[Retracted] Exogenous hydrogen sulfide protects against high glucose­induced apoptosis and oxidative stress by inhibiting the STAT3/HIF­1α pathway in H9c2 cardiomyocytes.

[This retracts the article DOI: 10.3892/etm.2019.8036.].

Hydrogen sulfide functions as a micro-modulator bound at the copper active site of Cu/Zn-SOD to regulate the catalytic activity of the enzyme.

The present study examines whether there is a mechanism beyond the current concept of post-translational modifications to regulate the function of a protein. A small gas molecule, hydrogen sulfide (H2S), was found to bind at active-site copper of Cu/Zn-SOD using a series of methods including radiolabeled binding assay, X-ray absorption near-edge structure (XANES), and crystallography. Such an H2S binding enhanced the electrostatic forces to guide the negatively charged substrate superoxide radicals to the catalytic copper ion, changed the geometry and energy of the frontier molecular orbitals of the active site, and subsequently facilitated the transfer of an electron from the superoxide radical to the catalytic copper ion and the breakage of the copper-His61 bridge. The physiological relevance of such an H2S effect was also examined in both in vitro and in vivo models where the cardioprotective effects of H2S were dependent on Cu/Zn-SOD.

S-propargyl-cysteine delays the progression of atherosclerosis and increases eNOS phosphorylation in endothelial cells.

The present study aimed to investigate the protective effect of S-propargyl-cysteine (SPRC) on atherosclerosis progression in mice. A mouse model of vulnerable atherosclerotic plaque was created in ApoE-/- mice by carotid artery tandem stenosis (TS) combined with a Western diet. Macrophotography, lipid profiles, and inflammatory markers were measured to evaluate the antiatherosclerotic effects of SPRC compared to atorvastatin as a control. Histopathological analysis was performed to assess the plaque stability. To explore the protective mechanism of SPRC, human umbilical vein endothelial cells (HUVECs) were cultured in vitro and challenged with oxidized low-density lipoprotein (ox-LDL). Cell viability was determined with a Cell Counting Kit-8 (CCK-8). Endothelial nitric oxide synthase (eNOS) phosphorylation and mRNA expression were detected by Western blot and RT-qPCR respectively. The results showed that the lesion area quantified by en face photographs of the aortic arch and carotid artery was significantly less, plasma total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) were reduced, plaque collagen content was increased and matrix metalloproteinase-9 (MMP-9) was decreased in 80 mg/kg per day SPRC-treated mice compared with model mice. These findings support the role of SPRC in plaque stabilization. In vitro studies revealed that 100 µmol/L SPRC increased the cell viability and the phosphorylation level of eNOS after ox-LDL challenge. These results suggest that SPRC delays the progression of atherosclerosis and enhances plaque stability. The protective effect may be at least partially related to the increased phosphorylation of eNOS in endothelial cells.

Structural analysis of traditional Chinese complex puzzle locks.

The development of barbed-spring locks in ancient China has a history of more than 2000 years. With the development of the design and manufacturing techniques in ancient China, the safety of locks has gotten better and better. Since the seventeenth century, the puzzle lock, with a complicated structure and a high difficulty in opening, was gradually developed and used. The puzzle lock needs specific steps to be opened. Based on the difficulty of the opening process, traditional Chinese puzzle locks can be partition into two sorts, namely, general puzzle locks and complex puzzle locks. As the structure of the puzzle lock will change during the opening process, the puzzle lock belongs to the reconfigurable mechanism. In this paper, a method of topology matrix is provided to analyze the structure of the complex puzzle lock during systematical operation. Firstly, the characteristics and types of general puzzle locks are explained, and then the topology matrix representation is introduced. Finally, four complex puzzle locks are taken as examples, to illustrate the opening process. There are various types of complex puzzle locks, and the mechanism designs are quite ingenious and interesting, which shows the extraordinary technique and ingenuity of the ancient craftsmen.

[Effect of improvement in antibiotic use strategy on the short-term clinical outcome of preterm infants with a gestational age of <35 weeks]. / æŠ—ç”Ÿç´ ä½¿ç”¨ç­–ç•¥æ”¹è¿›å¯¹èƒŽé¾„<35周早产儿近期临床结局的影响.

OBJECTIVES: To study the effect of improvement in antibiotic use strategy on the short-term clinical outcome of preterm infants with a gestational age of <35 weeks. METHODS: The medical data were retrospectively collected from 865 preterm infants with a gestational age of <35 weeks who were admitted to the Neonatal Intensive Care Unit of Xiangya Hospital of Central South University from January 1, 2014 to December 31, 2016. The improved antibiotic use strategy was implemented since January 1, 2015. According to the time of implementation, the infants were divided into three groups: pre-adjustment (January 1, 2014 to December 31, 2014; n=303), post-adjustment Ⅰ (January 1, 2015 to December 31, 2015; n=293), and post-adjustment Ⅱ (January 1, 2016 to December 31, 2016; n=269). The medical data of the three groups were compared. RESULTS: There were no significant differences among the three groups in gestational age, proportion of small-for-gestational-age infants, sex, and method of birth (P>0.05). Compared with the pre-adjustment group, the post-adjustment I and post-adjustment Ⅱ groups had a significant reduction in the rate of use of antibiotics and the duration of antibiotic use in the early postnatal period and during hospitalization (P<0.05), with a significant increase in the proportion of infants with a duration of antibiotic use of ≤3 days or 4-7 days and a significant reduction in the proportion of infants with a duration of antibiotic use of >7 days in the early postnatal period (P<0.05). Compared with the post-adjustment Ⅰ group, the post-adjustment Ⅱ group had a significant reduction in the duration of antibiotic use in the early postnatal period and during hospitalization (P<0.05), with a significant increase in the proportion of infants with a duration of antibiotic use of ≤3 days and a significant reduction in the proportion of infants with a duration of antibiotic use of 4-7 days or >7 days (P<0.05). Compared with the pre-adjustment group, the post-adjustment I and post-adjustment Ⅱ groups had significantly shorter duration of parenteral nutrition and length of hospital stay (P<0.05). There were gradual reductions in the incidence rates of grade ≥Ⅲ intraventricular hemorrhage (IVH) and late-onset sepsis (LOS) after the adjustment of antibiotic use strategy. The multivariate logistic regression analysis showed that the adjustment of antibiotic use strategy had no effect on short-term adverse clinical outcomes, and antibiotic use for >7 days significantly increased the risk of adverse clinical outcomes (P<0.05). CONCLUSIONS: It is feasible to reduce unnecessary antibiotic use by the improvement in antibiotic use strategy in preterm infants with a gestational age of <35 weeks, which can also shorten the duration of parenteral nutrition and the length of hospital stay and reduce the incidence rates of grade ≥Ⅲ IVH and LOS.

Potential Blood DNA Methylation Biomarker Genes for Diagnosis of Liver Fibrosis in Patients With Biopsy-Proven Non-alcoholic Fatty Liver Disease.

Background and objective: This pilot study aimed to identify potential blood DNA methylation (BDM) biomarker genes for the diagnosis of liver fibrosis in non-alcoholic fatty liver disease (NAFLD). Methods: We included a total of 16 NAFLD patients with significant (SLF, liver fibrosis stage ≥ 2) and 16 patients with non-significant liver fibrosis (NSLF, fibrosis stages 0-1). The association between BDM and liver fibrosis was analyzed. Genes were selected based on a stepwise-filtering with CpG islands containing significant differentially methylated probes. Results: The two groups of patients were distinguishable through both t-distributed stochastic neighbor embedding (t-SNE) analysis and unsupervised hierarchical clustering analysis based on their BDM status. BDM levels were significantly higher in the NSLF group than in the SLF group. The methylation levels in the island and shelf regions were also significantly higher in the NSLF group, as well as the methylation levels in the first exon, 3'-untranslated region, body, ExonBnd, non-intergenic region, transcription start site (TSS)1500, and TSS200 regions (all p < 0.05). BDM status was associated with greater histological liver fibrosis, but not with age, sex, or other histological features of NAFLD (p < 0.05). The methylation levels of the hypomethylated CpG island region of CISTR, IFT140, and RGS14 genes were increased in the NSLF group compared to the SLF group (all p < 0.05). Conclusion: BDM may stratify NAFLD patients with significant and non-significant liver fibrosis. The CISTR, IFT140, and RGS14 genes are potential novel candidate BDM biomarkers for liver fibrosis and these pilot data suggest further work on BDM biomarkers is warranted.

YB-1 Recruits Drosha to Promote Splicing of pri-miR-192 to Mediate the Proangiogenic Effects of H2S.

Aims: The genes targeted by miRNAs have been well studied. However, little is known about the feedback mechanisms to control the biosynthesis of miRNAs that are essential for the miRNA feedback networks in the cells. In this present study, we aimed at examining how hydrogen sulfide (H2S) promotes angiogenesis by regulating miR-192 biosynthesis. Results: H2S promoted in vitro angiogenesis and angiogenesis in Matrigel plugs embedded in mice by upregulating miR-192. Knockdown of the H2S-generating enzyme cystathionine γ-lyase (CSE) suppressed in vitro angiogenesis, and this suppression was rescued by exogenous H2S donor NaHS. Plakophilin 4 (PKP4) served as a target gene of miR-192. H2S up-regulated miR-192 via the VEGFR2/Akt pathway to promote the splicing of primary miR-192 (pri-miR-192), and it resulted in an increase in both the precursor- and mature forms of miR-192. H2S translocated YB-1 into the nuclei to recruit Drosha to bind with pri-miR-192 and promoted its splicing. NaHS treatment promoted angiogenesis in the hindlimb ischemia mouse model and the skin-wound-healing model in diabetic mice, with upregulated miR-192 and downregulated PKP4 on NaHS treatment. In human atherosclerotic plaques, miR-192 levels were positively correlated with the plasma H2S concentrations. Innovation and Conclusion: Our data reveal a role of YB-1 in recruiting Drosha to splice pri-miR-192 to mediate the proangiogenic effect of H2S. CSE/H2S/YB-1/Drosha/miR-192 is a potential therapeutic target pathway for treating diseases, including organ ischemia and diabetic complications. Antioxid. Redox Signal. 36, 760-783. The Clinical Trial Registration number is 2016-224.

Histone deacetylase 6 promotes skin wound healing by regulating fibroblast migration and differentiation in aged mice.

Skin wound healing tends to slow down with aging, which is detrimental to both minor wound recovery in daily life and the recovery after surgery. The aim of current study was to explore the effect of histone deacetylase 6 (HDAC6) on wound healing during aging. Cultured human dermal fibroblasts (HDFs) and mouse full-thickness skin wound model were used to explore the functional changes of replicative senescent dermal fibroblasts and the effect of aging on skin wound healing. Scratch wound healing assay revealed significantly decreased migration speed of senescent HDFs, and BrdU incorporation assay indicated their considerably retardant proliferation. The protein expression levels of collagen and HDAC6 were significantly decreased in both senescent HDFs and skin tissues from aged mice. HDAC6 activity inhibition with highly selective inhibitor tubastatin A (TsA) or HDAC6 knockdown with siRNA decreased the migration speed of HDFs and considerably suppressed fibroblast differentiation induced by transforming growth factor-ß1 (TGF-ß1), which suggests the involvement of HDAC6 in regulating fundamental physiological activities of dermal fibroblasts. In vivo full-thickness skin wound healing was significantly delayed in young HDAC6 knockout mice when compared with young wild type mice. In addition, the wound healing was significantly slower in aged wild type mice than that in young wild type mice, and became even worse in aged HDAC6 knockout aged mice. Compared to the aged wild type mice, aged HDAC6 knockout mice exhibited delayed angiogenesis, reduced collagen synthesis, and decreased collagen deposition in skin wounds. Together, these results suggest that delayed skin wound healing in aged mice is associated with impaired fibroblast function. Adequate expression and activity of HDAC6 are required for fibroblasts migration and differentiation.

D-bifunctional protein deficiency caused by HSD17B4 gene mutation in a neonate. / HSD17B4åŸºå› çªå˜è‡´D-双功能蛋白缺乏症.

A 15-day-old boy was admitted to the hospital due to repeated convulsions for 14 days. The main clinical manifestations were uncontrolled seizures, hypoergia, feeding difficulties, limb hypotonia, and bilateral hearing impairment. Clinical neurophysiology showed reduced brainstem auditory evoked potential on both sides and burst-suppression pattern on electroencephalogram. Measurement of very-long-chain fatty acids in serum showed that C26:0 was significantly increased. Genetic testing showed a pathogenic compound heterozygous mutation, c.101C>T(p.Ala34Val) and c.1448_1460del(p.Ala483Aspfs*37), in the HSD17B4 gene. This article reports a case of D-bifunctional protein deficiency caused by HSD17B4 gene mutation and summarizes the epidemiological and clinical features, diagnosis, and treatment of this disease, with a focus on the differential diagnosis of this disease from Ohtahara syndrome.

S-Propargyl-Cysteine Attenuates Diabetic Cardiomyopathy in db/db Mice Through Activation of Cardiac Insulin Receptor Signaling.

Background: Endogenous hydrogen sulfide (H2S) is emerging as a key signal molecule in the development of diabetic cardiomyopathy. The aim of this study was to explore the effect and underlying mechanism of S-propargyl-cysteine (SPRC), a novel modulator of endogenous H2S, on diabetic cardiomyopathy in db/db diabetic mice. Methods and Results: Vehicle or SPRC were orally administered to 8-month-old male db/db mice and their wild type littermate for 12 weeks. SPRC treatment ameliorated myocardial hypertrophy, fibrosis, and cardiac systolic dysfunction assessed by histopathological examinations and echocardiography. The functional improvement by SPRC was accompanied by a reduction in myocardial lipid accumulation and ameliorated plasma lipid profiles. SPRC treatment improved glucose tolerance in db/db mice, with fasting blood glucose and peripheral insulin resistance remaining unchanged. Furthermore, insulin receptor signaling involving the phosphorylation of protein kinase B (Akt/PKB) and glycogen synthase kinase 3ß (GSK3ß) were elevated and activated by SPRC treatment. Primary neonatal mice cardiomyocytes were cultured to explore the mechanisms of SPRC on diabetic cardiomyopathy in vitro. Consistent with the results in vivo, SPRC not only up-regulated insulin receptor signaling pathway in cardiomyocytes in dose-dependent manner in the basal state, but also relieved the suppression of insulin receptor signaling induced by high concentrations of glucose and insulin. Furthermore, SPRC also enhanced the expression of glucose transporter 4 (GLUT4) and 3H glucose uptake in cardiomyocytes. Conclusions: In this study, we found a novel beneficial effect of SPRC on diabetic cardiomyopathy, which was associated with activation of insulin receptor signaling. SPRC may be a promising medication for diabetic cardiomyopathy in type 2 diabetes mellitus patients.

[Current studies of cytokines in the pathogenesis of atherosclerosis and its therapeutic measures].

Atherosclerosis is a chronic inflammatory disease. Cytokine-related research provides an important direction for the prevention and treatment of atherosclerosis. Cytokines, produced by different types of cells and acting on a range of targets, play a key role in the pathogenesis and progression of atherosclerosis. This review summarizes the main pro-inflammatory and anti-inflammatory cytokines related to atherosclerosis and their underlying mechanism. We also outline current anti-atherosclerosis treatments targeting cytokines. The research and treatment prospects of cytokines in the prevention and treatment of atherosclerosis are discussed briefly as well.

Hydrogen sulfide accumulates LDL receptor precursor via downregulating PCSK9 in HepG2 cells.

Endogenous hydrogen sulfide (H2S) affects cholesterol homeostasis and liver X receptor α (LXRα) expression. However, whether low-density lipoprotein (LDL) receptor (LDLR), a key player in cholesterol homeostasis, is regulated by exogenous H2S through LXRα signaling has not been determined. We investigated the effects of sodium hydrosulfide (NaHS, H2S donor) on LDLR expression in the presence or absence of LXR agonists, T0901317 or GW3965 in HepG2 cells. We found that H2S strongly accumulated LDLR precursor in the presence of T0901317. Hence, LDLR transcription and the genes involved in LDLR precursor maturation and degradation were studied. T0901317 increased the LDLR mRNA level, whereas H2S did not affect LDLR transcription. H2S had no significant effect on the expression of LXRα and inducible degrader of LDLR (IDOL). H2S and T0901317 altered mRNA levels of several enzymes for N- and O-glycosylation and endoplasmic reticulum (ER) chaperones assisting LDLR maturation, but did not affect their protein levels. H2S decreased proprotein convertase subtilisin/kexin type 9 (PCSK9) protein levels and its mRNA level elevated by T0901317. T0901317 with PCSK9 siRNA also accumulated LDLR precursor as did T0901317 with H2S. High glucose increased PCSK9 protein levels and attenuated LDLR precursor accumulation induced by T0901317 with H2S. Taken together, H2S accumulates LDLR precursor by downregulating PCSK9 expression but not through the LXRα-IDOL pathway, LDLR transcriptional activation, or dysfunction of glycosylation enzymes and ER chaperones. These results also indicate that PCSK9 plays an important role in LDLR maturation in addition to its well-known effect on the degradation of LDLR mature form.

[Fetal pleural effusion in the uterus and dyspnea after birth].

Neonatal chylothorax is a common cause of neonatal congenital pleural effusion and is often caused by the accumulation of chylous fluid in the thoracic cavity due to the rupture of the thoracic duct and its branched lymphatic vessels for a variety of reasons. Neonatal chylothorax caused by malignant tumors is extremely rare, and this is the first case of neonatal mediastinal neuroblastoma with chylothorax in China. The boy was found to have pleural effusion in the left thoracic cavity in the uterus, and experienced apnea at birth, as well as dyspnea and cyanosis as the main manifestations after birth. He was diagnosed with left chylothorax based on conventional biochemical analysis of pleural effusion. After the treatment including persistent chest drainage and symptomatic and supportive treatment, the drainage of the left thoracic cavity reached a volume of 90-180 mL per day. Neonatal refractory chylothorax was considered. Chest radiograph on day 13 after birth showed lesions in the upper left lung field, and contrast-enhanced plain CT scan of the chest suggested the possibility of posterior mediastinal neuroblastoma. The autopsy confirmed giant posterior mediastinal neuroblastoma (poorly differentiated), which involved the C7-T6 spinal canal and the nearby erector spinae, with a small amount of tumor tissue in the liver and both adrenal glands. Mediastinal tumor is considered the underlying cause of chylothorax in this case.

[Comparison of the efficacy of domestic and imported caffeine citrate in the treatment of apnea in preterm infants: a prospective randomized double-blind controlled trial].

OBJECTIVE: To compare the efficacy of domestic and imported caffeine citrate in the treatment of apnea in preterm infants. METHODS: A total of 98 preterm infants with a gestational age of 28 - <34 weeks between April 2018 and December 2019 were enrolled. They were randomly administered with domestic (n=48) or imported caffeine citrate (n=50) within 6 hours after birth. The therapeutic effects, complications, adverse effects and clinical outcomes were compared between the two groups. RESULTS: There were no significant differences in the incidence of apnea within 7 days after birth, daily frequency of apnea, the time of apnea disappearance, the failure rate of intubation-surfactant-extubation strategy, the time of non-invasive assisted ventilation, the duration of oxygen therapy, the duration of caffeine citrate therapy, the length of hospital stay, blood gas analysis results, liver and kidney function testing results between the two groups (P>0.05). There were no significant differences in the incidence of complications and the mortality rate between the two groups (P>0.05). There was no significant difference in the incidence of adverse effects between the two groups (P>0.05). CONCLUSIONS: The efficacy and safety of domestic caffeine citrate in the treatment of apnea are similar to those of imported caffeine citrate in preterm infants.

[A multicenter survey of antibiotic use in very and extremely low birth weight infants in Hunan Province].

OBJECTIVE: To investigate the current status of antibiotic use for very and extremely low birth weight (VLBW/ELBW) infants in neonatal intensive care units (NICUs) of Hunan Province. METHODS: The use of antibiotics was investigated in multiple level 3 NICUs of Hunan Province for VLBW and ELBW infants born between January, 2017 and December, 2017. RESULTS: The clinical data of 1 442 VLBW/ELBW infants were collected from 24 NICUs in 2017. The median antibiotic use duration was 17 days (range: 0-86 days), accounting for 53.0% of the total length of hospital stay. The highest duration of antibiotic use was up to 91.4% of the total length of hospital stay, with the lowest at 14.6%. In 16 out of 24 NICUs, the antibiotic use duration was accounted for more than 50.0% of the hospitalization days. There were 113 cases with positive bacterial culture grown in blood or cerebrospinal fluid, making the positive rate of overall bacterial culture as 7.84%. The positive rate of bacterial culture in different NICUs was significantly different from 0% to 14.9%. The common isolated bacterial pathogens Klebsiella pneumoniae was 29 cases (25.7%); Escherichia coli 12 cases (10.6%); Staphylococcus aureus 3 cases (2.7%). The most commonly used antibiotics were third-generation of cephalosporins, accounting for 41.00% of the total antibiotics, followed by penicillins, accounting for 32.10%, and followed by carbapenems, accounting for 13.15%. The proportion of antibiotic use time was negatively correlated with birth weight Z-score and the change in weight Z-score between birth and hospital discharge (rs=-0.095, -0.151 respectively, P<0.01), positively correlated with death/withdrawal of care (rs=0.196, P<0.01). CONCLUSIONS: Antibiotics used for VLBW/ELBW infants in NICUs of Hunan Province are obviously prolonged in many NICUs. The proportion of routine use of third-generation of cephalosporins and carbapenems antibiotics is high among the NICUs.

Comparison of Serum Hepatitis B Virus RNA Levels and Quasispecies Evolution Patterns between Entecavir and Pegylated-Interferon Mono-treatment in Chronic Hepatitis B Patients.

Hepatitis B virus (HBV) RNA may independently predict virological and serological response. This study aimed to compare dynamic changes in serum HBV RNA levels and HBV quasispecies evolution patterns between entecavir and pegylated-interferon mono-treatment in chronic hepatitis B patients and to determine the clinical significance during treatment. TaqMan real-time PCR was used for quantitative analysis. HBV RNA levels were retrospectively determined in serial serum samples from 178 chronic hepatitis B patients who received either entecavir or pegylated-interferon treatment. Both serum HBV DNA and RNA quasispecies were analyzed via next-generation sequencing. Receiver operating characteristics (ROC) analysis was performed to evaluate the prediction value of individual biomarkers for hepatitis B e antigen (HBeAg) seroconversion. Patients who received pegylated-interferon treatment showed stronger declines in HBV RNA levels than did those who received entecavir treatment. Serum HBV RNA levels were lower in patients with subsequent HBeAg seroconversion. At baseline, the level of HBV RNA was better than other indicators in predicting HBeAg seroconversion. Moreover, the predictive value of serum HBV RNA levels was better in the entecavir group. Baseline HBV RNA exhibited a significantly higher genetic diversity than HBV DNA and had a significant decline after 4 weeks of entecavir treatment. Higher baseline genetic diversity may result in a better outcome in pegylated-interferon-treated patients. Serum HBV RNA levels showed different decline kinetics, and HBV RNA quasispecies showed different evolution patterns in entecavir and pegylated-interferon mono-treatment. Taken together, serum HBV RNA may serve as a promising biomarker of HBeAg seroconversion in patients during antiviral treatment.

eNOS-Nitric Oxide System Contributes to a Novel Antiatherogenic Effect of Leonurine via Inflammation Inhibition and Plaque Stabilization.

Leonurine (LEO) is a bioactive small molecular compound that has protective effects on the cardiovascular system and prevents the early progression of atherosclerosis; however, it is not clear whether LEO is effective for plaque stability. A novel mouse atherosclerosis model involving tandem stenosis (TS) of the right carotid artery combined with western diet (WD) feeding was used. Apolipoprotein E gene-deficient mice were fed with a WD and received LEO administration daily for 13 weeks. TS was introduced 6 weeks after the onset of experiments. We found that LEO enhanced plaque stability by increasing fibrous cap thickness and collagen content while decreasing the population of CD68-positive cells. Enhanced plaque stability by LEO was associated with the nitric oxide synthase (NOS)-nitric oxide (NO) system. LEO restored the balance between endothelial NOS(E)- and inducible NOS(iNOS)-derived NO production; suppressed the NF-κB signaling pathway; reduced the level of the inflammatory infiltration in plaque, including cytokine interleukin 6; and downregulated the expression of adhesion molecules. These findings support the distinct role of LEO in plaque stabilization. In vitro studies with oxidized low-density lipoprotein-challenged human umbilical vein endothelial cells revealed that LEO balanced NO production and inhibited NF-κB/P65 nuclear translocation, thus mitigating inflammation. In conclusion, the restored balance of the NOS-NO system and mitigated inflammation contribute to the plaque-stabilizing effect of LEO. SIGNIFICANCE STATEMENT: LEO restored the balance between endothelial NOS and inducible NOS in NO production and inhibited excessive inflammation in atherosclerotic "unstable" and rupture-prone plaques in apolipoprotein E gene-deficient mice. The protective effect of LEO for stabilizing atherosclerotic plaques was due to improved collagen content, increased fibrous cap thickness, and decreased accumulation of macrophages/foam cells. So far, LEO has passed the safety and feasibility test of phase I clinical trial.