator, plays a role in the regulation of LR development. However, we still have limited knowledge about melatonin-modulated signaling during LR development. Our recent study identifies that reactive oxygen species (ROS) acts as downstream signaling of melatonin to facilitate LR development. The recently identified receptor of melatonin in plants controls a signaling module involving G protein, ROS, and Ca2+. Based on these findings, we propose a novel signaling network for LR development controlled by melatonin.

Ozone exposure promotes pyroptosis in rat lungs via the TLR2/4-NF-κB-NLRP3 signaling pathway.

OBJECTIVE: Ozone has become a major air pollutant in recent years, which leading to a variety of lung diseases. This study aimed to explore the mechanisms of pyroptosis and related signaling pathways in ozone-induced lung injury. METHODS: We exposed 120 Wistar rats to ozone, 20 in each group (half male and half female). Ozone exposure concentrations were 0, 0.12, 0.5, 1.0, 2.0 and 4.0 ppm for 6 h. At the same time, we isolated and cultured type I alveolar epithelial cells, then intervened with high mobility group box 1 protein (HMGB1), hyaluronic acid (HA) and Toll-like receptors 2/4 (TLR2/4) inhibitor. In animal experiments, histopathological experiments, TUNEL, ELISA and biochemical indicators were performed. RT-qPCR and western blot experiments assay were used to detect the expression changes of key factors in relevant signal pathways in vivo and in vitro. RESULTS: After acute ozone exposure, the levels of lung cell injury indicators in bronchoalveolar lavage fluid (BALF), as well as the levels of inflammatory factors in BALF, blood, and lung tissue were significantly increased. Male rats were more sensitive to ozone exposure. Low-concentration ozone exposure caused mild interstitial inflammation in rat lung tissue. Severe inflammation and pulmonary edema appeared with increases in concentration. ELISA results in BALF showed that HMGB1 and HA expressions increased gradually with the increase of ozone exposure concentration. RT-qPCR and Western blot showed that when ozone concentrations increased above 0.5 ppm, the expression of nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing 3 (NLRP3), cleaved caspase-1, and N-gasdermin D (N-GSDMD) in the lung tissue increased significantly, suggesting that ozone exposure induces pyroptosis. At the same time, it was found that ozone exposure activated the nuclear factor kappa B (NF-κB) signal pathway, and increased the mRNA expressions of Toll-like receptors TLR2/4. The results of cell experiments showed that after the addition of HMGB1 and HA, the expression of NF-κB and pyroptosis related indexes increased in type I alveolar epithelial cells, while the corresponding expression decreased after the addition of TLR2/4 inhibitors. CONCLUSION: Ozone exposure causes lung injury in a dose- and gender-dependent manner, and is more severe in males. When injured, the levels of HMGB1 and HA in BALF increased, which interact with TLR 2/4 to activate the downstream NF-κB signaling pathway. Further activating the NLRP3 inflammasome complex and regulating the ozone-induced pyroptosis.

Acute ozone exposure can cause cardiotoxicity: Mitochondria play an important role in mediating myocardial apoptosis.

OBJECTIVE: To clarify the cardiotoxicity induced by acute exposure to different concentrations of ozone in both gender rats and explore the underlying mechanisms. METHODS: A total of 240 rats were randomly sorted into 6 groups with equal numbers of male and female rats in each group. The rats were subjected to ozone inhalation at concentrations of 0, 0.12, 0.5, 1.0, 2.0 and 4.0 ppm, respectively, for 6 h. After ozone exposure, function indicators, myocardial injury indexes and risk factors of cardiovascular disease in blood were assayed. RESULTS: High ozone exposure resulted in sustained ventricular tachycardia in male and female rats. Myocardial apoptosis in male rats started from 1.0 ppm ozone, and that in female rats started from 2.0 ppm ozone (p < 0.05). Caspase-9 increased significantly from 0.12 ppm ozone (p < 0.01) in both gender rats, while caspase-3 was initially activated at 0.5 ppm ozone. From 1.0 ppm ozone, mitochondrial cristae and myofilaments dissolved. The ratio of Bcl-2/Bax decreased significantly from 0.12 ppm and MRCC-IV decreased significantly from 2.0 ppm by ozone. CONCLUSION: Acute ozone exposure can cause paroxysmal ventricular tachycardia in rats. Moreover, the changes of inflammatory factors in the heart tissues of female and male rats after ozone exposure were greater than those of oxidative stress. This study reported for the first time that 6 h ozone exposure does not cause acute cardiomyocyte necrosis, but promotes cardiomyocyte apoptosis in a mitochondrial-dependent manner. Ozone could regulate caspases-3 dependent cardiomyocyte apoptosis by affecting the balance between caspase-9 and XIAP.

Exposure to copper oxide nanoparticles triggers oxidative stress and endoplasmic reticulum (ER)-stress induced toxicology and apoptosis in male rat liver and BRL-3A cell.

Copper oxide nanoparticles (Nano-CuO) toxicity has been researched widely in recent years. However, the relationship between oxidative stress and ER-stress and the possible mechanisms induced by Nano-CuO have been rarely studied. Here, the mechanism of hepatotoxicity and apoptosis through oxidative stress and ER-stress induced by Nano-CuO was investigated in vivo and in vitro. In in vivo experiments, male Wistar rats were intranasally instilled 10 µg Nano-CuO/g body weight daily for 60 days, which caused liver function impairment, oxidative stress, inflammatory response, histopathological and ultrastructural damage, ER-stress and apoptosis in liver tissue. in vitro experiments on rat hepatocytes BRL-3A cells showed that exposure to Nano-CuO for 24 h resulted in excess production of reactive oxygen species leading to decrease in mitochondria membrane potential causing cell death by inducing apoptosis. However, administration of n-acetyl cysteine decreased the apoptosis in Nano-cuo treated group. The in vivo and in vitro experiments confirmed that oxidative stress triggered ER-stress pathway, leading to the opening of apoptosis pathways of CHOP, JNK, and Caspase-12. In summary, treatment of Nano Cuo triggered oxidative stress by ROS, which in turn resulted in activation of ER stress pathways causing cell death in liver tissue and BRL-3A cells.

Neurotoxicity of aluminum oxide nanoparticles and their mechanistic role in dopaminergic neuron injury involving p53-related pathways.

The central nervous system is a potential target for Al2O3 nanoparticles (Nano-Al2O3). Here, we investigated the effects of intranasal instillation of Nano-Al2O3 on the distribution and damage in crucial functional sub-brain regions of rats. In vivo results show that Nano-Al2O3 was translocated into the brain via the olfactory nerve pathway. Nano-Al2O3 accumulated in the hippocampus, olfactory bulb, cerebral cortex, and striatum, causing ultrastructural changes, oxidative damage, inflammatory responses, and histopathological damage in sub-brain regions. As indicated by in vitro studies, cell viability decreased with the addition of Nano-Al2O3, which increased the levels of lactate dehydrogenase and oxidative stress. Nano-Al2O3 also impaired mitochondrial function, disturbed the cell cycle and induced apoptosis. In addition, Nano-Al2O3 decreased the expression of cyclin D1, bcl-2, Mdm2, and phospho-Rb and increased the expression of p53, p21, Bax, and Rb. Therefore, oxidative stress, mitochondrial dysfunction, and p53-related pathways might be important in the process of dopaminergic neurotoxicity induced by Nano-Al2O3. The current study establishes a striatum damage model and identifies molecular biomarkers of dopaminergic neuron damage induced by Nano-Al2O3. In brief, our study demonstrates that Nano-Al2O3 exposure can be a risk factor for neurodegenerative diseases and may negatively impact the hippocampus, striatum, and dopaminergic neurons.

Identification of apoptosis-associated protein factors distinctly expressed in cigarette smoke condensate-exposed airway bronchial epithelial cells.

Smoking is associated with an increased risk of respiratory diseases, including lung cancer and asthma. However, the mechanisms or diagnostic markers for smoking-related diseases remain largely unknown. Here we investigated the role of cigarette smoke condensate (CSC) in the regulation of human bronchial epithelial cell (BEAS-2B) behavior. We found that exposure to CSC significantly inhibited BEAS-2B cell viability, impaired cell morphology, induced cell apoptosis, triggered oxidative damage, and promoted inflammatory response, which suggests a deleterious effect of CSC on bronchial epithelial cells. In addition, CSC markedly altered the expression of apoptosis-associated protein factors, including p21, soluble tumor necrosis factor receptor 1, and Fas ligand. In sum, our study identified a panel of novel protein factors that may mediate the actions of CSC on bronchial epithelial cells and have a predictive value for the development and progression of smoking-related diseases, thus providing insights into the development of potential diagnostic and therapeutic strategies against these diseases.

Neurotoxicity and biomarkers of zinc oxide nanoparticles in main functional brain regions and dopaminergic neurons.

Manufactured zinc oxide nanoparticles (Nano-ZnO) are being used increasingly in many fields owing to their excellent physicochemical properties. Consequently, biosecurity has become a growing concern for human health and the environment. In the present study, Nano-ZnO neurotoxicity was investigated in vivo and in vitro. In vivo results showed that Nano-ZnO particles delivered through intranasal instillation were translocated to the brain, specifically deposited in the olfactory bulb, hippocampus, striatum, and cerebral cortex, and caused ultrastructural changes, oxidative damage, inflammatory responses, and histopathological damages there, which may be important for inducing Nano-ZnO neurotoxicity. Further in vitro studies on PC12 cell line illustrated that exposure to Nano-ZnO for 6 h affected cell morphology, decreased cell viability, increased lactate dehydrogenase and oxidative stress activity levels, impaired mitochondrial function, and disturbed the cell cycle. In addition, Nano-ZnO could destroy neuronal structure by affecting cytoskeleton proteins (tubulin-α, tubulin-ß and NF-H), resulting in the interruption of connection between nerve cells, which lead to nervous system function damage. Meanwhile, Nano-ZnO could induce neuronal repair and regeneration disorders by affecting the growth-related protein GAP-43 and delayed neurotoxicity by affecting the calcium/calcium-regulated kinase (CAMK2A/CAMK2B protein) signaling pathway.

UCH-L1 mitigates neurotoxicity induced by ZnO particles via stabilizing the inhibitor of NF-kappa B signaling, IκB-α.

Our study determined the toxic effects of zinc oxide (ZnO) particles with different diameters on dopaminergic (DA) neurons, the role of ubiquitin C-terminal hydrolase L1 (UCH-L1) for ZnO particles-induced neurotoxicity, and corresponding molecular mechanisms. We constructed an in vitro cell injury model for DA neurons to analyze the cytotoxicity of ZnO particles using SH-SY5Y cells. Following cell viability assays and flow cytometry, we found that the cytotoxicity of ZnO particles was affected by particle size, time, and dose of exposure. For example, the toxicity of ZnO particles with 50 nm or 100 nm diameter was stronger than that of ZnO particles with 1000 nm diameter. Furthermore, ZnO particles exposure resulted in a significant decrease in UCH-L1 expression in SH-SY5Y; whereas UCH-L1 overexpression led to a significant increase in cell viability and a sharp decrease in ROS level. Western blotting and adenovirus transfection found that exposure to ZnO particles with different diameters all activate the NF-κB signaling in SH-SY5Y cells; whereas UCH-L1 over-expression resulted in increased levels of IκBα, an endogenous inhibitor of NF-κB signaling pathway. ZnO particles with different diameters all induced cytotoxicity in DA neurons, which may be related to the free Zn2+ in the suspension. Regarding the neurotoxic effect of ZnO particles, UCH-L1 protects against and/or alleviates neuronal damage, possibly by deubiquitination of the endogenous inhibitor, IκBα, which leads to activation of NF-κB signaling. Therefore, one possible mechanism for ZnO particle-induced neurotoxicity may be mediated via the down-regulation of UCH-L1 expression in DA cells.

Strengthening Effect of Extruded Mg-8Sn-2Zn-2Al Alloy: Influence of Micro and Nano-Size Mg2Sn Precipitates.

In this study, Mg-8Sn-2Zn-2Al (TZA822) alloys with varying Mg2Sn contents prior to extrusion were obtained by different pre-treatments (without and with T4), and the strengthening response related to micro and nano-size Mg2Sn precipitates in the extruded TZA822 alloys was reported. The results showed that the morphology of nano-size Mg2Sn precipitates exhibits a significant change in basal plane from rod-like to spherical, owing to the decrement in the fraction of micro-size particles before extrusion. Meanwhile, the spherical Mg2Sn precipitates provided a much stronger strengthening effect than did the rod-like ones, which was ascribed to uniform dispersion and refinement of spherical precipitates to effectively hinder basal dislocation slip. As a consequence, the extruded TZA822 alloy with T4 showed a higher tensile yield strength (TYS) of 245 MPa, ultimate tensile strength (UTS) of 320 MPa and elongation (EL) of 26.5%, as well as a lower degree of yield asymmetry than their counterpart without T4. Detailed reasons for the strengthening effect were given and analyzed.