Cellular messengers involved in the inhibition of the Arabidopsis primary root growth by bacterial quorum-sensing signal N-decanoyl-L-homoserine lactone.

BACKGROUND: N-acyl-homoserine lactones (AHLs) are used as quorum-sensing signals by Gram-negative bacteria, but they can also affect plant growth and disease resistance. N-decanoyl-L-homoserine lactone (C10-HSL) is an AHL that has been shown to inhibit primary root growth in Arabidopsis, but the mechanisms underlying its effects on root architecture are unclear. Here, we investigated the signaling components involved in C10-HSL-mediated inhibition of primary root growth in Arabidopsis, and their interplay, using pharmacological, physiological, and genetic approaches. RESULTS: Treatment with C10-HSL triggered a transient and immediate increase in the concentrations of cytosolic free Ca2+ and reactive oxygen species (ROS), increased the activity of mitogen-activated protein kinase 6 (MPK6), and induced nitric oxide (NO) production in Arabidopsis roots. Inhibitors of Ca2+ channels significantly alleviated the inhibitory effect of C10-HSL on primary root growth and reduced the amounts of ROS and NO generated in response to C10-HSL. Inhibition or scavenging of ROS and NO neutralized the inhibitory effect of C10-HSL on primary root growth. In terms of primary root growth, the respiratory burst oxidase homolog mutants and a NO synthase mutant were less sensitive to C10-HSL than wild type. Activation of MPKs, especially MPK6, was required for C10-HSL to inhibit primary root growth. The mpk6 mutant showed reduced sensitivity of primary root growth to C10-HSL, suggesting that MPK6 plays a key role in the inhibition of primary root growth by C10-HSL. CONCLUSION: Our results indicate that MPK6 acts downstream of ROS and upstream of NO in the response to C10-HSL. Our data also suggest that Ca2+, ROS, MPK6, and NO are all involved in the response to C10-HSL, and may participate in the cascade leading to C10-HSL-inhibited primary root growth in Arabidopsis.

Jinlida granules ameliorate the high-fat-diet induced liver injury in mice by antagonising hepatocytes pyroptosis.

CONTEXT: Jinlida (JLD) as a traditional Chinese medicine formula has been used to treat type 2 diabetes mellitus (T2DM) and studies have shown its anti-obesity effect. OBJECTIVE: To investigate the therapeutic effects of JLD in a mouse model of non-alcoholic fatty liver (NAFL). MATERIALS AND METHODS: C57BL/6J mice were divided into three groups and fed a low-diet diet (LFD), high-fat diet (HFD), or HFD + JLD (3.8 g/kg) for 16 weeks, respectively. The free fatty acids-induced lipotoxicity in HepG2 cells were used to evaluate the anti-pyroptotic effects of JLD. The pharmacological effects of JLD on NAFL were investigated by pathological examination, intraperitoneal glucose and insulin tolerance tests, western blotting, and quantitative real-time PCR. RESULTS: In vivo studies showed that JLD ameliorated HFD-induced liver injury, significantly decreased body weight and enhanced insulin sensitivity and improved glucose tolerance. Furthermore, JLD suppressed both the mRNA expression of caspase-1 (1.58 vs. 2.90), IL-1ß (0.93 vs. 3.44) and IL-18 (1.34 vs. 1.60) and protein expression of NLRP3 (2.04 vs. 5.71), pro-caspase-1 (2.68 vs. 4.92) and IL-1ß (1.61 vs. 2.60). In vitro, JLD inhibited the formation of lipid droplets induced by 2 mM FFA (IC50 = 2.727 mM), reduced the protein expression of NLRP3 (0.74 vs. 2.27), caspase-1 (0.57 vs. 2.68), p20 (1.67 vs. 3.33), and IL-1ß (1.44 vs. 2.41), and lowered the ratio of p-IKB-α/IKB-α (0.47 vs. 2.19). CONCLUSION: JLD has a protective effect against NAFLD, which may be related to its anti-pyroptosis, suggesting that JLD has the potential as a novel agent in the treatment of NAFLD.

Long non-coding RNA HOXA11-AS induces type I collagen synthesis to stimulate keloid formation via sponging miR-124-3p and activation of Smad5 signaling.

Keloid, characterized by exuberant collagen deposition and invasive growth beyond original wound margins, results from abnormal wound healing. A recent microarray analysis identified homeobox (HOX) A11 antisense (HOXA11-AS) as a keloid-specific long non-coding RNA, although its potential role in keloid formation remains elusive. In this study, hematoxylin-eosin, Masson, and immunohistochemical staining of type I collagen (ColI) revealed abnormal arrangement and hyperplasia of fibers in keloid tissues along with increased ColI level. qRT-PCR and Western blot showed that HOXA11-AS and ColI were significantly upregulated, while miR-124-3p was decreased in both keloid tissues and human keloid fibroblasts (HKFs). Knockdown of HOXA11-AS inhibited cell proliferation (by CCK-8 and immunofluorescence staining of Ki67) and cell migration (by wound healing and transwell assays). Mechanistic experiments verified that HOXA11-AS acted as a sponge of micro-RNA (miR)-124-3p and Smad5 was a target of miR-124-3p. miR-124-3p sufficiently reversed the regulatory effects of HOXA11-AS, and Smad5 was involved in miR-124-3p-mediated biological functions. Furthermore, HOXA11-AS induced ColI synthesis via sponging miR-124-3p-mediated Smad5 signaling, thus promoting keloid formation. Overall, our study implied that HOXA11-AS induces ColI synthesis to promoted keloid formation via sponging miR-124-3p-mediated Smad5 signaling, which might offer a novel target for developing the therapy of keloid formation.

[Effect of Tongxinluo on Apoptosis of Rat Cardiac Microvascular Endothelial Cells].

OBJECTIVE: To observe the protective effects of Tongxinluo (TXL) on apoptosis of rat cardiac microvascular endothelial cells (RCMECs) resulting from homocysteine (Hcy) induced endoplasmic reticulum stress (ERS), and to determine the signaling pathway behind its protection. METHODS: Primary cultured RCMECs were isolated from neonatal rats using tissue explant method. The morphology of RCMECs was observed using inverted microscope, identified and differentiated by CD31 immunofluorescence method. Selected were well growing 2nd-4th generations of RCMECs. The optimal action time was determined by detecting the expression of glucose regulated protein 78 (GRP78) using immunofluorescence method. In the next experiment RCMECs were divided into 5 groups, i.e., the blank control group, the Hcy induced group (Hcy 10 mmol/L, 10 h), the Hcy + TXL group (Hcy 10 mmol/L + TXL 400 µg/mL), the Hcy +LY294002 group (Hcy 10 mmol/L + LY294002 5 µmol/L, LY294002 as the inhibitor of PI3K), the Hcy + LY294002 + TXL group (Hcy 10 mmol/L + LY294002 5 µmol/L + TXL 400 µg/mL). The apoptosis rate of RCMECs was detected by flow cytometry. mRNA and protein expressions of GRP78, C/ EBP homologous protein (CHOP), and cysteinyl aspartate specific proteinase-12 (caspase12) were detected by real-time reverse transcription PCR (RT-PCR) and Western blot respectively. Expression levels of phosphorylation of phosphatidylinositol 3-kinase (P-PI3K), total phosphatidylinositol 3-kinase (T- P13K) , phosphorylation of kinase B (P-Akt) , and total kinase B (T-Akt) were detected by Western blot. RESULTS: Ten hours Hcy action time was determined. Compared with the blank control group, the apoptosis rate was increased (22.77%), mRNA and protein expressions of GRP78, CHOP, and Caspase-12 were increased, protein expressions of P-PI3K and P-Akt,ratios of P-PI3K/T-PI3K and P-Akt/T-Akt were decreased in the Hcy induced group (P < 0.05, P < 0.01). Compared with the Hcy induced group, the apoptosis rate was decreased (10.17%), mRNA and protein expressions of GRP78, CHOP, and Caspase-12 were decreased, and expression levels of P-PI3K, P-Akt, P-PI3K/T-PI3K, and P-Akt/T-Akt were increased in the Hcy + TXL group (P < 0.05, P < 0.01). Compared with the Hcy + TXL group, the apoptosis rate was increased (17.9%), mRNA and protein expressions of GRP78, CHOP, and Caspase-12 were increased, expression levels of P-PI3K and P-Akt, ratios of P-PI3K/T-PI3K and P-Akt/T-Akt were decreased in the Hcy + TXL + LY294002 group (P < 0.05, P < 0.01). CONCLUSION: TXL could inhibit the apoptosis of RCMECs resulting from Hcy-induced ERS and its mechanism might be associated with activating PI3K/Akt signaling pathway.

[Research on Collateral-unblocking Medicines for Cardiovascular Diseases under the Guidance of Vessels Collateral Theory].

OBJECTIVE: "Vessels Collateral Theory", as the inherit and development of "blood vessels" in Huangdi Neijing, is a systematic system by integrating severe vascular diseases. In this article, by taking cardiovascular diseases (CVD) as a cut-in point, roles of "minute collateral-microvascular" lesions in the occurrence and development of CVD were further explored. The interventional effect of collateral-unblocking medicines under the guidance of Vessels Collateral Theory was also in-depth explored, hoping to leading Chinese medical prevention and treatment of CVD.

Glycyrrhetinic acid blocks SARS-CoV-2 infection by activating the cGAS-STING signalling pathway.

BACKGROUND AND PURPOSE: Traditional Chinese medicine (TCM) played an important role in controlling the COVID-19 pandemic, but the scientific basis and its active ingredients are still weakly studied. This study aims to decipher the underlying anti-SARS-CoV-2 mechanisms of glycyrrhetinic acid (GA). EXPERIMENTAL APPROACH: GA's anti-SARS-CoV-2 effect was verified both in vitro and in vivo. Homogeneous time-resolved fluorescence assays, biolayer interferometry technology, and molecular docking were employed to examine interactions of GA with human stimulator of interferon genes (hSTING). Immunofluorescence staining, western blot, and RT-qPCR were used to investigate nuclear translocation of interferon regulatory factor 3 (IRF3) and levels of STING target genes. Pharmacokinetics of GA was studied in mice. KEY RESULTS: GA could directly bind to Ser162 and Tyr240 residues of hSTING, thus up-regulating downstream targets and activation of the STING signalling pathway. Such activation is crucial for limiting the replication of SARS-CoV-2 Omicron in Calu-3 cells and protecting against lung injury induced by SARS-CoV-2 Omicron infection in K18-ACE2 transgenic mice. Immunofluorescence staining and western blot indicated that GA increased levels of phosphorylated STING, phosphorylated TANK-binding kinase-1, and cyclic GMP-AMP synthase (cGAS). Importantly, GA increased nuclear translocation of IRF3. Pharmacokinetic analysis of GA in mice indicated it can be absorbed into circulation and detected in the lung at a stable level. CONCLUSION AND IMPLICATIONS: Activation of the cGAS-STING pathway through the GA-STING-IRF3 axis is essential for the antiviral activity of GA in mice, providing new insights into the potential translation of GA for treating SARS-CoV-2 in patients.

Eldecalcitol protected osteocytes against ferroptosis of D-gal-induced senescent MLO-Y4 cells and ovariectomized mice.

BACKGROUND: Active vitamin D analog eldecalcitol is clinically applied in treatment of postmenopausal osteoporosis. This study aims to determine the role of eldecalcitol in the protection of osteocytes from senescence and the associated ferroptosis. METHODS: The MLO-Y4 osteocytes were exposed to D-gal inducing senescence. The ovariectomized (OVX) mice treated with D-gal using as an aging inducer were intraperitoneally injected with eldecalcitol. The multiplexed confocal imaging, fluorescence in situ hybridization and transmission electron microscopy were applied in assessing osteocytic properties. Immunochemical staining and immunoblotting were carried out to detect abundance and expression of molecules. RESULTS: The ablation of vitamin D receptor led to a reduction in amounts of osteocytes, a loss of dendrites, an increase in mRNA expression of SASP factors and in protein expression of senescent factors, as well as changes in mRNA expression of ferroptosis-related genes (PTGS2 & RGS4). Eldecalcitol reversed senescent phenotypes of MLO-Y4 cells shown by improving cell morphology and density, decreasing ß-gal-positive cell accumulation, and down-regulating protein expression (P16, P21 & P53). Eldecalcitol reduced intracellular ROS and MDA productions, elevated JC-1 aggregates, and up-regulated expression of Nrf2 and GPX4. Eldecalcitol exhibited osteopreserve effects in D-gal-induced aging OVX mice. The confocal imaging displayed its improvement on osteocytic network organization. Eldecalcitol decreased the numbers of senescent osteocytes at tibial diaphysis by SADS assay and attenuated mRNA expression of SASP factors as well as down-regulated protein expression of senescence-related factors and restored levels of ferroptotic biomarkers in osteocytes-enriched bone fraction. It reduced 4-HNE staining area, stimulated Nrf2-positive staining, and promoted nuclear translocation of Nrf2 in osteocytes of mice as well as inhibited and promoted protein expression of 4-HNE and Nrf2, respectively, in osteocytes-enriched bone fraction. CONCLUSIONS: The present study revealed the ameliorative effects of eldecalcitol on senescence and the associated ferroptosis of osteocytes, contributing to its preservation against osteoporosis of D-gal-induced senescent ovariectomized mice.

Tongxinluo Activates PI3K/AKT Signaling Pathway to Inhibit Endothelial Mesenchymal Transition and Attenuate Myocardial Fibrosis after Ischemia-Reperfusion in Mice.

OBJECTIVE: To investigate the potential role of Tongxinluo (TXL) in attenuating myocardial fibrosis after myocardial ischemia-reperfusion injury (MIRI) in mice. METHODS: A MIRI mouse model was established by left anterior descending coronary artery ligation for 45 min. According to a random number table, 66 mice were randomly divided into 6 groups (n=11 per group): the sham group, the model group, the LY-294002 group, the TXL group, the TXL+LY-294002 group and the benazepril (BNPL) group. The day after modeling, TXL and BNPL were administered by gavage. Intraperitoneal injection of LY-294002 was performed twice a week for 4 consecutive weeks. Echocardiography was used to measure cardiac function in mice. Masson staining was used to evaluate the degree of myocardial fibrosis in mice. Qualitative and quantitative analysis of endothelial mesenchymal transition (EndMT) after MIRI was performed by immunohistochemistry, immunofluorescence staining and flow cytometry, respectively. The protein expressions of platelet endothelial cell adhesion molecule-1 (CD31), α-smoth muscle actin (α-SMA), phosphatidylinositol-3-kinase (PI3K) and phospho protein kinase B (p-AKT) were assessed using Western blot. RESULTS: TXL improved cardiac function in MIRI mice, reduced the degree of myocardial fibrosis, increased the expression of CD31 and inhibited the expression of α-SMA, thus inhibited the occurrence of EndMT (P<0.05 or P<0.01). TXL significantly increased the protein expressions of PI3K and p-AKT (P<0.05 or P<0.01). There was no significant difference between TXL and BNPL group (P>0.05). In addition, the use of the PI3K/AKT pathway-specific inhibitor LY-294002 to block this pathway and combination with TXL intervention, eliminated the protective effect of TXL, further supporting the protective effect of TXL. CONCLUSION: TXL activated the PI3K/AKT signaling pathway to inhibit EndMT and attenuated myocardial fibrosis after MIRI in mice.

[Effects of Tomato Planting Years on Soil Physical and Chemical Properties and Microbial Communities].

Long-term continuous cropping of facility soils could influence soil properties; however, the differences in soil properties among different continuous cropping years are still not well understood. The objective of this study was to explore the effects of continuous cropping years of tomato on the physical and chemical properties and biological characteristics of facility soil. Conventional analysis, high-throughput sequencing, and other methods were used to examine the soil physicochemical properties, soil microbial community diversity, and enzyme activities in facility soil after continuous tomato cropping for 1-3 years, 5-7 years, and more than 10 years. As the continuous tomato cropping years increased, soil bulk density and pH decreased; soil maximal water holding capacity increased; and organic matter, total nitrogen, and total phosphorus accumulated. As continuous cropping years increased, the total salt and EC value decreased with continuous cropping for 5-7 years and increased from 5-7 years to more than 10 years continuous cropping and showed a trend of secondary soil salinization. There was a significant increase in alkaline phosphatase for 1-3 years to 5-7 years continuous tomato cropping. There were significant differences in fungal community abundance among different cropping years. The Simpson index and Shannon index of fungi showed a trend of increasing first and then decreasing with the extension of continuous cropping years and reached the maximum value at 5 years of continuous cropping. The Chao1 index decreased continuously following the cropping years. As continuous cropping years increased, Streptomyces became the dominant bacteria, and Aspergillus and Pseudaleuria became the dominant fungi. The key factors affected by continuous cropping years were available potassium and available nitrogen based on the redundancy analysis. The results of this study lay the foundation for future research on the influence of continuous cropping years on the health of facility soil.

Exhaustive swimming differentially inhibits P2X1 receptor- and α1-adrenoceptor-mediated vasoconstriction in isolated rat arteries.

AIM: To investigate the effects of exhaustive swimming exercise on P2X1 receptor- and α1-adrenoceptor-mediated vasoconstriction of different types of arteries in rats. METHODS: Male Wistar rats were divided into 2 groups: the sedentary control group (SCG) and the exhaustive swimming exercise group (ESEG). The rats in the ESEG were subjected to a swim to exhaustion once a day for 2 weeks. Internal carotid, caudal, pulmonary, mesenteric arteries and aorta were dissected out. Isometric vasoconstrictive responses of the arteries to α,ß-methylene ATP (α,ß-MeATP) or noradrenaline (NA) were recorded using a polygraph. RESULTS: The exhaustive swimming exercise did not produce significant change in the EC(50) values of α,ß-MeATP or NA in vasoconstrictive response of most of the arteries studied. The exhaustive swimming exercise inhibited the vasoconstrictive responses to P2X1 receptor activation in the internal carotid artery, whereas it reduced the maximal vasoconstrictive responses to α1-adrenoceptor stimulation in the caudal, pulmonary, mesenteric arteries and aorta. The rank order of the reduction of the maximal vasoconstriction was as follows: mesenteric, pulmonary, caudal, aorta. CONCLUSION: Exhaustive swimming exercise differentially affects the P2X1 receptor- and α1-adrenoceptor-regulated vasoconstriction in internal carotid artery and peripheral arteries. The ability to preserve purinergic vasoconstriction in the peripheral arteries would be useful to help in maintenance of the basal vascular tone during exhaustive swimming exercise.

The role of AMPKα in high-glucose-induced dysfunction of cultured rat mesangial cells.

AIM: Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) is known as a mammalian cell energy sensor, which could regulate cellular energy metabolism via sensing the alterations of energy balance, such as oversupply or lack of glucose and fatty acid. Recent studies have suggested that AMPK could also regulate many other biological processes, including cell cycling, inflammation, protein synthesis, and so on. In this study, AMPK signaling in high-glucose-induced dysfunction of mesangial cells (MCs) was investigated. METHODS: Established rat glomerular MCs were treated under normal glucose (5.6 mM glucose) or high-glucose conditions (30 mM glucose). mRNA levels of AMPK subunits were detected by reverse transcriptase-polymerase chain reaction. Expressions of AMPKα, phosphorylated AMPKα (p-AMPKα), phosphorylated acetyl-CoA carboxylase (p-ACC), and collagen IV were measured by Western blot. RESULTS: Under high-glucose conditions, AMPKα protein expression and mRNA levels were significantly decreased. High-glucose treatment also induced a notable decrease in p-AMPKα and p-ACC expression. AMPKα activation by 5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside (AICAR) effectively ameliorated high-glucose-induced dysfunction of MCs, including cell proliferation, cell-cycle progression, and collagen IV production. CONCLUSION: High glucose impaired AMPKα in its expression and activity; AICAR significantly ameliorated high-glucose-induced proliferation of MCs and collagen IV production, indicating a role of AMPKα in high-glucose-induced dysfunction of MCs.

[Effects of tongxinluo on angiogenesis and the volume of blood perfusion in ischemic stroke rats].

OBJECTIVE: To observe the effects of Tongxinluo (TXL) on angiogenesis and the volume of blood perfusion in ischemic stroke rats. METHODS: The model of middle cerebral artery occlusion (MCAO) was established using craniotomy ligation of the middle cerebral artery on one side. After screening, the male SD rats were randomly divided into the sham-operation group, the model group, the large dose TXL group, the middle dose TXL group, the low dose TXL group, and the Nimodipine group. The expression of microvascular density (MVD, CD31) of the MCAO rats was detected using immunohistochemical assay after 14 days of medication. The microvascular morphology and the volume of blood perfusion in the brain tissue were observed under laser scanning confocal microscope (LSCM). RESULTS: The positive CD31 expression was intense with significant coloring in the large dose TXL group, the middle dose TXL group, and the Nimodipine group, better than that of the model group. The blood perfusion volume in the ischemic brain cortex could be promoted in the large dose TXL group, the middle dose TXL group, and the Nimodipine group (P<0.01, P<0.05). The optimal effects were shown in the large dose TXL group (P<0.01). CONCLUSION: TXL significantly increased the MVD of the ischemic brain tissue, promoted the post-ischemic angiogenesis, and increased the volume of blood perfusion of ischemic brain tissue, playing certain blood flow compensatory roles.

Physiological significance of P2X receptor-mediated vasoconstriction in five different types of arteries in rats.

P2X(1) receptors, the major subtype of P2X receptors in the vascular smooth muscle, are essential for α,ß-methylene adenosine 5'-triphosphate (α,ß-MeATP)-induced vasoconstriction. However, relative physiological significance of P2X(1) receptor-regulated vasoconstriction in the different types of arteries in the rat is not clear as compared with α(1)-adrenoceptor-regulated vasoconstriction. In the present study, we found that vasoconstrictive responses to noncumulative administration of α,ß-MeATP in the rat isolated mesenteric arteries were significantly smaller than those to single concentration administration of α,ß-MeATP. Therefore, we firstly reported the characteristic of α,ß-MeATP-regulated vasoconstrictions in rat tail, internal carotid, pulmonary, mesenteric arteries, and aorta using single concentration administration of α,ß-MeATP. The rank order of maximal vasoconstrictions for α,ß-MeATP (E (max·α,ß-MeATP)) was the same as that of maximal vasoconstrictions for noradrenaline (E (max·NA)) in the internal carotid, pulmonary, mesenteric arteries, and aorta. Moreover, the value of (E (max·α,ß-MeATP)/E (max·KCl))/(E (max·NA)/E (max·KCl)) was 0.4 in each of the four arteries, but it was 0.8 in the tail artery. In conclusion, P2X(1) receptor-mediated vasoconstrictions are equally important in rat internal carotid, pulmonary, mesenteric arteries, and aorta, but much greater in the tail artery, suggesting its special role in physiological function.

[Effects of fatigue and restraint stress on the expression of carnitine palmitoyltransferase-I and 5-hydroxytryptamine receptors in aorta of rats].

OBJECTIVE: To investigate the effect of fatigue and restraint stress on the expressions of CPT (carnitine palmitoyltransferase)-I, PPAR (peroxisome proliferator-activated receptor) δ, 5-HT (hydroxytryptamine) 1D and 5-HT2A receptors in aorta of rats. METHODS: A total of 45 healthy male Wistar rats were randomly divided into control group, excessive fatigue group and restraint stress group (n = 15 each). The general condition, morphological changes of aortic endothelium cell and the blood levels of ET-1 (endothelin) and NO (nitric oxide) were observed. The real-time reverse transcription PCR (polymerase chain reaction) and Western blot were used to detect the gene and protein expressions of CPT-I, PPAR δ, 5-HT1D and 5-HT2A receptors in aorta. RESULTS: Compared with control group, the structural damages of endothelial cell were induced by excessive fatigue and restraint stress. The plasma levels of ET-1 increased [(124 ± 18) ng/L vs (161 ± 18) ng/L, (154 ± 17) ng/L] (P < 0.01, P < 0.05) while the serum levels of NO decreased [(63 ± 16) µmol/L vs (39 ± 8) µmol/L, (41 ± 7) µmol/L] (P < 0.05); the mRNA expressions of CPT-Iand PPARδ decreased in excessive fatigue rats, [(1.23 ± 0.21) vs (0.42 ± 0.05)], [(1.09 ± 0.10) vs (0.25 ± 0.07)] (P < 0.01); the protein expressions of CPT-Iand PPARδ decreased in excessive fatigue rats, [(1.32 ± 0.07) vs (0.83 ± 0.04)], [(1.41 ± 0.05) vs. (0.75 ± 0.06)]; the mRNA and protein expressions of 5-HT1D receptor decreased in excessive fatigue rats and restraint stress rats, [(1.10 ± 0.15) vs (0.46 ± 0.13), (0.45 ± 0.02)], [(1.19 ± 0.05) vs (0.71 ± 0.06), (0.70 ± 0.05)] (P < 0.01); the mRNA and protein expressions of 5-HT2A receptor increased in excessive fatigue rats and restraint stress rats, [(0.99 ± 0.08) vs (6.73 ± 0.46), (7.01 ± 1.56)], [(0.64 ± 0.03) vs (0.79 ± 0.05), (0.82 ± 0.03)] (P < 0.01). CONCLUSION: Excessive fatigue and restraint stress can injure the structure and function of endothelial cell. The changes in energy of abnormal carnitine metabolism and 5-HT receptors may play important roles.

Efficacy and safety of Lianhuaqingwen capsules, a repurposed Chinese herb, in patients with coronavirus disease 2019: A multicenter, prospective, randomized controlled trial.

BACKGROUND: Coronavirus disease 2019 (Covid-19) has resulted in a global outbreak. Few existing targeted medications are available. Lianhuaqingwen (LH) capsule, a repurposed marketed Chinese herb product, has been proven effective for influenza. PURPOSE: To determine the safety and efficacy of LH capsule in patients with Covid-19. METHODS: We did a prospective multicenter open-label randomized controlled trial on LH capsule in confirmed cases with Covid-19. Patients were randomized to receive usual treatment alone or in combination with LH capsules (4 capsules, thrice daily) for 14 days. The primary endpoint was the rate of symptom (fever, fatigue, coughing) recovery. RESULTS: We included 284 patients (142 each in treatment and control group) in the full-analysis set. The recovery rate was significantly higher in treatment group as compared with control group (91.5% vs. 82.4%, p = 0.022). The median time to symptom recovery was markedly shorter in treatment group (median: 7 vs. 10 days, p < 0.001). Time to recovery of fever (2 vs. 3 days), fatigue (3 vs. 6 days) and coughing (7 vs. 10 days) was also significantly shorter in treatment group (all p < 0.001). The rate of improvement in chest computed tomographic manifestations (83.8% vs. 64.1%, p < 0.001) and clinical cure (78.9% vs. 66.2%, p = 0.017) was also higher in treatment group. However, both groups did not differ in the rate of conversion to severe cases or viral assay findings (both p > 0.05). No serious adverse events were reported. CONCLUSION: In light of the safety and effectiveness profiles, LH capsules could be considered to ameliorate clinical symptoms of Covid-19.

[Effects of Tongxinluo on repeated hypoxic tolerance in mice].

OBJECTIVE: To observe the effects of Tongxinluo (TXL) on repeated hypoxic tolerance in mice and explore the underlying mechanisms. METHODS: Mice were randomly divided into groups of repeated hypoxia (control) and TXL according to body weights. The mice in each group were exposed to acute repeated hypoxia for 0 run (H0), 1 run (H1), 3 runs (H3) and 5 runs (H5). The animal's tolerance time of each hypoxic exposure was recorded. Western blot was used to measure the protein levels of hypoxia inducible factor-1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) in cortex tissue. RESULTS: The hypoxic tolerance time in control and TXL groups significantly increased run by run. Both HIF-1alpha and VEGF proteins in two groups increased gradually. Compared with control group, the tolerance time in H1 of TXL group [(18.0 +/- 2.4) minvs (15.6 +/- 2.0) min], H3 [(68.3 +/- 13.2) min vs (41.7 +/- 9.0) min) and H5 (85.9 +/- 7.0) min vs (51.4 +/- 14.4) min] increased (P < 0.05 or P < 0.01); the HIF-1alpha protein expression in H1 of TXL group (0.95 +/- 0.04 vs 0.79 +/- 0.02), H3 (1.01 +/- 0.03 vs 0.85 +/- 0.02), H5 (1.16 +/- 0.02 vs 0.92 +/- 0.03) increased (P < 0.05 or P < 0.01); the VEGF protein expression in H3 of TXL group (1.14 +/- 0.02 vs 0.89 +/- 0.03), H5 (1.34 +/- 0.05 vs 0.99 +/- 0.07) increased (P < 0.05 or P < 0.01). CONCLUSIONS: Under repeated hypoxia, an organism has a strong adaptive ability. The rises of HIF-1alpha and VEGF may be an adaptive mechanism. TXL can increase obviously the adaptive ability of mice to hypoxia.

Long non-coding RNA HOXA11-AS accelerates the progression of keloid formation via miR-124-3p/TGFßR1 axis.

Emerging evidence reveals the importance of long non-coding RNAs (lncRNAs) in the development and progression of keloid formation, whereas the underlying mechanisms are not well understood. In the present study, we investigated the biological effects and molecular mechanisms of lncRNA HOXA11-AS in keloid formation. First, the expression levels of HOXA11-AS, miR-124-3p, and transforming growth factor ß receptor type I (TGFßR1) were measured in both keloid tissues and human keloid fibroblasts (HKFs) using qRT-PCR and western blot analysis, respectively. Next, we adopted both gain- and loss-of-function strategies to explore the significance of HOXA11-AS. TUNEL, flow cytometry, DNA ladder, and tube formation assays were performed to measure cell apoptosis and angiogenesis, respectively. Besides, the potential binding relationship between HOXA11-AS and miR-124-3p, as well as miR-124-3p and TGFßR1 was identified using bioinformatic screening and verified by luciferase reporter assay. Furthermore, we explored the importance of miR-124-3p in HOXA11-AS-induced phenotypes and regulations on TGFß signaling or PI3K/Akt signaling. We found that HOXA11-AS and TGFßR1 were significantly up-regulated, while miR-124-3p was down-regulated both in keloid tissues or fibroblasts than in normal skin tissues or fibroblasts. Functionally, high expression of HOXA11-AS essentially inhibited cell apoptosis and promoted fibroblast-induced angiogenesis. Mechanistically, miR-124-3p was identified as a downstream effector to be involved in HOXA11-AS-mediated phenotypes through directly targeting TGFßR1, thus modulating PI3K/Akt signaling pathway. Taken together, our findings revealed that HOXA11-AS inhibits cell apoptosis and promotes angiogenesis through miR-124-3p/TGFßR1 axis, contributing to the progression of keloid formation, which might provide a novel target for keloid therapy.

N-3-oxo-hexanoyl-homoserine lactone, a bacterial quorum sensing signal, enhances salt tolerance in Arabidopsis and wheat.

BACKGROUND: N-acyl-homoserine lactones (AHLs) are the quorum sensing (QS) signal molecules to coordinate the collective behavior in a population in Gram-negative bacteria. Recent evidences demonstrate their roles in plant growth and defense responses. RESULTS: In present study, we show that the treatment of plant roots with N-3-oxo-hexanoyl-homoserine lactone (3OC6-HSL), one molecule of AHLs family, resulted in enhanced salt tolerance in Arabidopsis and wheat. We found that the growth inhibition phenotype including root length, shoot length and fresh weight were significantly improved by 3OC6-HSL under salt stress condition. The physiological and biochemical analysis revealed that the contents of chlorophyll and proline were increased and the contents of MDA and Na+ and Na+/K+ ratios were decreased after 3OC6-HSL treatment in Arabidopsis and wheat under salt stress condition. Molecular analysis showed that 3OC6-HSL significantly upregulated the expression of salt-responsive genes including ABA-dependent osmotic stress responsive genes COR15a, RD22, ADH and P5CS1, ABA-independent gene ERD1, and ion-homeostasis regulation genes SOS1, SOS2 and SOS3 in Arabidopsis under salt stress condition. CONCLUSIONS: These results indicated that 3OC6-HSL enhanced plant salt tolerance and ABA-dependent and ABA-independent signal pathways and SOS signaling might be involved in the induction of salt resistance by 3OC6-HSL in plants. Our data provide a new insight into the plant-microbe inter-communication.

Effect of serum from overfatigue rats on JNK/c-Jun/HO-1 pathway in human umbilical vein endothelial cells and the intervening effect of Tongxinluo superfine powder.

OBJECTIVE: To cultivate human umbilical vein endothelial cells (HUVECs) in the serum of overfatigue rats with the intervention of Tongxinluo superfine powder (TXLSP). By examining the variation of the activity of JNK/c-Jun/HO-1 pathway, the possible mechanisms of vascular endothelial dysfunction under overfatigue conditions and the intervening effect of TXLSP were explored. METHODS: The HUVECs were randomly divided into the normal control group, the model group, the SP600125 (a specific antagonist of JNK) group, the TXLSP group and the TXLSP + SP600125 group. The content of carboyhemoglobin (COHb) and the leak rate of lactic dehydrogenase (LDH) in different groups were measured. The mRNA and protein expression of JNK, c-Jun, HO-1 and the phosphorylation level of c-Jun (P-c-Jun) were detected using Western blot and PCR methods. RESULTS: Compared with the normal control group, the COHb level in supernatant was increased significantly in the model group, and the expression of HO-1, JNK, c-Jun mRNA and corresponding proteins and P-c-Jun were also increased remarkably. The increases in these parameters were significantly decreased by SP600125. TXLSP showed remarkable up-regulation on the expression of JNK, c-Jun, P-c-Jun and HO-1 mRNA and their protein expression. Compared with the SP600125 group, the expressions of JNK, c-Jun, P-c-Jun and HO-1 mRNA and its protein in the TXLSP+SP600125 group were significantly increased at different time points (P<0.05, P<0.01). CONCLUSIONS: The vascular endothelial dysfunction under overfatigue conditions is related to the activity of the JNK/c-Jun/HO-1 pathway. One of the mechanisms of TXLSP in improving the vascular endothelial function is to adjust the activity of the JNK/c-Jun/HO-1 pathway at gene and protein levels.

Interaction between COX-2 and iNOS aggravates vascular lesion and antagonistic effect of ginsenoside.

AIM OF THE STUDY: Ginseng, the root of Panax ginseng C.A.Meyer (Araliaceae), is one of the most widely used Chinese herbs with hypotensive and cardiotonic actions for thousands of years, but the underlying mechanisms have not been well determined. Ginsenoside, the effective components of ginseng, has anti-inflammatory and anti-oxidative effects. Cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) are key enzymes in inflammation and oxidative stress, respectively, which have close interaction, aggravating their damaging effects. This study investigated whether COX-2 interacted with iNOS in vascular endothelial lesion and the protective effect of ginsenoside. MATERIALS AND METHODS: SD male rats were fed with high l-methionine (3%, w/w) to induce vascular endothelial lesion, and the rats in ginsenoside group were fed ginsenoside solution (0.8 mg kg(-1)d(-1)). The mRNA expression and protein contents of COX-2 and iNOS were detected by RT-PCR and Western blotting, respectively. The interaction between COX-2 and iNOS was analyzed by co-immunoprecipitation and laser confocal microscopy. The content of NT, a specific marker of peroxynitrite, was evaluated by Western blotting. The morphological changes of vascular endothelium were observed. RESULTS: Compared with control group, the transcription and protein levels of both COX-2 and iNOS increased obviously and their interaction enhanced significantly in model group, in accord with the increased NT content and the pathological alterations of aorta. In ginsenoside group, all these alterations were attenuated significantly (P < 0.01 or P < 0.05). CONCLUSIONS: It is proved that there exists interaction between COX-2 and iNOS, aggravating endothelial lesion through peroxynitrite and ginsenoside might antagonize their interaction, playing a protective role.