N-acetyl- l-cyste ine im proves function of islet Β cell in hyperl ip idem ic rats and its m echan ism. V o l 36 N o , NA C PCR

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1 ( ) JOU RNAL O F ZH EJ IAN G UN IV ERS IT Y (M ED ICAL SC IEN CES) V o l 36 N o h ttp: www. journals. zju. edu. cngm ed Β 1, 1, 1, 1, 1, 1, 2 ( 1. 2., ) [ ] : 2N 2 (N 2acetyl2l2cysteine, NA C) Β : 59 8 SD (N C ) (H F ) + NA C (NA C ) 20, (1) (M DA ) (GSH ) ; (2), ; (3), Β ; (4) PCR 21 ( IR S21) 22 ( IR S22) 22 (Glu t22) mrna : (1)H F M DA N C, GSH N C, NA C ; (2)H F (G IR ) N C (P < 0101), NA C G IR (P < 0101) ; H F (GS IS), NA C ; (3) H F IR S21 IR S22 Glu t22mrna 4213% 2811% 2219% ( P < 0105 ) ; NA C IR S21 IR S22 Glu t22 mrna H F 4012% 3012%, 1911% (P < 0105) :,, NA C [ ] ; g ; g ; N 2 ; Β ; PCR [ ] R [ ] A [ ] (2007) N-acetyl- l-cyste ine im proves function of islet Β cell in hyperl ip idem ic rats and its m echan ism W AN G B ing, L I Hong2liang, YAN G W en2ying, et al (D ep a rtm en t of E nd ocrinology, Ch ina J ap an F riend sh ip H osp ita l, P ek ing U n ion M ed ica l Colleg e, B eij ing , Ch ina) [Abstract ] Objective: To investigate the effect of N 2acetyl2l2cysteine (NA C ) on islet Β cell function in hyperlip idem ic rats and its m echan ism. M ethods: F ifty2n ine m ale SD rats of 8 w eek o ld w ere random ly divided in to 3 group s: no rm al diet group (N C, n= 20), h igh fat diet group (H F, n= 20) and NA C treated group (NA C, n= 19,NA C 300 m g kg - 1 d - 1 and h igh fat diet). A t the : : : ( ). : (1977- ),,, ; E2m ail: w angbing 000@ sina. com. cn. : (1946- ),,,, ; Em ail: w fl921@yahoo. com. cn

2 576 ( ) 36 end of 20 w eek s, fasting serum in su lin ( In s), gluco se (Glu), m alonaldehyde (M DA ) and reduced glu tath ione (GSH ) w ere determ ined in p lasm a and pancreas tissue. T he gluco se infu sion rate (G IR ) w as m easu red by euglycem ic hyperin su linem ia clamp to evaluate the peripheral in su lin resistance. Pancreatic islets w ere iso lated and sub jected to a perifu sion m edium con tain ing 3. 3 mmo1gl gluco se fo r 15 m in, fo llow ed by mmo1gl gluco se fo r 30 m in, in su lin con ten t of perifu sion m edium w as m easu red by R IA. T he exp ression s of IR S21, IR S22, Glu t22 gene in islets w ere detected by real tim e PCR. Results: (1) T he in su lin, gluco se and M DA concen tration in H F group w ere h igher than tho se in N C group, bu t GSH levels in p lasm a and pancreas w ere low er. NA C in terven tion cou ld reverse these effectṡ ( 2) T he G IR w as decreased sign ifican tly in H F group compared w ith N C group [ ( ) vs ( )m g m in - 1 kg - 1, P < ], NA C in terven tion reversed these effect: G IR [ ( ) vs ( )m g m in - 1 kg - 1, P < ]. (3) mmo lgl gluco se increased the in su lin secretion in the islet cells of the th ree group s, bu t the peak w as low er in H F group. NA C in terven tion reversed these effectṡ (4) T he gene exp ression of IR S21 w as sign ifican tly decreased by 42. 3% in H F group (P < 0. 05), and the exp ression s of IR S22 and Glu t22 w ere decreased by 28. 1% and 22. 9% (P < 0. 05) compared w ith N C group. In con trast, the exp ression s of IR S21, IR S22, Glu t22 in NA C group increased by 40. 2%, 30. 2% and 19. 1%, respectively than tho se in H F group. Conclusion: NA C can reverse functional diso rder of islet Βcells induced by h igh2fat2diet feeding. T h is an tiox idan t effect m igh t be associated w ith upgrading gene exp ression of in su lin signal tran sduction mo lecu les in islet Β cells. [ Key words ] A cetylcysteine; Islets of langerhan sgcyto l; In su lingsecret; N 2acetyl2l2cysteine; Islet Βcells; R eal2tim e PCR [ J Zhejiang U n iv (M edical Sci), 2007, 36 (6) : ] Β 2 ( T 2DM ), T 2DM,,, T 2DM [122 ] N 2 (N 2acetyl2l2cysteine, NA C ),, Β, PCR, NA C SD 59 (SPF, ), (N C, n = 20) (H F, n= 20) + NA C (NA C, n = 19) N C ( 64% 23% 13%, ), H F ( 20% 21% 66%,, ), NA C NA C 300 m g kg - 1 d - 1 (NA C Sigm a, 115K01261), , L inco (m alonaldehyde, M DA ) ( reduced

3 6,. Β 577 glu tath ione, GSH ) % (50 m ggkg),,,,, 12 mu kg - 1 d - 1, 20% mmo lgl, 5 6 (gluco se infu sion rate, G IR ),, 1. 4 (1) 3% (50 m ggkg),,,, 5,,,, 1 m ggm l p 8 m l,,, 6 m l H ank s (2) m in, 15 s,, 4 10 m l 4 H ank s 30 m l (3) 600 Λm, 50 m l rgm in 4 1 m in,, 4 H ank s,, H ank s 2 15 m l, rgm in 4 2 m in, (4) 25% F ico ll 4 m l, 23% 20% 11% F ico ll H ank s 2 m l, rgm in, 4 20 m in, 23% 20% 20% 11%, 4 H ank s 50 m l m l EP 1. 5 Pharm acia U pp sala 50, : (0 15 m in), 3. 3 mmo lg L, ; (16 45 m in), mmo lgl 30 m in [3 ] 1. 6 PCR T rizo l ( Invitrogen, ) RNA, ; RNA R evert ra A ce2 ΑcDNA (TO YOBO, ) cdna [4 ] AB I R PR ISM 7300 ( AB I B io system ) PCR SYBR R Green PCR M aster M ix TO YOBO ( ) PCR : SYBR R Green R ealtim e PCR M aster M ix 25 Λl, 18 Λl, 1 Λl, 1 Λl, cdna 5 Λl : s, 1 ; s; s; s; CT (GA PDH, sen se 5 2T GGGT GT G AA CCA CGA GAA 23, an tisen se 2GGCA T G GA CT GT GGTCA T GA 23, 145 bp; IR S21, sen se 5 2CA CCCA GT T T T TCGA CA CCT 23, an tisen se 5 2GCT T GT TCT T GGA GTCA GCC 23, 284 bp; IR S22, sen se 5 2CTA CCCA CT G A GCCCAA GA G23, an tisen se 5 2CCA GGG A T GAA GCA GGA CTA 23, 152 bp; Glu t22, sen se 5 2TA GCAA CT GGGTCT GCAA T 23, an tisen se 292 bp ) 5 2GTA GTCCTA CA CTCA T G23, 1. 7 SPSS x θ s,, ( ), SN K2q , H F (FBG) (F IN S) N C, NA C H F ( 1) 2. 2 H F M DA N C, GSH N C,NA C ( 2) m in [ ( 4. 80

4 578 ( ) 36 1 Table 1 Comparison of body w eigh t, visceral fat ratio, fasting b lood gluco se and in su lin level in the th ree group s of rats [x θ s o r m edian (range) ] g ( g FBGg F IN Sg g ) g% (mmo l L - 1 ) (m IU L - 1 ) N C ( ) H F # # # ( ) # NA C ( ) # F H F vs N C o r NA C vs H F, 3 P < 0. 05, # P < Table 2 Comparison of ox idative stress indexes in p lasm a and pancreas in the th ree group s of rats (x θ s) M DA g (nmo l m l - 1 ) GSH g (m g m l - 1 ) M DA g (mmo l m g - 1 p ro t) GSH g (m g g - 1 p ro t) N C H F # # # # NA C F H F vs N C o r NA C vs H F, 3 P < 0. 05, # P < )mmo lgl ], H F N C G IR [ ( ) m g m in - 1 kg - 1 vs ( )m g m in - 1 kg - 1, P < ] NA C G IR H F [ ( )m g m in - 1 kg - 1 vs ( )m g m in - 1 kg - 1, P < ], Β (gluco se stim u late in su lin secretion, GS IS) H F, F = , P < 0. 01; vs H F, 3 P < 0105, N C [ ( )m IU gl vs ( )m IU gl, P < 0. 01), GS IS,, 1g 4,, # P < F ig. 1 Comparison of gluco se infu sion rate in the th ree group s NA C H F [ ( ) 4213%, IR S22 Glu t % m IU gl vs ( ) m IU gl, P < 0. 01), 2219% (P < 0. 05) NA C IR S21 GS IS H F 2, 2 mrna H F 40. 2%, IR S PCR N C Glu t % 1911% ( P <, H F IR S21 mrna 0105), 3

5 6,. Β 579, F = , P < 0. 01; vs H F, 3 P < 0105, # P < F ig. 2 Changes of in su lin secretion du ring perifu sion in the th ree group s of rats H F N C NA C H F, 3 P < PCR F ig. 3 Changes of gene exp ression 3, Β, Β,, Β, [5 ], T 2DM Β,, T 2DM,, Β [6 ],, N C, H F, ; NA C, H F, 76. 8%, NA C, Β,,,, H F N C, mmo lg L H F, ; NA C, GS IS H F, NA C GS IS Β, Β Β Kubo ta [7 ], IR S22, Β, Β, : Β IR S21, IR S22 Glu t22mrna, H F IR S21 IR S22 Glu t22 N C 57. 7%, 71. 9% 77. 1%,, NA C IR S21 IR S 22 Glu t22 H F 40. 2%, 30. 2% 19. 1%, Β, Β, NA C, [8 ], Β? Calsson,, GS IS, ( reactive oxygen species, RO S), GSH, [9 ]

6 580 ( ) 36 M DA, GSH g,, C E, NA C, RO S, NA C, ;,NA C GSH,, GSH, GSH, C E,NA C, RO S,, NA C, M DA, GSH,, Β IR S21 IR S22 GLU T 22 mrna, NA C Β,, T 2DM References: [1 ] M A RCH IOL I R, SCHW E IGER C, L EVAN T ESi G, et al. A ntioxidant vitam ins and p revention of cardiovascular disease: ep idem io logical and clinical trial data [J ]. L ip ids, 2001, 36: S532S63. [2 ] O ZK IL IC A C, CEN G IZ M, O ZA YD IN, A et al. T he ro le of N 2acetylcysteine treatm ent on antioxidative status in patients w ith type II diabetes m ellitus [ J ]. Basic Cl in Physiol Pharmacol, 2006, 17 (4): [3 ] WU Yun2hong, L I X iu2jun, L I Hong2liang, et al (,,, ). A study on the m echanism of islet cell insulin resistance in h igh2 fat2diet obese rats [J ]. National M edical Journal of Ch ina ( ), 2005, 85 (27): (in Ch inese) [4 ] DU R ui2qin, L I Hong2liang, YAN G W eng2ying, et al (,,, ). T he exp ression of insulin signal transduction m o lecules and islet Αcell insulin resistance [J ]. National M edical Journal of Ch ina ( ), 2006, 86 (36): (in Ch inese) [5 ] M ULL ER DAN Y GUO CA I HUAN G, ST EPHAN IE AM IEL. Identification of insulin signaling elem ents in hum anβcells autocrine regulation of insulin gene exp ression [ J ]. D iabetes, 2006, (55): [6 ] GER ICH J E. T he genetic basis of type 2 diabetes m ellitus: im paired insulin secretion versus im paired insulin sensitivity [ J ]. Endocr Rev, 1998, (19): [7 ] KUBO TA N, TOBE K, T ERAU CH I Y, et al. D isrup tion of insulin recep to r substrate22 causes type 2 diabetes because of liver insulin resistance and lack of compensato ry Β2cell hyperp lasia [J ]. D iabetes, 2000, 49: [8 ] M A IESE K, M ORHAN S D, CHON G Z Z. O xidative stress bio logy and cell injure during type 1 and type 2 diabetes m ellitus [ J ]. Curr Neurovasc Res, 2007, 4 (1): [9 ] CA RL SSON C, BOR G L A,W EL SH N. Sodium palm itate induces partial m itochondrial uncoup ling and reactive oxygen species in rat pancreatic islets in v itro [ J ]. Endocr inology, 1999, (140): [ ]