KLF9 regulates hepatic lipid metabolism via inducing CD36 expression / 生理学报

The development of nonalcoholic fatty liver disease (NAFLD) is closely related to the fatty acid (FA) uptake. This study aimed to investigate the effect of Krüppel-like factor 9 (KLF9) on CD36 (typical fatty acid translocase), hepatocellular lipid metabolism as well as the development and progression of nonalcoholic fatty liver. High-fat diet-induced obese C57BL/6J mice and db/db mice were used to test the expression levels of Klf9 and Cd36 in the livers. The primary hepatocytes were isolated from C57BL/6J mice, treated with Ad-GFP, Ad-Klf9, Ad-shCtrl or Ad-shKlf9, and then incubated with oleic acid and palmitic acid for 24 h. Liver-specific knockout of Klf9 mice were established. The protein levels and relative mRNA levels were examined by Western blot and real-time PCR, respectively. Triglyceride content was determined by using an assay kit. Lipid content was determined by Oil Red O staining. The results showed that: (1) Klf9 expression levels were increased in the livers of high-fat diet-induced obese mice and db/db mice, compared to their respective control mice. (2) Adenovirus-mediated overexpression of Klf9 in primary hepatocytes increased Cd36 expression and cellular triglyceride contents. (3) In contrast, adenovirus-mediated knockdown of Klf9 expression in primary hepatocytes by Ad-shKlf9 decreased Cd36 expression and cellular triglyceride contents. (4) Finally, Klf9 deficiency decreased liver Cd36 expression and alleviated fatty liver phenotype of high-fat diet-induced obese mice. These results suggest that KLF9 can regulate hepatic lipid metabolism and development of NAFLD by promoting the expression of CD36.

Histamine stimulates thermogenesis of brown and beige fat / 生理学报

β3-adrenergic agonists induce adaptive thermogenesis and promote beiging of white fat. However, it remains unclear which metabolites mediate the stimulatory effects of β3-adrenergic agonists on thermogenesis of brown and beige fat. In this study, adipose tissue was isolated from 8-week-old C57/BL6J male mice by intraperitoneal administration of β3-adrenergic agonist CL316,243 for RNA-Seq, which revealed that histidine decarboxylase, a key enzyme in histamine synthesis, was strongly induced in adipose by CL316,243. Therefore, we speculated that histamine might be involved in the process of thermogenesis in adipose tissue. We determined the physiological role and mechanism by which histamine promotes fat thermogenesis by intravenous administering histamine to C57BL/6J mice fed a normal or a high-fat diet. The results showed that intravenous injection of histamine into C57BL/6J mice fed a normal diet stimulated the expression of thermogenic genes, including peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) and uncoupling protein 1 (UCP1), in brown adipose tissue (BAT) and inguinal white adipose tissue (iWAT). H&E staining also suggested that histamine treatment decreased the size of lipid droplets in adipocytes. Moreover, histamine treatment also enhanced thermogenesis of fat in high-fat diet induced obese mice, and improved glucose intolerance and fatty liver phenotype. Finally, we demonstrated that the effects of histamine on the thermogenic program were cell autonomous. Our data suggest that histamine may mediate the effects of β3-adrenergic agonists on thermogenesis of fat.

Detection of novel mutations related to maturity-onset diabetes of the young / 基础医学与临床

Objective To find novel genes related to maturity-onset diabetes of the young(MODY)or novel muta-tions of known MODY related genes and provide the basis for MODY diagnosis. Methods Taking the major clinical features of MODY as screening criteria,we selected four patients from the Endocrine Department of Peking Union Medical College Hospital and prepared their genomic DNA sample for whole exome sequencing.PCR and Sanger se-quencing are used to validate the sequencing results. Results Two novel mutations of the GCK gene and HNF4α gene, c.1348G>T(p.Ala450Thr)and c.758G>A(p.Arg253Gln)were found in two patients. Conclusions These two patients are both MODY patients and this is the first time the novel mutations were found.

Expression of microRNA-29b2-c cluster is positively regulated by MyoD in L6 cells / 中国医学科学杂志(英文版)

<p><b>OBJECTIVES</b>To evaluate the expression profile of myoD microRNA-29 (miR-29) family in L6 myoblast differentiated to myotube of L6 myotube treated by glucose and insulin, and to further probe the molecular mechanism of myoD regulating the expression of miR-29 clusters.</p><p><b>METHODS</b>The expression of myoD and miR-29 family was detected by using real-time PCR and Western blot analysis. The potential promoter and transcription factors binding sites of miR-29 clusters were predicted by Promoter scan and transcriptional factor search. The promoter sequence of miR-29b1-a and miR-29b2-c cluster was cloned into a luciferase reporter plasmid and the regulatory effect of myoD was analyzed by using dual luciferase reporter assay. Electrophoretic mobility shift assay was further conducted to indicate the binding of myoD on specific sequence. Moreover, overexpression of myoD was achieved by a recombinant adenovirus system (Ad-myoD). L6 cells were infected with Ad-myoD and real-time PCR was conducted to analyze the expression of miR-29b and miR-29c.</p><p><b>RESULTS</b>The expression levels of myoD, miR-29a, miR-29b, and miR-29c were increased in L6 myoblast differentiated to myotube. The expression of myoD, miR-29b, and miR-29c was up-regulated in L6 myotube treated with glucose and insulin, but miR-29a depicted no significant change. Dual luciferase reporter gene assay showed that myoD functioned as a positive regulator of miR-29b2-c expression and myoD could bind to the specific sequence located at the promoter region of miR-29b2-c cluster. Enforced expression of myoD led to a marked increase of miR-29b and miR-29c levels in L6 cells.</p><p><b>CONCLUSION</b>MyoD might act as a crucial regulator of myogenesis and glucose metabolism in muscle through regulating the expression of miR-29b2-c.</p>

Adenovirus construction of expression and its function of connective tissue growth factor / 中国医学科学院学报

<p><b>OBJECTIVE</b>To construct and identify a adenovirus vector of the expression of connective tissue growth factor (CTGF) and to explore the role of CTGF in the metabolism of glucose and lipid.</p><p><b>METHODS</b>The over-expressed plasmid of CTGF was cloned, and then the CTGF sequences were cloned into pAdTrack-CMW vector. The reformed E. coli BJ5183-sensitive bacteria that contain pAdEasy-1 were transformed with lined vector cut by Pme I enzyme. The recombinant adenovirus vector was cut with Pac I enzyme and obtained, then transfected 293A cells to produce virus. Through three times of amplification, the adenovirus infected the primary hepatocytes to determine the infection efficiency and CTGF expression. The mice were starved for several time periods, and then the liver RNA was extracted for real-time PCR to detect the expressions of CTGF under different nutritional conditions.</p><p><b>RESULTS</b>The adenovirus of CTGF was successfully produced with an infection efficiency of 90%. The expressions of the CTGF were different under different nutritional conditions and showed a coincidence with the expression of peroxisome proliferators-activated receptor gamma coactivator 1 alpha. After the mice were starved for 24h, the expression of CTGF increased by (2.38 +/- 0.51) folds; after the mice were starved for 48 h, the expression of CTGF increased by (2.95 +/- 0.57) folds (P < 0.05).</p><p><b>CONCLUSION</b>CTGF is speculated to be involved in the metabolism of glucose and lipids.</p>

A second protein marker of caveolae: caveolin-2 / 中国医学科学杂志(英文版)

Caveolin-2, a protein about 20 kD, is a major component of the inner surface of caveolae, small invaginations of the plasma membrane. Similar with caveolin-1 and caveolin-3, it serves as a protein marker of caveolae. Caveolin-1 and -2 are located next to each other at 7q31.1 on human chromosome, the proteins encoded are co-localized and form a stable hetero-oligomeric complex, distributing similarly in tissue and cultured cells. Caveolin-3 is located on different chromosomes but confirmed to interact with caveolin-2. Caveolin-2 is similar to caveolin-1 in many respects but differs from the latter in functional domains, especially in G-protein binding domain and caveolin scaffolding domain. The mRNAs of both caveolin-1 and caveolin-2 are most abundantly expressed in white adipose tissue and are induced during differentiation of 3T3-L1 cells to adipocytes. Caveolin-2-deficient mice demonstrate clear pulmonary defects, with little or no change in caveolin-1 expression and caveolae formation, suggesting that caveolin-2 plays a selective role in lung functions. Caveolin-2 is also involved in lipid metabolism and human cancers.

Current situation researching of methylation in tumor / 中国医学科学院学报

The disorders of DNA and histone methylation have a close relationship with the development and progression of tumors. Epigenetic regulation is critical in maintaining the stability and integrity of the expression profiles of different cell types by modifying DNA methylation and histone methylation. However, the abnormal changes of methylation often result in the development and progression of tumors. This review summarized the theory of tumor genomic and histone methylation, detection methods of methylation and their applications, and the clinical application of methylation as biological markers and drug targets.

Research advances in Sirt1 gene / 中国医学科学院学报

As the most homologic homologue of silent information regulator 2 of yeast, Sirt1 gene is extensively expressed in mature tissues, and is rich in early embryo and reproductive cells. It is involved in the regulation of gene transcription, energy metabolism and cell aging. It promotes fat mobilization in adipocytes and glucose production in liver and regulates insulin secretion in islet beta cell. Furthermore, Sirt1 gene is an essential endogenous apoptosis inhibitor. In future, it may be used as new drug targets or applied in other disease management modalities.

Coactivators in energy metabolism: peroxisome proliferator-activated receptor-gamma coactivator 1 family / 中国医学科学院学报

Peroxisome proliferator-activated receptor gamma coactivator 1 (PGC1) family is highly expressed in tissues with high energy metabolism. They coactivate transcription factors in regulating genes engaged in processes such as gluconeogenesis, adipose beta-oxydation, lipoprotein synthesis and secretion, mitochondrial biogenesis, and oxidative metabolism. Protein conformation studies demonstrated that they lack DNA binding domains and act as coactivators through physical interaction with transcription factors. PGC1 activity is regulated at transcription level or by multiple covalent chemical modifications such as phosphorylation, methylation and acetylation/deacetylation. Abnormal expression of PGC1 coactivators usually is closely correlated with diseases such as diabetes, obesity, hyperglycemia, hyperlipemia, and arterial and brain neuron necrosis diseases.

Gene expression profile of human skeletal muscle and adipose tissue of Chinese Han patients with type 2 diabetes mellitus / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To study the differential patterns of gene expression in skeletal muscle and adipose tissue between type 2 diabetes mellitus (T2DM) patients and healthy subjects using DNA microarray analysis.</p><p><b>METHODS</b>T2DM patiens were divided into female group, young male group and old male group. DNA microarray analysis and quantitative real-time PCR were carried out to analyze the relation between gene expressions and T2DM.</p><p><b>RESULTS</b>The mRNA expression of 298, 578, and 350 genes was changed in the skeletal muscle of diabetes mellitus patients compared with control subjects. The 1320, 1143, and 2847 genes were modified in adipose tissue of the three groups. Among the genes surveyed, the change of 25 and 39 gene transcripts in skeletal muscle and adipose tissue was > or = 2 folds. These differentially expressed genes were classified into 15 categories according to their functions.</p><p><b>CONCLUSION</b>New genes are found and T2DM can be prevented or cured.</p>

Identification and expression of non-coding RNAs NC28 and NC119 in human tumors / 中国医学科学院学报

<p><b>OBJECTIVE</b>To explore and identify the non-coding RNAs related to tumors.</p><p><b>METHODS</b>We used RT-PCR and Northern blot to analyze non-coding RNAs in tumor tissues and cell lines.</p><p><b>RESULTS</b>Two predicted non-coding RNAs were confirmed to be expressed in cancer tissues and cell lines by RT-PCR and DNA sequencing. We detected the expression of two non-coding RNA transcripts by Northern blot. The length of NC28 was about 1800 nt, and that of NC119 was about 1200nt.</p><p><b>CONCLUSIONS</b>NC28 and NC119 have a tumor-associated expression pattern. The non-coding RNAs may play a role in the development of tumors.</p>

Liver X receptor: crucial mediator in lipid and carbohydrate metabolism / 中国医学科学院学报

Liver X receptors (LXRs) are members of the nuclear receptor superfamily and are activated by oxysterols and intermediates in the cholesterol synthetic pathway. The pivotal role of LXRs in the metabolic conversion of cholesterol to bile acids has been well established. Furthermore, insulin induces LXRa in hepatocytes, resulting in the suppression of key enzymes in gluconeogenesis, including phosphoenolpyruvate carboxykinase, glucose-6-phosphatase, and fructose-1, 6-bisphosphatase (FBPase). LXRs also play an important role in fatty acid metabolism by activating the sterol regulatory element-bing protein 1c gene (SREBP1c). This articles reviews the molecular mechanisms by which LXRs act to influence the lipid and carbohydrate metabolism.

A cross-sectional study on injuries in residents from the countryside of Huanghe Delta rural areas / 中华流行病学杂志

<p><b>OBJECTIVE</b>To study the incidence of injuries among residents living in the countryside of Huanghe delta area.</p><p><b>METHODS</b>A household questionnaire survey was conducted to 15 276 residents in 20 villages of Dongying municipality of Shandong province with a stratified-cluster sampling on their injuries from March 1, 2002 to February 28, 2003. Data were analyzed with Excel 2000 and SPSS 11.0 software.</p><p><b>RESULTS</b>The crude incidence of injuries was 5.90% in total, and the standardized incidence was 5.93%. It was higher in men (7.79%) than in women (4.03%). There were 19 deaths with 20 cripples. The standardized death rate was 122.56 per 100 000 with leading causes of injuries was blunt or by sharp articles (24.61%), traffic accident (24.17%), falls (22.62%) and animal bites (13.08%). Peak incidence of age group was high in 25 - 54 age group and 0 - 4 age group (> 6.0%). 267 cases (29.60%) inpatients had had about 15.89 days hospitalization for each case. Rest of each case with injury had 19.20 days of rest. Direct economic loss for treatment would cost 904.85 RMB Yuan and 10.15 days with care takers and 221.88 RMB for other cost. The potential years of life lost was 24 years, the working years of life lost was 19.6 year, the valued years of life lost was 8.7 year, and the standardized period expected years of life lost was 31.73 year.</p><p><b>CONCLUSION</b>Injury was common and frequently occurred among residents in the countryside of rural Huanghe delta areas, that seriously endangered the health care systems and burden on families.</p>

Serial analysis of gene expression in the pituitary adenomas and para-tumor normal pituitary tissues / 中国医学科学院学报

<p><b>OBJECTIVE</b>To observe the characteristics and difference of gene expression in the pituitary adenomas and para-tumor normal pituitary tissues.</p><p><b>METHODS</b>Using serial analysis of gene expression (SAGE), two SAGE libraries were generated. Forty clones from each SAGE library were sequenced, and the results were analyzed by SAGE2000 software and compared with the SAGE map at NCBI.</p><p><b>RESULTS</b>A total of 655 gene tags, representing 43 genes, were extracted from the 40 sequence files of the para-tumor normal pituitary tissues and 737 gene tags, representing 53 genes, were extracted from the 40 sequence files of the pituitary adenomas. Of these tags, 13 were not reported before. The genes related to pituitary hormone secretion and energy metabolism were highly expressed in the two kinds of tissues. Some growth factors and cytokines were also expressed, including those involved in the immunological system. But there were also much difference of gene expression in the two tissues. Thirty-one and five tags were only detected in para-tumor normal pituitary tissues and pituitary adenomas, respectively.</p><p><b>CONCLUSIONS</b>Genes involved in hormones secretion and energy metabolism were highly expressed in the pituitary adenomas and para-tumor normal pituitary tissues. Many growth factors and cytokines were also expressed in pituitary. There was also much difference of gene expression in the two kinds of tissues. SAGE can be used not only in understanding the quantity information of gene expression, but also in finding new genes.</p>

PPAR gamma--the master of thrifty genes / 中国医学科学院学报

Peroxisome proliferation is a cellular response to many chemical compounds affects including natural and modified fatty acids, phthalate and adipate ester plasticizers, leukotriene antagonists, acetylsalicylic acid and certain pathophysiological conditions including dramatic change of cellular morphology and enzymatic activity. Peroxisome proliferation phenomenon is seen primarily in liver and kidney. Hormones and nutritional factor can regulate peroxisome proliferation response. Sustained peroxisome proliferation can lead to hepatocarcinogenesis. The three types of peroxisome proliferator activated receptor, termed PPAR alpha, PPAR beta, and PPAR gamma, expressed in specific tissue, are consisted of a specific a nuclear receptor superfamily. After more than 10 years world wide research, the function of PPAR is clarified, as PPAR gamma, the master of thrifty genes, controls the expression of genes relative to adipogenesis, diabetes mellitus and obesity. The receptor is involved in transcriptional control of numerous cellular processes including cell cycle control, inflammation, immunoregulation and carcinogenesis.

A novel gene involved in blood glucose regulation / 中国医学科学院学报

<p><b>OBJECTIVE</b>To clone a novel gene relative to blood glucose regulation.</p><p><b>METHODS</b>Rat modes of autonomous regulation of blood glucose was made by intra jugular vein to right atrium injection of high concentration of glucose solution, and the control rats were injected with 0.9%NaCl both before skeletal muscles were separated for gene analysis. The differentially expressed fragments were identified by differential display technology (DDRT-PCR). After slot blot and Northern blot analysis, the artificial positive fragments were excluded and the true EST (expression sequence tag) differentially expressed was obtained. These positive EST were used as probes to screen cDNA library of rat skeletal muscle.</p><p><b>RESULTS</b>A novel full-length cDNA, named as Fang-2 was obtained. GenBank Accession No. was AF399874. Fang-2 was found rat homologue of human troponin T by blast software (NCBI). It shared 78% identical nucleotides, which showed the family proteins were conservative. After high concentration of glucose stimulation of rats, the expression of Fang-2 was down-regulated.</p><p><b>CONCLUSIONS</b>A novel gene relative to blood glucose regulation was cloned from rat skeletal muscle. The gene can regulate blood glucose level by effect certain mechanisms unknown yet with down-regulation expression.</p>

A novel gene involved in blood glucose regulation / 中国医学科学院学报

<p><b>OBJECTIVE</b>To clonea a novel gene related to blood glucose regulation.</p><p><b>METHODS</b>Contrast rat model of autonomous regulation through jugular vein right atrium intubation of high concentration of glucose, the control rats were injected with 0.9% NaCl and skeletal muscle was separated. The differentially expressed fragments were identified by differential display technology (DDRT-PCR). After slot blot and Northern blot analysis, we excluded the artificial positive fragments and got the true EST (expression sequence tag) differentially expressed. These positive EST were used as probes to screen cDNA library of rat skeletal muscle, the coding region of full-length gene was cloned to pEGFP and L-6TG cell line was cultured and transiently transfected with the lipofectamine reagent. After 48 h, intact cells were examined with fluorescence microscope.</p><p><b>RESULTS</b>We got a novel full-length cDNA, named as Fang-1. GenBank Accession No. is AF399873. Using blast software (NCBI), we found Fang-1 is rat homologue of human AK001644. They share 82% identical nucleotides, indicating the family proteins are very conserved. After high concentration of glucose stimulation compared with control rats, the expression of Fang-1 was up-regulated and the expression product of Fang-1 was localized in both cytoplasm and cell nucleus.</p><p><b>CONCLUSIONS</b>A novel gene related to blood glucose regulation was cloned from rat skeletal muscle. The expression product of Fang-1 was localized in both cytoplasm and cell nucleus. The gene can regulate blood glucose level by controlling some mechanisms unknown now with down-regulation expression.</p>