Analysis of genotypes and biochemical phenotypes of neonates with abnormal metabolism of butyrylcarnitine / 浙江大学学报·医学版

OBJECTIVES@#To investigate the genotypes and biochemical phenotypes of neonates with abnormal metabolism of butyrylcarnitine (C4).@*METHODS@#One hundred and twenty neonates with increased C4 levels detected by tandem mass spectrometry in the neonatal screening at Children's Hospital, Zhejiang University School of Medicine from January 2018 to June 2023 were included. The initial screening data and recalled data of C4 and C4/C3 were collected and converted into multiples of C4 reference range. Next generation sequencing was performed and the exons with adjacent 50 bp regions of ACAD8 and ACADS genes were captured by liquid phase capture technique. Variant information was obtained by bioinformatic analysis and the pathogenicity were classified according to the American College of Medical Genetics and Genomics criteria. The Wilcoxon rank sum test was used to analyze the differences in C4 levels among neonates with different variation types.@*RESULTS@#In total, 32 variants in ACAD8 gene were detected, of which 7 variants were reported for the first time; while 41 variants of ACADS gene were detected, of which 17 variants have not been previously reported. There were 39 cases with ACAD8 biallelic variations and 3 cases with ACAD8 monoallelic variations; 34 cases with ACADS biallelic variations and 36 cases with ACADS monoallelic variations. Furthermore, 5 cases were detected with both ACAD8 and ACADS gene variations. Inter group comparison showed that the multiples of C4 reference range in initial screening and re-examination of the ACAD8 biallelic variations and ACADS biallelic variations groups were significantly higher than those of the ACADS monoallelic variations group (all P<0.01), while the multiples in the ACAD8 biallelic variations group were significantly higher than those in the ACADS biallelic variations group (all P<0.01). The multiples of C4 reference range in the initial screening greater than 1.5 times were observed in all neonates carrying ACAD8 or ACADS biallelic variations, while only 25% (9/36) in neonates carrying ACADS monoallelic variations.@*CONCLUSIONS@#ACAD8 and/or ACADS gene variants are the main genetic causes for elevated C4 in newborns in Zhejiang region with high genotypic heterogeneity. The C4 levels of neonates with biallelic variations are significantly higher than those of neonates with monoallelic variations. The cut-off value for C4 level could be modestly elevated, which could reduce the false positive rate in tandem mass spectrometry neonatal screening.

The Long Chain Acyl-coenzyme A Synthetase Family and Malignant Tumors / 中国生物化学与分子生物学报

Disorders of the fatty acid metabolism can lead to cancer. The long chain acyl-coenzyme A synthetase (ACSL) family is important in fatty acid metabolism and is responsible for activating long chain fatty acids. In cancer cells, the regulatory effect of ACSLs is often disrupted, and the distribution, type, and quantity of fatty acids are altered. These alterations can lead to cancer development and other metabolic diseases. ACSLs include five subtypes in mammals, namely ACSL1, 3, 4, 5, and 6. ACSL1 is important in the synthesis and distribution of triglycerides. ACSL3 contributes to the formation of lipid droplets, which are important for maintaining lipid homeostasis. The expression of ACSL4 is related to steroid hormones and plays an important role in ferroptosis. ACSL5 can catalyze the metabolism of exogenous fatty acids but not the metabolism of de novo fatty acids. ACSL6 is important in fatty acid metabolism in the brain, spermatogenesis, and ovary. The regulatory factors of ACSLs include transcription factors, coactivators, hormone receptors, protein kinases, and small non-coding RNAs. These factors regulate mitochondria-mediated energy metabolism, endoplasmic reticulum stress, and the tumor inflammatory microenvironment through fatty acid metabolism. In addition, ACSLs serve as independent prognostic factors, biomarkers for clinical diagnosis, and therapeutic targets for various cancers. In recent years, accumulating evidence has demonstrated the important roles of ACSLs in the occurrence and development of cancer. This article focuses on the ACSL family, the relationship between ACSL and malignant tumors, and tumor therapies based on lipid metabolism by ACSLs. The information provides a theoretical basis for the further study of the ACSL family as molecular candidates for the targeted therapy of tumors.

Study on Improvement Effect of Selenium-enriched Ganoderma lucidum Crude Extract on Lipid Metabolism ，Liver Function and Inflammatory Response in Type 2 Diabetic Model Rats and Its Mechanism / 中国药房

OBJECTIVE： To study the effects of selenium-enriched Ganoderma lucidum crude extract on lipid metabolism， liver function and inflammatory response in type 2 diabetic model rats. METHODS： Totally 120 rats were randomly divided into normal control group （n＝20， normal saline） and model group （n＝100）. Normal control group was fed with normal diet， and model group was fed with high-fat diet. 4 weeks later， model group was given intraperitoneal injection of Streptozotocin solution （30 mg/kg） to induce T2DM model. After modeling， 90 rats were randomly subdivided into model control group （normal saline）， positive control group （metformin， 200 mg/kg） and selenium-enriched G. lucidum crude extract low-dose， medium-dose and high-dose groups （300， 600， 1 200 mg/kg， calculated by extract）， with 18 rats in each group. They were given medicine intragastrically， once a day， from Monday to Saturday. Half of rats in each group were selected 4， 8 weeks after medication； the serum levels of glucose and insulin were detected， and islet resistance index were calculated. The serum levels of liver function indexes （AST， ALT， AKP）， blood lipid indexes （FFA， TC， TG， LDL-C） and inflammatory factors （TNF-α， IL-6， IL-1β） were detected by ELISA. After HE staining， the histopathological changes of liver tissue were observed by microscopy. mRNA and protein expressions of peroxisome proliferator activated receptor α （PPARα） and peroxidase acyl coenzyme A oxidase 1 （ACOX1） in liver tissue were detected by RT-qPCR and Western blot assay. RESULTS： Compared with normal control group， glucose， insulin serum levels and islet resistance index were significantly increased （P＜0.01）； serum liver function indexes， blood lipid indexes and inflammatory factor levels of model control group were increased significantly in model control group after 4 and 8 weeks medication （P＜0.05 or P＜0.01）. The hepatocyte swelling of model control group was round and the volume was significantly larger than that of blank control group.  The liver had different degrees of steatosis and vacuolization， accompanied by a small amount of inflammatory cell infiltration. mRNA and protein expressions of PPARα and ACOX1 in liver tissue were decreased significantly （P＜0.05 or P＜0.01）. Compared with model control group， except that there was no significant decreased in islet resistunce index and AST， ALT， IL-6， IL-1β serum levels after 4 weeks of medication， and glucose， insulin， ALT serum levels after 8 weeks of medication and the levels of 4 blood lipid indexes after 4 and 8 weeks of medication in selenium-enriched G. lucidum crude extract low-dose group （P＞0.05）， above serum indexes of other groups were decreased significantly after 4 and 8 weeks of medication （P＜0.05 or P＜0.01）. After 4 and 8 weeks of medication， the pathological changes of liver tissue in rats were alleviated in varying degrees. protein and mRNA expressions of PPARα and ACOX1 in liver tissue were increased significantly after 4 and 8 weeks of medication （P＜0.05 or P＜0.01）. CONCLUSIONS： Selenium-enriched G. lucidum crude extract can up-regulate protein and mRNA expressions of PPARα and ACOX1 in liver tissue， promote the excretion of accumulated fatty acid and significantly improve fatty acid metabolism， inflammatory response and liver function in T2DM model rats.

Lipid storage myopathy / 中华神经科杂志

Lipid storage myopathy (LSM) is an etiologically heterogeneous group of lipid metabolic disorders characterized by accumulation of light microscopic lipid droplets in muscle fibers.This disease seems to be more common in Chinese population accounting for 3％-5％ of total muscle biopsies in several large neuromuscular centers in China.The pathogenesis of LSM is the impairment of fatty acid oxidation in muscle fibers.Late-onset multiple acyl-coenzyme A dehydrogenase deficiency (MADD) caused by electron transfer flavoprotein dehydrogenase (ETFDH) gene mutation has been demonstrated to be the main molecular defect in China.Three frequent ETFDH mutations were identified:c.250G＞A in patients from South China,and c.770A＞G and c.1227A＞C in those from both South and North China.More importantly,almost all late-onset MADD are dramatically responsive to riboflavin supplementation.Neutral lipid storage disease with myopathy (NLSDM) caused by mutations in PNPLA2 gene is the second common cause of Chinese LSM.Distal muscle involvement and asymmetrical muscle weakness and atrophy are common in primary symptoms of NLSDM which may be the first clue indicating the diagnosis of NLSDM.There were also a few case reports showing that LSM may be caused by carnitine transport defect and other deficiencies of acyl-coenzyme A dehydrogenase involved in fatty acid beta oxidation.Increased lipid droplets accumulation in muscle fibers may also be a secondary consequence of mitochondrial myopathy (mtDNA depletion syndrome or MELAS),dermatomyositis and steroid treatment.

Silibinin ameliorates hepatic lipid accumulation and oxidative stress in mice with non-alcoholic steatohepatitis by regulating CFLAR-JNK pathway

Non-alcoholic steatohepatitis (NASH) is a chronic metabolic syndrome and the CFLAR-JNK pathway can reverse the process of NASH. Although silibinin is used for the treatment of NASH in clinical, its effect on CFLAR-JNK pathway in NASH remains unclear. This study aimed to investigate the effect of silibinin on CFLAR-JNK pathway in NASH models both and . The study was performed using male C57BL/6 mice fed with methionine- choline-deficient diet and simultaneously treated with silibinin for 6 weeks. The study was performed by using mouse NCTC-1469 cells which were respectively pretreated with oleic acid plus palmitic acid, and adenovirus-down for 24 h, then treated with silibinin for 24 h. After the drug treatment, the key indicators involved in CFLAR-JNK pathway including hepatic injury, lipid metabolism and oxidative stress were determined. Silibinin significantly activated CFLAR and inhibited the phosphorylation of JNK, up-regulated the mRNA expression of and , reduced the activities of serum ALT and AST and the contents of hepatic TG, TC and MDA, increased the expression of NRF2 and the activities of CAT, GSH-Px and HO-1, and decreased the activities and expression of CYP2E1 and CYP4A . These effects were confirmed by the experiments. Silibinin prevented NASH by regulating CFLAR-JNK pathway, and thereby on one hand promoting the -oxidation and efflux of fatty acids in liver to relieve lipid accumulation, and on the other hand inducing antioxidase activity (CAT, GSH-Px and HO-1) and inhibiting pro-oxidase activity (CYP2E1 and CYP4A) to relieve oxidative stress.

A family affected by multiple acyl-CoA dehydrogenase deficiency due to compound heterozygous mutations on ETFDH and literature review / 中国新生儿科杂志

Objective To improve the understanding of clinical phenotype and genotype of multiple acyl-CoA dehydrogenase deficiency (MADD) in neonates.Method The clinical data of a neonates with the diagnosis of MADD and treated in the Neonatal Department of Children's Hospital of Capital Institute of Pediatrics in December 2016 were analyzed.The literature collected from Wanfang database,CNKI and PubMed database from 1976 January to 2017 June was retrieved.Using "glutaric acidemia type Ⅱ ","multiple acyl CoA dehydrogenase deficiency","infant" and "neonate" as the key words.The phenotype and genotype characteristics were summarized.Result This boy was a full-term low birth weight infant with abnormal family history.He was admitted to hospital with recurrent episodes of poor response,respiratory distress and hyperlactacidemia.B-mode ultrasound abdominal examination suggested polycystic kidney disease.Laboratory tests revealed non-kenotic hypoglycemia,refractory metabolic acidosis,elevated lactate and muscle enzymes,hyperammonemia,abnormal coagulation function test.Mass spectrometry analysis showed that multiple acyl-carnitine increased.Urine gas chromatography-mass spectrometry showed significantly increased levels of lactic,glutaric,2-hydroxypentanedioic,dicarboxylic,and 4-hydroxybenzene lactic acids.The infant was given high doses of vitamin B2,L-carnitine,and other symptomatic treatments,but the condition did not improve.He died 5 days later.The gene test showed ETFDH gene compound heterozygous mutations,one missense mutations from the father with normal phenotype c.770A ＞ G (p.Y257C),a frameshift mutation from the mother with normal phenotype c.1281-1282 deletion mutation of AA (p.I428Rfs6).The protein structures of the mutations were predicted to be deleterious.Frameshift mutation c.1281-1282 deletion mutation of AA (p.I428Rfs6) were not included in the gene bank.A total of 21 cases with MADD were found from the literature.The clinical characteristics including:male (76.2％),dyspnea (52.4％),poor response (52.4％),hypoglycemia (47.6％),hepatomegaly (47.6％),elevated muscle enzymes (42.9％),immediate onset within 24 hour of birth (42.9％),abnormal family history (38.1％),malformation (38.1％),hyperammonemia (33.3％),metabolic acidosis (28.6％).81.0％of the patients were given vitamin B2 treatment,71.4％ of carnitine,28.6％ of coenzyme Q10,28.6％ of low fat,low protein and high carbohydrate feeding.However,the prognosis of these patients was poor,76.2％ died,and 42.9％ died within 1 week after birth,and 23.8％ survived.But all showed different degrees of mental retardation during follow-up periods.Conclusion Neonatal onset MADD can be characterized by dyspnea,poor response,hypoglycemia,hepatomegaly and elevated muscle enzymes.The disease is more common in early male neonates.It can be treated with vitamin B2 and L-carnitine,but with poor prognosis and high mortality.In this case,there were 2 sites in the ETFDH gene that formed complex heterozygous mutation:c.770A ＞ G (p.Y257C) and c.1281-1282 deletion mutation of AA (p.I428Rfs6),while the latter is a new mutation.

Effect of silencing ACAT1 gene by RNA interference on proliferation, migration and invasion of human colon cancer cells / 实用医学杂志

Objective To evaluate the effect of silencing ACAT1 gene on colon cancer cells proliferation,migration,invasion and colon cancer development by using the small interference RNA (siRNA) in colon cancer cell line HT-29.Methods Acyl coenzyme A cholesterol acyltransferase 1 (ACAT1) gene was silenced in HT-29 cell lines using Hiperfect transfection reagent.The expression level of ACAT1 was detected by real time PCR.CFSE and transwell assays were used to evaluate the effect of ACAT1 gene interfering on cells proliferation,mi gration and invasion.Result ACAT1 mRNA expression decreased obviously after siRNA interference.Compared with pre-transfection,proliferation,migration and invasion of colon cancer cells have been significantly inhibited (P ＜ 0.05).Conclusion ACAT1 gene interference reduced proliferation,migration and of invasion of HT29 cells,which provide a new potential target for colon cancer treatment.

Study on the Inhibitory Effect of Ligustrazine on the Formation of Macrophage-derived Foam Cells / 中国药房

OBJECTIVE:To investigate the inhibitory effect of ligustrazine on the formation of macrophage-derived foam cells and its possible mechanism. METHODS:Human acute mononuclear cells (THP-1) were incubated with 160 nmol/L phorbol ester (PMA) for 24 h to differentiate into macrophages;and the macrophages were incubated with oxidized low-density lipoprotein (ox-LDL)culture solution containing 80 mg/L for 24 h to differentiate into macrophage-derived foam cells. And then the cells were randomly divided into blank control group(ox-LDL),model(AngⅡ)group,positive control(valsartan)group,and ligustrazine low,medium and high concentration groups(the mass concentration were 0.025,0.05 and 0.1 g/L). After all cells were respective-ly incubated with 80 mg/L ox-LDL culture solution for 48 h,oil red O staining was adopted to observe the transformation rate of foam cells,enzyme chemical method was used to determine the content of cholesterol,and real-time quantitative polymerase chain (RT-PCR)and Western blot were conducted to detect expression levels of Acyl-coenzyme A cholesterol acyltransferase-1(ACAT-1) mRNA and its protein. RESULTS:Compared with blank control group,the transformation rate of foam cells and content of choles-terol in model group were increased,and the expression levels of ACAT-1 mRNA and its protein were obviously strengthened,with significant differences(P<0.01 or P<0.05). Compared with model group,the transformation rate of foam cells and content of cho-lesterol in positive control group(valsartan)and ligustrazine low,medium and high concentration groups were decreased,and the expression levels of ACAT-1 mRNA and its protein were obviously weakened,with significant differences (P<0.01 or P<0.05). CONCLUSIONS:Ligustrazine can inhibit the macrophages differentiating into foam cells,by a mechanism that may be related to inhibiting expression of ACAT-1,and reducing content of cholesterol to reduce formation of foam cells.

Effects of total flavonoids from Dracocephalum moldovica on formation of mouse peritoneal macrophage-derived foam cells and study on their mechanisms / 中草药

Objective: To study the effects of total flavonoids from Dracocephalum moldovica on the formation of mouse peritoneal macrophage-derived foam cells and their possible mechanisms. Methods: The mouse peritoneal macrophages were induced with oxidized low density lipoprotein (Ox-LDL) to establish foam cell model, and were intervened with different concentration of total flavonoids from D. moldovica. The foam cells were observed by oil red O staining and the contents of cholesterol ester in cells were detected by fluorescence spectrophotometric method. The expression levels of acyl-coenzyme A, cholesterol acyltransferase-1 (ACAT-1) mRNA in cells and scavenger receptor A (SR-A) on cells were determined using real-time quantitative PCR (RT-qPCR) and flow cytometry, respectively. Results: The Ox-LDL-induced formation of foam cells, accumulation of cholesterol ester in cells, and the expression levels of ACAT-1 mRNA in cells and SR-A on cells were significantly inhibited by total flavonoids from D. moldovica, which showed a tendency toward a dose-dependent manner. Conclusion: These data suggest that total flavonoids from D. moldovica could inhibit the formation of macrophage-derived foam cells, which is probably one of the underlying mechanisms of the total flavonoids for clinical treatment of atherosclerosis.

Hot spot mutations in electron transfer flavoprotein dehydrogenase gene of riboflavin responsive lipid storage myopathy in 20 Chinese families / 中华神经科杂志

Objective To report the spectrum of electron transfer flavoprotein dehydrogenase (ETFDH)gene mutations in 20 Chinese RR-LsM families.Methods Twenty-four RR-LSM patients in the First Hospital of Peking University from January 2003 to May 2010 were collected and the clinical characteristics were analyzed.These patients came from 20 families in North Mainland China.Sixteen families had 1 patient each.and the other 4 families had 2 patients.ETFDH gene analysis was performed in all patients,11 family members and 100 healthy controls.Results The mean onset age was(27.9±9.9)years.The main symptoms were limb weakness(21,87.5%),dysmasesia(15,62.5%),neck weakness (14,58.3%)and myalgia(14,58.3%).Eighteen patients had high level of acyleamitine.Fifteen of 17patients had glutaric aciduria.Seventeen ETFDH mutations,including 13 missense mutations,2 splice mutations,and 2 nonsense mutations,were identified in 19 families:c.998A>G,c.1450T>C,c.1703T>C,c.1717C>T,c.821G>A,c.643G>A,c.251C>T,c.1763A>T,c.IVS7+2T>C and c.IVS6+1G>A were Hovel mutations which were not found in 100 healthy controls.Nine families had the mutation of c.770A>G(P.Y257C)and 5 families had the mutation of c.1227A>C(P.L409F).Conclusions The numerous novel mutations in ETFDH gene indicate that Chinese RR-LSM might have special mutation pattern.c.770A>G(P.Y257C)and c.1227A>C(p.L409F)may be hot spot mutations in North Mainland China.

Clinical features and electron transfer flavoprotein dehydrogenase gene mutation analysis in 35 Chinese patients with lipid storage myopathy / 中华神经科杂志

Objective To investigate the clinical features and electron transfer flavoprotein dehydrogenase (ETFDH) gene mutations in 35 Chinese patients with lipid storage myopathy. Methods The clinical data of 35 cases with lipid storage myopathy confirmed by muscle biopsy were collected. The sequences of all 13 exons of ETFDH were analyzed. Results All 35 patients showed proximal weakness. Ten of them demonstrated masseter weakness and 28 of them showed weakness in neck flexion. Twenty-nine of 32 patients who were followed up showed improvement after treatment with VitB2 and CoQ10. Mutations of ETFDH were found in 30 of 35 patients,which included 8 homozygosises,20 compound heterozygosises and 2 single heterozygosises. Fourteen novel mutations were found, including 9 missense mutations ( c. 3G > C, c. 152G>A, c. 191G > A, c.349G>C, c.433G>C, c. 949C > A, c. 1454C > G, c. 1744A >T and c. 1763A>G), 1 nonsense mutation(c. 172G>T), 2 deletions(c. 1282_1283del and 1773_1774del) and 2 splice mutations (c. 405 + 1G > T and c. 1691 -3C > G). Nine of them showed c. 250G > A mutation and 6 of them showed c. 770A > G mutation. Conclusions Lipid storage myopathy is presented as proximal weakness. Multiple acyl-CoA dehydrogenase deficiency caused by mutations of ETFDH is the major cause of lipid storage disease in this group. ETFDH c. 250G > A and c. 770A > G mutations show a high frequency.

Impact of acyl-coenzyme A cholesterol acyltransferase inhibitor on cholesterol homeostasis in human mesangial cells / 中华肾脏病杂志

Objective To study the effects of acyl-coenzyme A cholesterol acyltransferase inhibitor(ACATI, 58-035) on cholesterol homeostasis in lipid-loaded human mesangial cell line (HMCL) . Methods Oil red O was used to examine the lipid droplet . MTT was performed to detect the cell proliferation . High performance liquid chromatography (HPLC) was used to measure intracellular free cholesterol (FC) and cholesterol ester (CE) . Western blot was used to demonstrate the protein level . Real-time PCR was performed to detect the mRNA expression .Results In HMCL loaded with 100 mg/L of low density lipoprotein (LDL), 10 mg/L 58-035 significantly inhibitted the formation of lipid droplets and CE mass (ratios over control, 1 .91±0 .36 and 1 .07±0 .30 respectively, P＜0 .01) without toxic effect . It further enhanced the ACAT1 protein expression (ratios over control, 1 .27 and 1 .77 respectively) without significant influence on mRNA level, since the activity of ACAT1 pl promoter by transient transfection was not affected by either LDL or ACATI . 100 mg/L LDL down-regulated the mRNA of LDL receptor (LDLR) (ratio over control, 0 .08±0 .02, P＜0 .01)and up-regulated that of ATP binding cassette transporter 1(ABCA1)(ratio over control, 2 .97±0 .39, P＜0 .01) . The mRNA expression of ABCA1 was further upregulated (ratio over control 4 .41±1 .27, compared with LDL group P＜0 .05), and LDLR was further down-regulated (ratio over control 0 .04±0 .005, compared with LDL group P＜0 .01) in the presence of 10 mg/L ACATI 58-035 . Conclusions The CE mass in lipid-loaded HMCL can be decreased in the presence of ACATI, accompanied by further down-regulation of LDLR mRNA expression and up-regulation of ABCA1 protein and mRNA expression . It is physiological feedback to inhibit free cholesterol accumulation to maintain cholesterol homeostasis .

Effect of Tumor Necrosis Factor-α on Acyl Coenzyme A: Cholesteryl Acyltransferase Activity and ACAT1 Gene Expression in THP-1 Macrophages / 华中科技大学学报（医学）（英德文版）

In order to explore the effect and mechanisms of tumor necrosis factor-α (TNF-α) on the activity of the acyl coenzyme A: cholesteryl acyltransferase (ACAT), THP-1 monocytes were cultured and induced to differentiate into macrophages with phorbol ester. TNF-α (60 ng/mL) was added at different time points into the macrophage-containing medium and the ACAT enzyme activity was measured by quantifying the incorporation of [1-14C] oleoyl CoA into cholesteryl esters. The expression of ACAT-1 protein and mRNA was respectively detected by Western blotting and RT-PCR in THP-1 macrophages 24 h after treatment with TNF-α (60 ng/mL). The results indicated that ACAT activity in THP-1 macrophages treated with TNF-α was increased in a time-dependent manner. The expression levels of ACAT-1 protein and mRNA were significantly increased in THP-1 macrophages after treatment with TNF-α (P＜0.05). It was suggested that TNF-α could increase the activity of ACAT in THP-1 macrophages by up-regulating the expression of ACAT-1 gene.

Study of the inborn errors of mitochondrial fatty acid ?-oridation deficiency / 北京大学学报(医学版)

Mitochondrial fatty acids ?-oxidation is a repetitive process of four steps which provides the major source of energy for heart, liver and skeletal muscle. Several enzymes are involved in this spiral cycle. The medium-chain acyl-CoA dehydrogenase ( MCAD) , the short-chain acyl-CoA dehydrogenase (SCAD) , the long-chain 3-hydroxy acyl-CoA dehydrogenase (LCHAD) and the carnitine-palmitoyl-CoA transferase Ⅱ ( CPT Ⅱ ) deficiency have been recognized as the most common inborn errors of metabolism and frequently reported in their association with sudden infant death ( SID) . The prevalent mutations in these genes need further investigation in different populations.

Effects of advanced glycation end-products on expression of acyl coenzyme A:cholesterol acyltransferase-1 in THP-1 macrophages / 第三军医大学学报

Objective To investigate the effects of advanced glycosylation end-products (AGEs) on the expression of acyl coenzyme A: cholesterol acyltransferase-1 (ACAT-1) in cultured THP-1 macrophages. Methods THP-1 cells were differentiated into macrophages after cultured in RPMI1640 media containing 0.1 ?mol/L PMA for 72 h. THP-1 macrophages were then exposed to AGE-modified bovine serum albumin (AGE-BSA, at concentration of 50, 100, 200 mg/L) for 24 h or to 200 mg/L AGE-BSA for 0, 12, 24, 36 h. Expression of ACAT-1 mRNA and protein in THP-1 macrophages was measured by RT-PCR and Western blot, respectively. Results After induced by 0.1 ?mol/L PMA, THP-1 cells stopped proliferation and differentiated into macrophages. Treatment of THP-1 macrophages with AGE-BSA resulted in an increase in the mRNA and protein levels for ACAT-1 in a dose-and time-dependent manner. Conclusion AGEs upregulate the expression levels of mRNA and protein for ACAT-1 in cultured THP-1 macrophages, which might be partly involved in the atherogenesis in diabetic patients.

Effects of valsartan on the expression of acyl-coenzyme A:cholesterol acyltransferase-1 and peroxisome proliferator activated receptor-gamma in experimental atherosclerosis in rabbits / 解放军医学杂志

Objective To investigate the effects of valsartan on the expression of acyl-coenzyme A: cholesterol acyltransferase-1(ACAT-1) and peroxisome proliferator activated receptor-gamma(PPAR-?) in atherosclerotic plaques on rabbit aortic wall.Methods Twenty-four male Japanese white rabbits were randomly assigned into three groups(8 each): control group,valsartan group and high cholesterol feeding group.All rabbits were fed according to the experimental protocol for 12 weeks.Blood samples were taken from vein for measurement of serum lipids.The ratio of intima/media thickness of the aorta was measured.ACAT-1 mRNA/protein and PPAR-? mRNA/protein were determined by reverse transcription polymerase chain reaction(RT-PCR) and Western blotting,respectively.Results After 12 weeks,the levels of serum total cholesterol(TC),triglyeride(TG) and low density lipoprotein-cholesterol(LDL-C) in valsartan group and cholesterol group were significantly higher than those in control group(P0.05).The intima thickness and the ratio of intima/media in carotid arteries in cholesterol group were significantly higher than those in control group and valsartan group(P

APOPTOSIS OF HUMAN OVARY GRANULOSA CELLS INDUCED BY THE METABOLITES ACYL-COA OF SATURATED FREE FATTY ACIDS / 解放军医学杂志

To investigate the apoptotic effect of free fatty acids (FFAs) on human ovary granulosa cells, the granulosa cells were treated with FFAs in various concentrations for three days, and then the cell viability was determined by trypan blue exclusion method. DNA fragmentation was examined by DNA ladder formation and Annexin V GFP/PI staining of the cells. We also determined if the apoptotic effect of saturated FFAs was mediated by their acly CoA metabolites and measured the expression of apoptosis related genes, Bcl 2 and Bax by Western blot. The results indicated that saturated FFAs induced apoptosis of granulosa cell by their acly CoA metabolites in a dose dependent manner, and led to the down regulation of Bcl 2 and the up regulation of Bax. It is concluded that saturated FFAs can induce apoptosis of human ovary granulosa cells, suggesting that the sexual disorder of obese and insulin resistant women is related to it.

Effect of acyl coenzyme A:cholesteryl acyltransferase 1 antisense oligonucleotides on the formation of foam cells / 中国病理生理杂志

AIM:To study the effect of acyl coenzyme A:cholesteryl acyltransferase 1(ACAT1)antisense oligonucleotides on the formation of foam cells(FC).METHODS:THP-1 cells were cultured and differentiated into macrophages(MP)by phorbol myristate acetate(PMA).Over-expressing ACAT1 gene THP-1 cells were constructed.The ACAT1 antisense and missense oligonucleotides conducted by LipofectamineTM 2000 were incubated with above cells.Ac-LDL was added 6 h later and incubated for 24 h.The expression of ACAT1 protein was detected by Western blotting.The ACAT activity was measured by quantifying the incorporation of 1-14C oleoyl CoA into cholesteryl esters.The formation of foam cells was detected by oil red O staining.RESULTS:The ACAT1 antisense oligonucleotides inhibited the activity of ACAT in macrophages and over-expressing ACAT1 gene THP-1 cells.It also inhibited the formation of foam cell in macrophages and over-expressing ACAT1 gene THP-1 cells with lipid loading.The missense oligonucleotides did not show the inhibitory effects.CONCLUSION:The ACAT1 antisense oligonucleotides inhibit the activity of ACAT and the formation of foam cells.

ACTIVITY ASSAY OF ACYL COENZYME A OXIDASE(ACO) FROM CANDIDA TROPICALIS / 微生物学通报

Acyl Coenzyme A Oxidase(ACO) is the rate-limiting enzyme of the ?-oxidation of dicarboxylic acid in Candida tropicalis . Using strains with low Acyl Coenzyme A Oxidase activity to produce dicarboxylic acid on an industrial scale will facilitates the improvement of the yield and purity of dicarboxylic acid. In this research, we improved the method of Allain, so as to determine the activity of Acyl Coenzyme A Oxidase from Candida tropicalis easily and correctly. This method can be used as quantity criterion in the screen of industrial strains of dicarboxylic acid production, and this work established the basic of character study of Acyl Coenzyme A Oxidase. We determined the activities of Acyl Coenzyme A Oxidase from a series of Candida tropicalis strains with different ability of dicarboxylic acid production, and found the obvious relationship of ACO activity and ability of acid production.

Alteration of the acyl chain composition of free fatty acids, acyl coenzyme A and other lipids by dietary polyunsaturated fats.

Dietary alterations were used to demonstrate selective handling of fatty acids during their redistribution in vivo. Differences in the mol per cent of individual acyl chains in the non-esterified fatty acid, acyl-coenzyme A and phospholipid fractions reflected a result of relative precursor abundance combined with enzymic selectivities. Selective distributions were observed in the utilization of individual acyl chains between 16:0 and 18:0, 18:1 and 18:2, and among 20:3, 20:4 and 20:5, 22:6 by ligase(s), hydrolase(s) and acyl-transferases. The variations in the mol per cent of linoleate present in the acyl-coenzyme A fraction of liver relative to that in the non-esterified fatty acids suggested an in vivo regulation of the level of linoleoyl-coenzyme A that influenced the synthesis of both arachidonoyl-coenzyme A and lipids. The greater abundance of eicosapentaenoic acid in the free fatty acid fraction relative to that in the acyl-coenzyme A fraction may increase the ability of dietary 20: 5n-3 to be an effective inhibitor of the synthesis of prostaglandins derived from 20:4n-6.