Originl Pper Phrmcology 27;79:243 249 DOI: 1.1159/11989 Received: June 22, 26 Accepted: December 22, 26 Published online: April 23, 27 Nongenetic Model of Type 2 Dibetes: A Comprtive Study Md. Shhidul Islm Hymie Choi Deprtment of Food nd Nutrition, Seoul Ntionl University, Seoul, South Kore Key Words Rt dibetes model Nicotinmide Streptozotocin High-ft diet diet-fed, 4-mg/kg BW STZ-injected SD rt is better thn the LF diet-fed NA-STZ-injected rt s n niml model of humn type 2 dibetes. Copyright 27 S. Krger AG, Bsel Abstrct Seven-week-old mle Sprgue-Dwley (SD) rts were divided into six groups (LFC, LFD, HFC, HFD3, HFD4, HFD5) to determine whether nimls receiving low-ft (LF) diet plus nicotinmide-streptozotocin (NA-STZ) injection or nimls receiving high-ft (HF) diet plus STZ injection provide better model of type 2 dibetes. After 2 weeks of feeding, dibetes ws induced by intrperitonel injection of NA (23 mg/kg BW) nd STZ (65 mg/kg BW) in LFD, nd STZ 3, 4, 5 mg/kg BW to HFD3, HFD4, HFD5 groups, respectively. Fsting blood glucose t 48 72 h nd nonfsting blood glucose t 1 week fter STZ injection were 1 2 nd 1 6 mg/ dl, respectively, in HFD4 nd HFD5 groups while no significnt difference ws observed mong other groups. Serum insulin concentrtion ws significntly (p!.5) decresed in LFD, HFC, HFD3, nd HFD4 groups compred to LFC nd HFD5 groups. One niml died nd other nimls of the HFD5 group were in criticl condition. Serum lipid nd liver glycogen were incresed in HFD groups compred to other groups. The results of this study suggest tht the HF The bstrct of this rticle ws presented t the 49th Annul Meeting of the Jpn Dibetes Society, Tokyo Interntionl Forum, My 25 27, 26 nd ws lso presented t the 19th World Congress of Dibetes, Interntionl Convention Center, Cpe Town, South Afric, December 3 7, 26. Introduction Dibetes is mjor thret to globl public helth tht is rpidly getting worse nd the biggest impct on dults of working ge in developing countries. At lest 177 million subjects worldwide hve dibetes nd this figure is likely to more thn double by 23 to rech 366 million [1]. Of the two mjor types of dibetes, type 2 is the more prevlent in dults with different rnge of ge nd grdully incresing in young children nd dolescents. According to recent review [2], type 2 dibetes is incresing mong children ll over the world nd ppers to hve incresed significntly in the lst 15 yers. Type 2 dibetes ccounts for up to 45% of new cses mong dolescents. In Jpn, 8% of new cses of peditric dibetes [3] nd 7% of new cses mong Ntive Americns re type 2 dibetes [3, 4]. An epidemiologicl study suggested tht the number of ptients with dibetes is mrkedly incresing in the Asi-Pcific region [5]. In Europe, 3% of the totl popultion is suffering from type 2 dibetes nd the cost for mngement mounts to 5% of totl helth cre expenditure [6]. Type 2 dibetes is heterogeneous disorder chrcterized by progressive decline in insulin ction (insulin resistnce), followed by the inbility of -cells to com- Fx +41 61 36 12 34 E-Mil krger@krger.ch www.krger.com 27 S. Krger AG, Bsel 31 712/7/794 243$23.5/ Accessible online t: www.krger.com/ph Md. Shhidul Islm, PhD Deprtment of Nutrition, Fculty of Helth Sciences North-West University (Potchefstroom Cmpus) Potchefstroom 252 (South Afric) Tel. +27 18 299 2466, Fx +27 18 299 2464, E-Mil sislm1974@yhoo.com
T b l e 1. Compositions of experimentl diets Ingredients, g/kg Low-ft diet High-ft diet Corn strch 527. 377. Sucrose 1. 1. Csein 2. 2. Soyben oil 7. 2. Lrd. 2. Cellulose 5. 5. Vitmin fortified mixture 1. 1. Minerl premix (AIN-93M) 4. 4. L-Methionine 3. 3. Totl 1,. 1,. penste for insulin resistnce ( -cell dysfunction) [7]. The prevlence of type 2 dibetes is much higher thn tht of type 1 nd it ccounts for bout 9% of totl existing dibetic cses in the world. Animl models in dibetes reserch re very common, the rt being the first choice. Besides genetic models, nongenetic rt models re widely used in dibetes reserch for their low cost nd wide rnge of vilbility. Chemicl inducers, such s streptozotocin (STZ) nd lloxn, re used for the induction of dibetes in nimls, while nicotinmide (NA)-STZ model hs been proposed especilly for type 2 dibetes [8]. A high-ft diet with STZ for induction of type 2 dibetes in the rt hs been found to be better model for dibetes reserch [7, 9]. Moreover, the pthogenesis of dibetes in the rt model is most likely similr to the pthogenesis in humns. The dose nd type of inducers nd the composition of diets lrgely ffect the pthogenesis of dibetes in experimentl niml models. Higher doses of inducer cuse severe -cell dmge wheres lower doses fil to induce dibetes [1]. Conversely, lthough low-ft diet is unble to induce insulin resistnce [9], very-high-ft diet my cuse pncretic -cell dmge s well s permnent insulin resistnce. Although mny spects of the bove models [7 9] of type 2 dibetes hve been discussed, no comprtive study hs been done to clrify which model is better to study type 2 dibetes in humns. The present study ws initited to clrify which model of type 2 dibetes for humns is better: low-ft diet plus NA-STZ injection or high-ft diet plus STZ injection. Three different doses of STZ (3, 4, nd 5 mg/kg BW) combined with diet contining modertely high mount of ft were compred with low-ft diet nd NA- STZ injections to identify the optimum dose of STZ. Mterils nd Methods Animls Seven-week-old mle Sprgue-Dwley rts with men body weight of 217.6 8 14.74 g were provided from Animl Cre Fcility of Seoul Ntionl University. Animls were rndomly divided into six subgroups: (1) low-ft control (LFC), (2) low-ft dibetic (LFD), (3) high-ft control (HFC), (4) high-ft dibetic-3 (HFD3), (5) high-ft dibetic-4 (HFD4), nd (6) high-ft dibetic-5 (HFD5) with similr men body weight in ech group. Animls were housed in polycrbonted cges in temperturend humidity-controlled room with 12-hour light-drk cycle. Either low-ft (7% soyben oil) or high-ft (2% lrd nd 2% soyben oil) diet ws provided d libitum to corresponding groups ( tble 1 ) during 3-week experimentl period. Animls were mintined ccording to the rules nd regultions of the Seoul Ntionl University committee for experimentl nimls. Induction of Dibetes After exposure to respective diets for 2 weeks, STZ (Sigm, St. Louis, Mo., USA) dissolved in citrte buffer (ph 4.5) ws intrperitonelly injected (65 mg/kg body weight BW) 15 min fter the injection of NA (23 mg/kg BW dissolved in norml sline) to fsted nimls of the LFD group. Similrly 3, 4, nd 5 mg STZ/ kg BW ws injected to nimls of HFD3, HFD4, nd HFD5 groups, respectively, wheres norml sline nd citrte buffer were injected to control nimls insted of NA nd STZ, respectively. After 48 h but before 72 h fter injection, fsting blood glucose (FBG) ws mesured in whole blood collected from the til vein by portble Glucometer (Accu-Check Active, Roche Dignostics Limited, Germny), the mximum mesuring cpcity of which ws 6 mg/dl. Rts with n FBG level bove 2 mg/dl were considered dibetic. Nonfsting blood glucose (NFBG) level of ech niml ws lso mesured fter 1 week of injection. Food intke ws mesured dily nd body weight chnge ws recorded weekly during the experimentl period. Collection of Blood nd Liver At the end of the experimentl period nimls were scrificed fter light diethyl ether nesthesi nd blood nd liver were smpled. Liver ws wshed in cold sline, wiped with filter pper, nd weighed. Then wiped liver ws immeditely frozen in liquid nitrogen nd preserved t 8 C for subsequent nlysis. Blood ws centrifuged t 3, rpm for 15 min, serum ws seprted into individul microtubes nd preserved t 8 C for further nlysis. Anlyticl Methods Serum insulin ws nlyzed by using multiplte ELISA reder (Biord-68, Biord Ltd., Jpn) nd n ultrsensitive rt insulin ELISA kit (Mercodi AB, Uppsl, Sweden, lot No. 14154) ws used for this nlysis. Serum lipid profile ws determined photometriclly by using commercil kits purchsed from ACE Chemicls Ltd., Seoul, Kore. Liver glycogen level ws mesured by phenol-sulfuric cid method s described by Lo et l. [11]. Sttisticl Anlysis Dt re presented s mens nd their stndrd devitions. Dt were nlyzed by sttisticl softwre (Sttview, version 5, USA) by using Tukey-Krmer multiple comprison test. Sttisticl significnce of the difference between groups ws considered t p!.5. 244 Phrmcology 27;79:243 249 Islm/Choi
Fi g. 1. Food intke nd body weight chnge during the experimentl period. Dt re shown s men 8 SD. Brs with different letters re significntly different from ech other (Tukey-Krmer multiple rnge posthoc test, p!.5). Body weight (g) 35 3 25 2 15 1 5 b b BW before STZ injection BW 1 week fter STZ injection Food intke b bc LFC LFD HFC HFD3 HFD4 HFD5 c 2 18 16 14 12 1 8 6 4 2 Food intke/rt/dy (g) Fi g. 2. Blood glucose nd serum insulin concentrtions in different groups of rts during the experimentl period. Dt re shown s men 8 SD. Vlues (brs nd curve) with different letters re significntly different from ech other (Tukey- Krmer multiple rnge post-hoc test, p!.5). Blood glucose (mg/dl) 7 6 5 4 3 2 1 FBG t 48 72 h fter STZ injection NFBG 1 week fter STZ injection Serum insulin 1 week fter STZ injection b b c b c b b b b b LFC LFD HFC HFD3 HFD4 HFD5 45 4 35 3 25 2 15 1 5 Serum insulin (pmol/l) Results Although food intke ws not significntly different between experimentl groups, it ws lower in the highft groups, decresing with incresing dose of STZ injected ( fig. 1 ). This mens tht the dose of STZ is directly proportionl to the food intke especilly in groups fed high-ft diet. Although body weight before STZ injection ws not significntly different between groups, it significntly decresed in the HFD5 group compred to other groups (except the HFD4 group) 1 week fter STZ injection ( fig. 1 ). The highest reduction in body weight ws found in the HFD5 group nd 1 niml of this group died. On the other hnd, body weight of the HFD4 group ws significntly lower thn tht of HFC nd low-ft groups, but ll the nimls of this group were norml nd fed normlly fter 2 3 dys of STZ injection. FBG or NFBG levels of LFD nd HFD3 groups were not significntly (p!.5) different from corresponding nondibetic control groups ( fig. 2 ), but the FBG levels of HFD4 nd HFD5 groups were significntly higher ( 1 2 mg/dl) thn those of the HFC nd low-ft groups. The men FBG nd NFBG levels of the NA-STZ-injected LFD group were 97. 8 2. nd 162.3 8 18.2 mg/dl, respectively. The NFBG levels for HFD4 nd HFD5 groups were considerbly higher ( 1 6 mg/dl) thn those of other groups. Serum insulin concentrtion ws significntly lower for ll groups except for the HFD5 group compred to the LFC group ( fig. 2 ). Nongenetic Model of Type 2 Dibetes Phrmcology 27;79:243 249 245
Fi g. 3. Serum lipid profile t the end of the experimentl period. Dt re shown s men 8 SD. Brs with different letters re significntly different from ech (Tukey- Krmer multiple rnge post-hoc test, p!.5). Serum lipid profile (mg/dl) 18 16 14 12 1 8 6 4 2 b b b Totl cholesterol HDL cholesterol LDL cholesterol Triglyceride b b b b b b b b b LFC LFD HFC HFD3 HFD4 HFD5 b bc c T b l e 2. Liver weight, reltive liver weight, nd liver glycogen level t the end of the experimentl period LFC LFD HFC HFD3 HFD4 HFD5 Liver weight, g 8.338.85 8.78.36 7.98.1 9.181.28 8.881.3 8.581.21 Reltive liver weight, % 2.768.28 2.78.11 2.48.3 3.18.42 3.238.48 3.328.48 Liver glycogen, g/1 g tissue.668.3 1.318.94.698.11 12.68.13 b 9.4282.14 c 13.38.72 b Dt re shown s men 8 SD. Vlues with different superscript letters within row re significntly different from ech other (Tukey-Krmer multiple rnge post-hoc test, p <.5). Reltive liver weight (%) = (liver weight in g/totl body weight in g)! 1. Serum totl cholesterol nd LDL cholesterol were significntly (p!.5) incresed in the HFD groups compred to other groups while serum triglyceride ws significntly incresed only in the HFD5 group ( fig. 3 ). No significnt difference ws observed for serum HDL cholesterol mong the groups. Liver glycogen level ws significntly incresed (p!.5) in the HFD groups compred with ll other groups wheres significntly lower increment ws observed in the HFD4 group compred to HFD3 nd HFD5 groups ( tble 2 ). No significnt difference ws observed for liver weight or reltive liver weight. Discussion Type 2 dibetes is heterogeneous disorder chrcterized by progressive decline in insulin ction (insulin resistnce), followed by the inbility of -cells to compenste for insulin resistnce ( -cell dysfunction) [7]. Severl niml models hve lredy been developed to study humn type 2 dibetes [8, 9]. Animl models for type 2 dibetes usully involve using chemicl inducers, which re mostly responsible for -cell dysfunction but not for insulin resistnce. Insulin resistnce cn be obtined by feeding high-ft diet s ws found in severl previous studies [7, 9, 12]. Moreover, high-ft diet lone cuses insulin resistnce but does not ffect -cells in rodents [12]. In contrst, s the blood glucose level in type 2 dibetes is not higher s in type 1, the effect of dibetes inducer ws minimized by NA in nother type 2 dibetes model [8]. Therefore, the present study ws initited to determine whether high-ft diet plus STZ injection model or low-ft diet plus NA-STZ injection model is better to study type 2 dibetes. Norml FBG level hs recently been defined s less thn 1 mg/dl, more thn 1 mg/dl being considered s type 2 dibetes [13] ; however, sometimes levels up to 246 Phrmcology 27;79:243 249 Islm/Choi
126 mg/dl hve been considered norml for humns [14]. In our study, the FBG level for the NA-STZ-injected LFD nd HFC3 groups ws 97. 8 2. nd 84. 8 6.2 mg/dl, respectively, which is not significntly different from the corresponding nondibetic control groups ( fig. 2 ). Therefore none of these groups cn be considered s type 2 dibetic models, especilly with respect to their FBG levels. On the other hnd, the FBG levels of HFD4 nd HFD5 groups were 22. 8 4.4 nd 36.5 8 46.5 mg/ dl, respectively, which is significntly higher thn the level of ll other groups. Animls of the HFD5 group showed frnk hyperglycemi compred to the control group, nd their serum insulin levels were significntly higher thn those of other dibetic groups. The pthogenesis of type 2 dibetes usully includes moderte increment of blood glucose with insulin deficiency, but the results of the HFD5 group differ from type 2 dibetes. With regrd to blood glucose nd serum insulin the HFD4 group is closer to the pthogenesis of type 2 dibetes thn LFD nd other HFD groups. Moreover, the degree of insulin response to glucose loding nd the effects of ntidibetic gents in similr high-ft diet plus STZ injection model hve been tested successfully in previous report [7]. NFBG levels 1 3 mg/dl were considered dibetic in some previous investigtions [7, 9]. In our study, the NFBG levels of LFD nd HFD3 groups were 162.3 8 18.2 nd 37.7 8 224.6 mg/ dl, respectively ( fig. 2 ). The NFBG level of our NA-STZinjected LFD group is consistent with the result of previous NA-STZ model (155 8 3 mg/dl) [8], but it differs from the type 2 dibetes model recently presented by Srinivsn et l. [7]. This cn be clled predibetes but not type 2 dibetes model from the point of view of two recent reports [15, 16]. In ddition, Msiello et l. [8] reported tht 35-mg/kg BW NA dose completely prevented ltertions induced by 65 mg/kg BW STZ, but in nother study [17] it ws found tht intrperitonel dministrtion of NA t 5 mg/kg BW/dy for 45 consecutive dys could not prevent type I dibetes in BB rts. A similr result ws lso found in nother report [18]. Shim et l. [19] found tht NA (35 mg/kg BW) increses -cell mss nd control blood glucose in prtilly pncretectomized OLETF rts. Therefore, the effect of NA in experimentl nimls is contrdictory. Although Nkmur et l. [2] recently developed nonobese mouse model of type 2 dibetes with STZ nd NA injection, their experimentl sitution is different from our experiment. They developed nonobese type 2 dibetes mouse model specilly trgeting t the type of dibetes in Fr-Est Asin countries, e.g. Jpn, Chin, nd Kore. But in our study we compred two generl models of type 2 dibetes which re usully used in most type 2 dibetes reserch. However, lthough the NFBG level of the HFD3 group ws considerbly dibetic (37.7 8 224.6 mg/dl), the intersubject devition of this group ws very high. Moreover, the FBG level of this group ws in the norml rnge. Therefore, nimls of the HFD3 group cnnot be considered s dibetic t lest in these experimentl conditions. On the other hnd, food intke of the nimls of the HFD5 group grdully decresed ( fig. 1 ) nd 1 of the nimls of this group died within 1 week of STZ injection while the physicl condition of other nimls of this group ws rther poor. The color of their eyes ws chnged, nd body weight ws drsticlly reduced just 1 week fter STZ injection. In contrst, neither food intke nor body weight ws ffected in nimls of the LFD group dministered NA-STZ nd in control groups. On the other hnd, the physicl condition of the nimls of the HFD4 group ws quite norml; nimls moved nd fed normlly nd did not show ny bnormlities fter STZ injection. Therefore, the HFD4 group in this study cn be regrded s better niml model for type 2 dibetes thn the HFD5 group. In ddition, the HFD4 group showed considerbly incresed FBG level s in type 2 dibetes nd its high-ft diet incresed body ft content [7] s observed in overweight or obese dibetic humn subjects. Moreover, it ws reported tht diets high in sturted ft cuse severe modifictions of kidney glomeruli, which could be n independent risk fctor of dibetic nephritis [12]. However, the mount of ft in the diet of the high-ft group ws not s high s in some previous reports [7, 9], becuse very high ft content might reduce the pltbility s well s the consumption of the diet. In our experiment, 22% ft with 1% sugr ws used to increse the pltbility of the diet nd this composition did not ffect the food intke when compred with low-ft diet groups. The high-ft diet plus STZ injection model lso showed bnormlities in lipid metbolism, which re closely relted to crdiovsculr diseses nd re common finding in ptients with type 2 dibetes. The hyperlipoproteinemi observed in these high-ft/stz rts ( fig. 3 ) my be due to incresed bsorption nd formtion of low-density lipoprotein (LDL) in the form of chylomicrons following exogenous consumption of diet rich in ft or through incresed endogenous production of LDL nd its decresed uptke in peripherl tissue [21]. Similr mechnisms cn be considered for the grdully incresing Nongenetic Model of Type 2 Dibetes Phrmcology 27;79:243 249 247
level of serum triglycerides in proportion with STZ dose. In ddition, the concentrtion of totl cholesterol nd HDL cholesterol ws grdully reduced with the incresed dose of STZ. However, other lipoproteins re lso involved in triglyceride formtion in the blood strem. Moreover, the presence of higher serum lipoprotein due to high-ft diet intke could constitute source of incresed ftty cid vilbility nd oxidtion [7]. The preferentil use of incresed ftty cid for oxidtion blunts the insulin-medited reduction of heptic glucose output nd reduces the glucose uptke or utiliztion in skeletl muscle [22 24] leding to incresed glycogen synthesis in liver nd incresed liver glycogen levels in the rts of HFD groups ( tble 2 ). But using very high mount of dietry ft (31% or more) s used in some recent studies [7, 9] my cuse permnent insulin resistnce during long-term studies, nd therpeutic effects of ntidibetic food or drugs cnnot be well understood on the bsis of tht dibetic model. Therefore modertely incresed mount of ft (22%) with moderte dose of STZ (4 mg/kg BW) were used in our study. We found tht using diet with moderte ft content (22%) ws effective enough to produce insulin resistnce in our study. The concentrtion of serum insulin in the HFC group ws significntly lower thn in the LFC group ( fig. 2 ). It confirmed tht 22% ft content is enough to induce insulin resistnce compred to low-ft diet [7, 9]. Lrd ws used s principl source of ft in this study becuse sturted-ft diet induces moderte dibetes through severl mechnisms, e.g. reduction of insulin secretion nd modifiction of pncretic -cell structure [12]. High sturted-ft diet is lso n effective mens of cusing other dibetes-relted complictions (dibetic nephropthy) in rodents [12]. However, only 2% soyben oil with 2% lrd ws used to mke the highft diet in our study in order to cover the requirement of essentil ftty cids s lrd does not contin most of them. The results of our study suggest tht 7-week-old mle Sprgue-Dwley rts fed high-ft diet nd injected with 4 mg/kg BW STZ provide better niml model for short-term phrmcologicl studies of humn type 2 dibetes thn NA-STZ-injected rts fed low-ft diet. Nevertheless, it should be noted tht no informtion is vilble in this report on the use of this model in long-term studies. Acknowledgment This study ws supported by grnt from the Kore Helth 21 R nd D Project, Ministry of Helth nd Welfre, Republic of Kore (3-PJ1-PG1-CH12-2). References 1 World Helth Orgniztion Report: Globl Strtegy on Diet, Physicl Activity, nd Helth. WHO, 2. 2 Pinhs-Mmiel O, Zeitler O: The globl spred of type 2 dibetes mellitus in children nd dolescents. J Peditr 25; 146: 693 7. 3 Moore KR, Hrwell TS, McDowll JM, Helgerson SD, Gohdes D: Three-yer prevlence nd incidence of dibetes mong Americn Indin youth in Montn nd Wyoming, 1999 to 21. J Peditr 23; 143: 368 371. 4 Krosmick A: The dibetes nd obesity epidemic mong the Pim Indins. N Engl J Med 2; 97: 31 37. 5 Cockrm CS: The epidemiology of dibetes mellitus in the Asi-Pcific region. Hong Kong Med J 2; 6: 43 52. 6 Wllemcq C, Vn Gll LF, Scheen AJ: The cost of type 2 dibetes: summry of the Cost of Dibetes in Europe-Type II study (CODE- 2) nd nlysis of the sitution in Belgium (in French). Rev Med Liege 25; 6: 278 284. 7 Srinivsn K, Viswnd B, Lydi A, Kul CL, Rmro P: Combintion of high-ft dietfed nd low-dose streptozotocin-treted rt: model for type 2 dibetes nd phrmcologicl screening. Phrmcol Res 25; 52: 313 32. 8 Msiello P, Broc C, Gross R, Roye M, Mntegnetti M, Hillire-Buys D, Novelli M, Ribes G: Experimentl NIDDM: development of new model in dult rts dministered streptozotocin nd nicotinmide. Dibetes 1998; 47: 224 229. 9 Reed MJ, Meszros K, Entes LJ, Clypool MD, Pinkett JG, Gdbois TM, Reven GM: A new rt model of type 2 dibetes: the ftfed, streptozotocin-treted rt. Metbolism 2; 49: 139 1394. 1 Nishigki A, Nom H, Kkizw T: The reltions between doses of streptozotocin nd pthosis in induced dibetes mellitus (in Jpnese). Shikw Gkuho 1989; 89: 639 662. 11 Lo S, Russell JC, Tylor AW: Determintion of glycogen in smll tissue smples. J Appl Physiol 197; 28: 234 236. 12 Popov D, Simioneseu M, Shepherd PR: Sturted-ft diet induces moderte dibetes nd severe glomerulosclerosis in hmsters. Dibetologi 23; 46: 148 1418. 13 Tirosh A, Shi I, Tekes-Mnov D, Isreli E, Pereg D, Shocht T, Kochb I, Rudich A; Isreli Dibetes Reserch Group: Norml fsting plsm glucose levels nd type 2 dibetes in young men. N Engl J Med 25; 353: 1454 1462. 14 Tenenbum A, Fishmn EZ, Adler Y, Motro M, Boyko V, Behr S: Smoking nd development of type 2 dibetes in ptients with decresed functionl cpcity. Int J Crdiol 25; 14: 275 281. 15 Butler AE, Jnson J, Bonner-Weir S, Ritzel R, Rizz RA, Butler PC: Bet-cell deficit nd incresed bet-cell poptosis in humns with type 2 dibetes. Dibetes 23; 52: 12 11. 248 Phrmcology 27;79:243 249 Islm/Choi
16 Yoon KH, Ko SH, Cho JH, Lee JM, Ahn YB, Song KH, Yoo SJ, Kng MI, Ch BY, Lee KW, Son HY, Kng SK, Kim HS, Lee IK, Bonner- Weir S: Selective bet-cell loss nd lph-cell expnsion in ptients with type 2 dibetes mellitus in Kore. J Clin Endocrinol Metbol 23; 88: 23 238. 17 Hermitte L, Vilettes B, Atlef N, Pyn MJ, Doll N, Scheimnn A, Vgue P: High dose of nicotinmide fils to prevent dibetes in BB rts. Autoimmunity 1989; 5: 79 86. 18 Pn J, Chn EK, Chet D, Schrnz V, Chrles MA: The effects of nicotinmide nd glimepiride on dibetes prevention in BB rts. Life Sci 1995; 57: 1525 1532. 19 Shim K, Zhu M, Kuwjim M: A role of nicotinmide-induced increse in pncretic bet-cell mss on blood glucose control fter discontinution of the tretment in prtilly pncretectomized OLETF rts. Dibetes Res Clin Prct 1998; 41: 1 8. 2 Nkmur T, Terjim T, Ogt T, Ueno K, Hshimoto N, Ono K, Yno S: Estblishment nd pthophysiologicl chrcteriztion of type 2 dibetic mouse model produced by streptozotocin nd nicotinmide. Biol Phrm Bull 26; 29: 1167 1174. 21 Srinivsn K, Ptole PS, Kul CL, Rmro P: Reversl of glucose intolernce by pioglitzone in high-ft diet fed rts. Methods Find Exp Clin Phrmcol 24; 26: 327 333. 22 Belfiore F, Innello S: Insulin resistnce in obesity: metbolic mechnisms nd mesurement methods. Mol Genet Metb 1998; 65: 121 128. 23 Rosholt MN, King PA, Horton ES: High-ft diet reduces glucose trnsporter responses to both insulin nd exercise. Am J Physiol 1994; 266:R95 R11. 24 Iwnishi M, Kobyshi M: Effect of pioglitzone on insulin receptors of skeletl muscles from high-ft-fed rts. Metbolism 1993; 42: 117 121. Nongenetic Model of Type 2 Dibetes Phrmcology 27;79:243 249 249