研究背景及目的︰糖尿病病患常有心血管病變的問題，且因周圍神經病變使得感覺遲鈍，容易有傷口而受感染導致內毒血症，造成嚴重低血壓並使心肺功能衰退，伴隨全身性發炎反應，大量產生的腫瘤壞死基因（TNF-α）等發炎前趨物質還會造成肺部損傷。而透過運動訓練或熱休克能在大鼠體內誘發熱休克蛋白72（HSP72），可以藉由保護蛋白質架構而保護組織。於是我們假設在脂多醣（LPS）誘發的內毒血症過程中，第一型糖尿病鼠的心肺功能和存活時間會比正常鼠差。而接受運動訓練後，第一型糖尿病鼠與正常鼠體內均能誘發大量HSP72，並在內毒血症發生時，延長存活時間、維持心肺功能並保護肺組織。研究方法︰將Wistar大鼠分成誘發內毒血症組以及生理食鹽水對照組，其下又各細分四小組︰運動訓練的正常鼠和糖尿病鼠，未訓練的正常鼠和糖尿病鼠。運動組會接受為期三週、時速1英哩的漸進式跑步機運動訓練（每週五日，每日30-60分）。最後一次運動訓練後24小時，將大鼠麻醉後注射LPS誘發內毒血症，監測平均動脈壓、心跳、心輸出量及心搏量。誘發內毒血症後四小時，偵測血液氣體變化、血清及肺灌洗液中腫瘤壞死基因α （TNF-α）濃度、肺部病理切片、肺灌洗液中白蛋白含量以及肺濕乾重比。除此之外，還會觀察心臟、肺臟、肝臟、腎臟、骨骼肌及延腦背側區之孤獨核中HSP72的含量。結果︰誘發內毒血症後血液動力學參數明顯降低，血清及肺灌洗液中TNF-α量、肺中白蛋白量、肺濕乾重比明顯提升，動脈氧氣分壓明顯提升而動脈二氧化碳分壓明顯降低。病理切片中發現肺泡間隙變厚後且有白血球浸潤的現象。第一型糖尿病鼠存活時間及存活率明顯較正常鼠低，血液及肺灌洗液中TNF-α明顯較高，血液動力學參數衰退明顯較快，但兩組在肺中白蛋白量、肺濕乾重比、血液氣體分析無顯著差異。經過運動訓練的正常鼠與糖尿病鼠，在上述各個器官皆能發現大量的HSP72生成，而且存活時間明顯較未運動組延長，TNF-α明顯被抑制，血液動力參數降低的明顯較慢，肺中白蛋白量及肺乾溼重比也明顯較低，血液氣體分析無明顯變化。結論：注射LPS誘發內毒血症時，第一型糖尿病鼠存活時間明顯比正常鼠短，心肺功能亦較正常鼠差。但若事前給予糖尿病鼠和正常鼠運動訓練能在發生內毒血症過程中，有效保護組織、增加存活時間、維持心臟功能。這些效果可能與各器官組織大幅度產生的HSP72有關。

Background and purposes: Diabetic patients usually have cardiovascular complications and nerve system disease. Impaired sensation easily results in ulcers and serious infections such as endotoxemia accompanying severe hypotension and cardiopulmonary failure. Pro-inflammatory cytokines, ie. tumor necrosis factor -α (TNF-α), increase significantly with systemic inflammation and lead to pulmonary injury. However, heat shock protein 72 (HSP72) overexpression induced by exercise or heat shock in several organs of rats could attenuate tissue damages. The objective of this study is to reveal whether streptozotocin- induced diabetic rats tend to develop more severe symptoms than normal rats during lipopolysacharide (LPS)-induced endotoxemia. We also validated the hypothesis that exercise preconditioning may induce the overexpression of HSP72 in multiple organs and confer cardiopulmonary protection during endotoxemia in normal and diabetic rats. Methods: Wistar rats were randomly assigned to endotoxemia group and saline group. Each group was separated into following subgroups: sedentary normal rats, sedentary diabetic rats, normal rats with exercise and diabetic rats with exercise. The trained rats run gradually on a treadmill 5 days/week, 30-60 min/day with intensity 1.0 mile/hour for 3 weeks. At 24 hour after the last training session, we compared mean arterial pressure, heart rate, cardiac output, and stroke volume of urethane-anesthetized rats in all groups after injection of LPS. At the 4th hour after LPS injection, we determined arterial blood gas, TNF-α level in serum and lavage, histopathology of lung tissue, albumin content in lavage, and wet/dry weight ratio of lung. In addition, the HSP72 expression in multiple organs including heart, lung, liver, kidney, muscle, and nucleus of the tractus solitarius were determined in different groups. Results: After administration of LPS, we found that heart rate, cardiac output and stroke volume index dropped significantly in the diabetic animals as compared to those in the normal rats (P<0.05). In addition, the increased level of TNF-α in serum and lavage were significantly higher in diabetic rats than those in normal rats. The arterial blood gas, albumin content, and wet/dry weight ratio of lung no different among diabetic and normal rats. Finally, the survival time of diabetic rats was significantly shorter than that of normal rats after LPS administration. After 3-week exercise training, we found that HSP72 expression in multiple organs were significantly greater in rats with exercise training than sedentary rats, regardless of diabetic or normal rats. During endotoxemia, prior exercise training could significantly attenuate TNF-α level in serum and lavage, reduce albumin in lung, diminish lung wet-dry weight ratio, maintain hemodynamic functions, and prolong the survival time of rats. Conclusion: The cardiopulmonary dysfunction of diabetic rats was worse than that in normal rats during endotoxemia. Additionally, TNF-α in diabetic rats were greater in amount than that among normal rats, resulting in a higher mortality rate in diabetic rats than in normal rats. However, exercise preconditioning conferred significant protection against high mortality rate and improved cardiopulmonary function of diabetic rats during endotoxemia. This protective effect could be correlated with the overexpression of HSP72 in multiple organs in diabetic rats.

1. Ostergaard J, Hansen TK, Thiel S and Flyvbjerg A. Complement activation and diabetic vascular complications. Clinica Chimica Acta 361: 10-19, 2005.
2. Wiegel J and Mayer F. Isolation of lipopolysaccharides and the effect of polymyxin B on the outer membrane of Corynebacterium autotrophicum. Archives of Microbiology 118: 67-69, 1978.
3. Dauphinee SM and Karsan A. Lipopolysaccharide signaling in endothelial cells. Laboratory Investigation 86: 9-22, 2006.
4. Fan H and Cook JA. Molecular mechanisms of endotoxin tolerance. Journal of Endotoxin Research 10: 71-84, 2004.
5. Robert W, Schrier RW and Wang W. Acute renal failure and sepsis. New England Journal of Medicine 351: 159-169, 2004.
6. Vayssettes-Courchay C, Bouysset F and Verbeuren TJ. Sympathetic activation and tachycardia in lipopolysaccharide treated rats are temporally correlated and unrelated to the baroreflex. Autonomic Neuroscience-Basic & Clinical 120: 35-45, 2005.
7. Andersson A, Fenhammar J, Frithior R, Sollevi A and Hjelmqvist H. Haemodynamic and metabolic effects of resuscitation with Ringer's ethyl pyruvate in the acute phase of porcine endotoxaemic shock. Acta Anaesthesiologica Scandinavica 50: 1198-1206, 2006.
8. Ghosh S, Latimer RD, Gray BM, Harwood RJ and Oduro A. Endotoxin-induced organ injury. Critical Care Medicine 21: S19-S24, 1993.
9. Yoshinari D, Takeyoshi I, Koibuchi Y and Matsumoto K. Effects of a dual inhibitor of tumor necrosis factor-alpha and interleukin-1 on lipopolysaccharide-induced lung injury in rats: involvement of the p38 mitogen-activated protein kinase pathway. Critical Care Medicine 29: 628-634, 2001.
10. Shimada H, Hasegawa N, Koh H, Tasaka S and Shimizu M. Effects of initial passage of endotoxin through the liver on the extent of acute lung injury in a rat model. Shock 26: 311-315, 2006.
11. Takao Y, Mikawa K, Nishina K and Obara H. Attenuation of acute lung injury with propofol in endotoxemia. Anesthesia & Analgesia 100: 810-816, 2005.
12. Maytin EV. Heat shock proteins and molecular chaperones: implications for adaptive responses in the skin. Journal of Investigative Dermatology 104: 448-455, 1995.
13. Kregel KC. Heat shock proteins: modifying factors in physiological stress responses and acquired thermotolerance. Journal of Applied Physiology 92: 2177-2186, 2002.
14. Shephard RJ and Balady GJ. Exercise as cardiovascular therapy. Circulation 99: 963-972, 1999.
15. Rusko H. The effect of training on aerobic power characteristics of young cross-country skiers. J Sports Sci 5: 273-286, 1987.
16. Febbraio MA, Ott P, Nielsen HB, Steensberg A and Keller C. Exercise induces hepatosplanchnic release of heat shock protein 72 in humans. Journal of Physiology 544: 957-962, 2002.
17. Ascensao A, Magalhaes J, Soares JM and Ferreira R. Endurance training limits the functional alterations of rat heart mitochondria submitted to in vitro anoxia-reoxygenation. International Journal of Cardiology 109: 169-178, 2006.
18. Hung CH, Chang NC, Cheng BC and Lin MT. Progressive exercise preconditioning protects against circulatory shock during experimental heatstroke. Shock 23: 426-433, 2005.
19. Delogu G, Signore M, Mechelli A and Famularo G. Heat shock proteins and their role in heart injury. Current Opinion in Critical Care 8: 411-416, 2002.
20. Chicco AJ, Schneider CM and Hayward R. Voluntary exercise protects against acute doxorubicin cardiotoxicity in the isolated perfused rat heart. American Journal of Physiology - Regulatory Integrative & Comparative Physiology 289: R424-R431, 2005.
21. Koh Y, Lee YM, Lim CM, Lee SS and Shim TS. Effects of heat pretreatment on histopathology, cytokine production, and surfactant in endotoxin-induced acute lung injury. Inflammation 25: 187-196, 2001.
22. Van MW, Wielockx B, Mahieu T and Takada M. HSP70 protects against TNF-induced lethal inflammatory shock. Immunity 16: 685-695, 2002.
23. Lau SS, Griffin TM and Mestril R. Protection against endotoxemia by HSP70 in rodent cardiomyocytes. American Journal of Physiology - Heart & Circulatory Physiology 278: H1439-H1445, 2000.
24. Wong HR, Ryan M, Menendez IY, Denenberg A and Wispe JR. Heat shock protein induction protects human respiratory epithelium against nitric oxide-mediated cytotoxicity. Shock 8: 213-218, 1997.
25. Niu CS, Lin MT, Liu IM and Cheng JT. Role of striatal glutamate in heatstroke-induced damage in streptozotocin-induced diabetic rats. Neuroscience Letters 348: 77-80, 2003.
26. Srinivasa V, Gerner P, Haderer A, Abdi S and Jarolim P. The relative toxicity of amitriptyline, bupivacaine, and levobupivacaine administered as rapid infusions in rats. Anesthesia & Analgesia 97: 91-95, 2003.
27. Smolka MB, Zoppi CC, Alves AA, Silveira LR and Marangoni S. HSP72 as a complementary protection against oxidative stress induced by exercise in the soleus muscle of rats. American Journal of Physiology-Regulatory Integrative and Comparative Physiology 279: R1539-R1545, 2000.
28. Skidmore R, Gutierrez JA, Guerriero V and Kregel KC. Hsp70 Induction During Exercise and Heat-Stress in Rats - Role of Internal Temperature. American Journal of Physiology-Regulatory Integrative and Comparative Physiology 37: R92-R97, 1995.
29. Harris MB and Starnes JW. Effects of body temperature during exercise training on myocardial adaptations. Am J Physiol Heart Circ Physiol 280: H2271-H2280, 2001.
30. Shinohara T, Takahashi N, Ooie T, Hara M and Shigematsu S. Phosphatidylinositol 3-kinase-dependent activation of akt, an essential signal for hyperthermia-induced heat-shock protein 72, is attenuated in streptozotocin-induced diabetic heart. Diabetes 55: 1307-1315, 2006.
31. Liu YF, Lormes W, Wang LL, Reissnecker S and Steinacker JM. Different skeletal muscle HSP70 responses to high-intensity strength training and low-intensity endurance training. European Journal of Applied Physiology 91: 330-335, 2004.
32. Ensor JE, Wiener SM, McCrea KA, Viscardi RM and Crawford EK. Differential effects of hyperthermia on macrophage interleukin-6 and tumor necrosis factor-alpha expression. American Journal of Physiology 266: C967-C974, 1994.
33. Meldrum DR, Meng X, Shames BD, Pomerantz B and Donnahoo KK. Liposomal delivery of heat-shock protein 72 into the heart prevents endotoxin-induced myocardial contractile dysfunction. Surgery 126: 135-141, 1999.
34. Chen HW, Kuo HT, Wang SJ, Lu TS and Yang RC. In vivo heat shock protein assembles with septic liver NF-kappaB/I-kappaB complex regulating NF-kappaB activity. Shock 24: 232-238, 2005.
35. Criswell DS, Henry KM, DiMarco NM and Grossie VB, Jr. Chronic exercise and the pro-inflammatory response to endotoxin in the serum and heart. Immunology Letters 95: 213-220, 2004.
36. Chatterjee S, Premachandran S, Shukla J and Poduval TB. Synergistic Therapeutic Potential of Dexamethasone and l-arginine in Lipopolysaccharide-Induced Septic Shock. Journal of Surgical Research 140: 99-108, 2007.
37. Mikami K, Otaka M, Goto T, Miura K and Ohshima S. Induction of a 72-kDa heat shock protein and protection against lipopolysaccharide-induced liver injury in cirrhotic rats. Journal of Gastroenterology & Hepatology 19: 884-890, 2004.
38. Rojas M, Woods CR, Mora AL, Xu J and Brigham KL. Endotoxin-induced lung injury in mice: structural, functional, and biochemical responses. American Journal of Physiology - Lung Cellular & Molecular Physiology 288(2):L333-41, 2005.
39. Hangai-Hoger N, Nacharaju P, Manjula BN, Cabrales P and Tsai AG. Microvascular effects following treatment with polyethylene glycol-albumin in lipopolysaccharide-induced endotoxemia. Critical Care Medicine 34: 108-117, 2006.
40. Boczkowski J, Dureuil B, Branger C, Pavlovic D and Murciano D. Effects of sepsis on diaphragmatic function in rats. American Review of Respiratory Disease 138: 260-265, 1988.
41. Nin N, Cassina A, Boggia J, Alfonso E and Botti H. Septic diaphragmatic dysfunction is prevented by Mn(III)porphyrin therapy and inducible nitric oxide synthase inhibition. Intensive Care Medicine 30: 2271-2278, 2004.
42. McDeigan GE, Ladino J, Hehre D, Devia C and Bancalari E. The effect of Escherichia coli endotoxin infusion on the ventilatory response to hypoxia in unanesthetized newborn piglets. Pediatric Research 53: 950-955, 2003.
43. Hansen PD, Coffey SC and Lewis FR, Jr. Changes in oxygen consumption relative to oxygen delivery in endotoxemic dogs given adrenergic agents. Journal of Surgical Research 57: 156-163, 1994.
44. Okamoto H, Ito O, Roman RJ and Hudetz AG. Role of inducible nitric oxide synthase and cyclooxygenase-2 in endotoxin-induced cerebral hyperemia. Stroke 29: 1209-1218, 1998.
45. Baker HJ, Lindsey JR and Weisbroth SH. The laboratory rat. UK: Elsevier's science & Technology, 2006.