EFFECTS OF RAPAMYCIN ON INTRACELLULAR CHOLESTEROL HOMEOSTASIS OF GLOMERULAR MESANGIAL CELL IN THE PRESENCE OF INTERLEUKIN-1β

Objective To investigate the effects of rapamycin on cholesterol homeostasis of glomerular mesangial cells and the underlying mechanisms.
Methods Intracellular cholesterol accumulation was measured by Oil Red O staining and high performance liquid chromatography. The effects of rapamycin on interleukin-1β（1L-1β）-induced mRNA and protein changes of low-density lipoprotein receptor （LDLR） and ATP-binding cassette transporter Al （ABCAl） were assayed by quantitative real-time PCR and Western blot. Transient expressions of 3 types of mammalian target of rapamycin （mTOR）, including mTOR-WT （wild type）, mTOR-RR （rapamycin resistant, with kinase activity）, and mTOR-RR-KD （rapamycin resistant, without kinase activity）, were obtained by plasmid transfection.
Results Rapamycin had no significant influence on intracellular cholesterol concentration trader normal condition, but it significantly decreased the intracellular cholesterol concentration in the presence of IL-1β. Rapamycin dose-dependently suppressed the increased expression of LDLR induced by IL-1β and up-regulated the suppressed expression of ABCAl caused by IL-1β Transient expression of 3 types of mTOR all reduced ABCAl mRNA expression significantly, which all could be overroded by rapamycin.
Conclusions Rapamycin may contribute to the maintaining of glomerular mesangial cell intracellular cholesterol homeostasis under inflammatory state by both reducing cholesterol uptake and increasing cholesterol effiux. And the effect may be not completely mediiated by mTOR.     

EFFECTS OF PDGF-BB ON INTRACELLULAR CALCIUM CONCENTRATION AND PROLIFERATION IN CULTURED GLOMERULAR MESANGIAL CELLS

Objective To investigate the relationship between the alteration of intracellular calcium concentration and proliferation in cultured glomerular mesangial cells. Methods Rat mesangial cells were cultured. lntracellular calcium concentrations were measured by confocal Laser Scanning Microscopy and Fura-3 fluorescence dyeing techniques. Cell growth was measured by MTT assay. Results PDGF-BB increased intracellular calcium concentrations in a dose-dependent manner, and at the same time promote the proliferation of mesangial cells. After preincubation with calcium channel blocker nifedipine or angiotensin converting enzyme inhibitor captopril, both the increase of intracellular calcium concentrations and cell proliferations induced by PDGF-BB were inhibited. Tripteriglum Wilfordii Glycosides （TMG） significantly inhibited the mesangial cell proliferations, but it had no significant effect on intracellular calcium concentrations. Conclusion There was a positive relationship between the elevation of intracellular calcium concentration and cell proliferation in glomerular mesangial cells, but the increase of intracellular calcium concentrations wasn＇t the only way for proliferation.     

EFFECTS OF DYNORPHIN A （ 1-17） ON MOTOR FUNCTION AND SPINAL INTRACELLULAR MESSENGER SYSTEMS IN RAT

The effect of intrathecal injection of dynorphin A(1-17) on second messenger systems of spinal cord relative to behavioral change in rats was studied. Dynorphin A(1-17) 5,10 (20nmol) caused dose-depen-dent flaccid paralysis of hindlimbs. Dynorphin A (1-17) 10, 20 nmol do,e-dependently decreased spinal adenylate cyclase (AC) activity, cyclic AMP production, calmodulin (CAM) level and cyclic-nucleotidephosphodiesterase(PDE) activity 10 rain after intrathecal injection. They recovered to a varying extent two hours later. Pretreatment -with selective K-opioid receptor antagonist nor-BN| 30 nmol 10 min before dynorphin A(1-17) markedly antagonized the effects of dynorphin A(1-17) at 20 nmol on hindlimb paralysis and inhibition of intraeelhilar second messengers. The L-type calcium channel blocker verapamil (100nmol) also played a role in blocking dynorphin neurotoxicity. The NMDA receptor antagonist APV toddpartially or completely block dynorphin inhibition of CaM level and PDE activity without affecting paralysis and decrease of AC-cAMP level induced by dynorphin A(1-17) 10 rain after intrathecal injection.     

HYDROCORTISONE POTENTIATES THE INHIBITORY EFFECT OF INTERLEUKIN-1 ON PANCREATIC BETA-CELL FUNCTION IN VITRO

To test the possible influence of hydrocortisone on the inhibitory effect of interleukin-1 on the pancreatic beta-cell function, insulin release and insulin content of newborn rat islets incubated without or with interleukin-1 and/or hydroc-ortisone were measured. Interleukin-1 (500 ng/1) resulted in a decrease of insulin release during day 1～3, but a partial recovery from inhibition during days 4～7. The insulin content of the islets was not changed significantly. Hydrocortisone had an inhibitory effect on insulin release (at 10~(-7) M) or both insulin release and insulin content of islets (at 10~(-6) M). Both 10~(-7) and 10~(-6) M hydrocortisone seem to prevent the initial decrease in insulin release exerted by interleukin-1 (500 ng/1) during day 1-3,but thereafter resulted in an even more marked decrease of insulin release correspon-ding to 51% (at 10~(-7) M) or 25% (at 10~(-6) M) of control islets, respectively, duringdays 4～7. Rebound of insulin release after interleukin-1 was not seen. Insulin c     

EFFECTS OF GLUCAGON ON ISLET β CELL FUNCTION IN PATIENTS WITH DIABETES MELLITUS

Objective To evaluate islet β cell response to intravenous glucagon(a non-glucose secretagogue)stimulation in diabetes mellitus.Methods Nineteen patients with type 1 diabetes(T1D)and 131 patients with type 2 diabetes(T2D)were recruited in this study.T2D patients were divided into two groups according to therapy:36 cases treated with insulin and 95 cases treated with diet or oral therapy.The serum C-peptide levels were determined at fasting and six minutes after intravenous injection of 1 mg of glucagon.Results Both fasting and 6-minute post-glucagon-stimulated C-peptide levels in T1D patients were significantly lower than those of T2D patients(0.76±0.36 ng/mL vs.1.81±0.78 ng/mL,P<0.05;0.88±0.42 ng/mL vs.3.68±0.98 ng/mL,P<0.05).In T1D patients,the C-peptide level after injection of glucagon was similar to the fasting level.In T2D,patients treated with diet or oral drug had a significantly greater fasting and stimulated C-peptide level than those patients received insulin therapy(2.45±0.93 ng/mL vs.1.61±0.68 ng/mL,P<0.05;5.26±1.24 ng/mL vs.2.15±0.76 ng/mL,P<0.05).The serum C-peptide level after glucagon stimulation was positively correlated with C-peptide levels at fasting in all three groups(r=0.76,P<0.05).Conclusions The 6-minute glucagon test is valuable in assessing the function of islet β cell in patients with diabetes mellitus.It is helpful for diagnosis and treatment of diabetes mellitus.     

EFFECTS OF TGF-β_1 ON THE EXPRESSION OF PLASMINOGEN ACTIVATOR INHIBITOR TYPE 1 IN CULTURED HUMAN RENAL INTERSTITIAL FIBROBLASTS

ffeSllm6 Objectif POur studier ies effets de facteur gi transfonnant de croimnce (tranSforming growth factor-ac,~ ) sur l' exPlession de nzARN de l' inhibiteur d' activateur de Plermi~ne type 1 (PAl-1 ) dens la fibrose interstitiellerdnale in vitro. met~ bofibroblastes rdnaux humans ~t iSOIds et cultivds in yi irc. ac cellules ~t stimuldes per TGFPI de diverses concentrations differenteS (de 0 d 10ng/ml ) et dens une durde diffhente (de 0 d 48h ). L' expression de ~ dePAl-1 est exclude…     

EFFECTS OF SYSTEMIC FLUCONAZOLE THERAPY ON IN VITRO ADHESION OF CANDIDA ALBICANS TO BUCCAL EPITHELIAL CELLS AND CHANGES OF THE CELL SURFACE PROTEINS OF THE EPITHELIAL CELLS

This paper presented the effects of systemic fluconazole therapy via intravenous (IV) and oral (PO) administrations on the adhesion of Candida albicans (C. albicans) to the huccal epithelial ceils (BEC) from five treated patients with three candidosis, one mucornlycosis and one sporotrichosis and at the same time,an analysis of the cell surface proteins involving candidal adherent receptor in the BEC of the patients in the course of 7 days were exposed to ^3H-leucine radiolabaled C. atbicans for in vitro eandidal adherent assay,and the BEC from first intake day and the last intake day of the patients were extracted by dithiothreitol(DTT)-iodoacetamide treatment for SDS-PAGE. These results indicate that the systemic iluconazole therapy resuks in the inhibitory effect of candldal adhesion to BEC of treated patients to prevent them from oral candidosis for a prolonged time, which is based on the absent surface protein (35KDa) of the BEC.     

EFFECTS OF NIMODIPINE ON PLATELET AGGREGATION AND THE ACTIVITY OF ENZYMES IN ARACHIDONIC ACID METABOLISM

The effects of nimodipine on platelet aggregation and arachidonic acid(AA)metabolismwere studied in order to explore its effect on patients with thrombosis or cardiovas-cular disease.The results indicate that nimodipine(50-350μmol/L)significantly inhibitsplatelet aggregation induced by ADP,AA,and ionophore A23187 in a dose dependentmanner.The inhibitory effects induced by ionophore A23187 could be partially antagonizedby calcium(1 mmol/L).When the substrate was AA and the enzyme was supplied bypig lung microsomes,nimodipine(50-400μmol/L)significantly reduced the generation ofTXB_2 and 6-keto-PGF_(1a) in parallel.When the substrate was prostaglandin endoperoxide,however,the levels of TXB_2 and 6-keto-PGF_(1a)were not significantly altered in the sameconcentration range.The results suggest that nimodipine is a cyclooxygenase inhibitor,andits ability to inhibit platelet aggregation is related to its calcium blocking effect.     

THE EFFECTS OF DEPRENYL AND 1-METHYL-4-PHENYL-1, 2, 3, 6-TETRAHYDROPYRIDINE (MPTP) ON 2-DEOXYGLUCOSE UPTAKE IN THE MOUSE BRAIN

~3H-2-deoxyglucose (2-DG) autoradiographic technique was used to study the ef feets of a monoamine-oxidase-B (MAO-B) inhibitor deprenyl and the neurotoxin Ⅰ-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) on 2-DG uptake in the mouse brain. Following MPTP intoxication, 2-DG uptake was increased in the substantia nigra and lo(?)us ceruleus. At the same time, obvious abnormal behavior of the animals was induced. In the mice pretreated with deprenyl, 2-DG uptake was similar to that of control animal. Ab normal behavior. though present, was significantly milder than in mice given MPTP alone. It is concluded that MPTP interferes with the glucose metabolism in the substantia nigra and locus ceruleus and induces remarkable abnormal behavioral syndrome of mice. These deleterious effects can be blocked by pretreatment with deprenyl.     

EFFECTS OF THE HOE 694 ON TRANSIENT INWARD CURRENT AND NA~ - CA~(2 ) EXCHANGE IN GUINEA PIG CARDIOMYOCYTES

NTRODUCTION Several studies have provided evidences that blockade of the Na＋- H＋ exchange with amiloride or its congeners is associated with the protection of heart from arrhythmias and calcium overload during ischemia and reperfusion. It also has been shown that amiloride attenuates the rise of cytosolic Ca2＋ during ischemia and prevents Na＋ and Ca2＋ overload on reperfusion(1, 2).How-ever, although the protective effects of amiloride against cardiac arrhythmias and contractile dysf…     

EFFECT OF INTERLEUKIN-1β ON GROWTH HORMONE GENE EXPRESSION AND ITS POSSIBLE MOLECULAR MECHANISM IN RAT MtT/S SOMATOTROPH CELLS

Objective To elucidate the effect of interleukin-1β (IL-1β) on human growth hormone (hGH) gene expression in a rat somatotropic pituitary cell line MtT/S. Methods Stably transfected MtT/S cells were firstly established by transfecting 484-Luc1 plasmid which contained hGH gene promoter -484 to 30 bp and luciferase reporter gene. The effect of IL-1β on hGH gene expression was determined by assaying the luciferase activities. RT-PCR method was also used to determine whether IL-1 recepor mRNA was expressed in MtT/S cells. Results The 103 U/mL IL-1β stimulated secretion and synthesis of GH, and promoted the 5’-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.38 times above the control. Among inhibitors of signaling transduction pathways, mitogen-activated protein kinase kinase (MAPKK/MEK) inhibitor PD98059 (40 μmol/L) and p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 (5 μmol/L) completely blocked the stimulatory effect of IL-1β, and phosphatidylinositol-3-kinase (PI3-K) inhibitor LY294002 partly abolished the effect of IL-1β. Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells. Neither over-expression of Pit-1 nor inhibition of Pit-1 expression affected induction of hGH promoter activity by IL-1β. A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-1β, and results showed that the stimulatory effect of IL-1β was abolished following deletion of the -196 to -132 bp fragment. Conclusions IL-1β promotes GH secretion and synthesis in rat MtT/S somatotroph cells. The stimulatory effect of IL-1β on hGH gene promoter appears to require the activation of MEK, p38 MAPK, PI3-K, and a fragment of promoter sequence that spans the –196 to –132 bp of the gene, but it may be unlinked with Pit-1 protein.