在現今社會，化學物質對苯二胺 (para-phenylenediamine; PPD) 因具有使毛髮氧化脫色之能力，因此常會被添加於一般民眾所使用之永久性染髮劑中。然而，PPD的使用長期以來被認為對於膀胱功能有所損害以及發生接觸性皮膚炎。為了釐清PPD對於膀胱造成的可能損害，我們使用腹腔注射的方式將PPD施打進入實驗動物體內為期一個月，以探討PPD對於膀胱功能的影響。藉由膀胱頂部插管以固定流速注入生理食鹽水的方式，進行膀胱壓力檢測；除此之外，為了更深入探討PPD所引起的傷害性，我們抽取了實驗動物的血液以及尿液樣本進行自由基數量的檢測，以及進行膀胱活體自由基數量檢測，此外也採得實驗動物的檢體進行病理切片觀察與使用西方墨點法進行計劃性細胞死亡之相關蛋白定量。而實驗結果顯示出，PPD會使膀胱收縮變得頻繁，也會造成血液與膀胱內部的氧化壓力增加。從病理切片發現到，PPD會使膀胱產生發炎。而西方墨點法結果也顯示：PPD會誘發兩種計畫性細胞死亡(細胞凋亡與細胞自我吞噬)。綜合上述結果可得知，PPD不僅會造成膀胱收縮功能的異常，也會促使血液中氧化壓力的上升以及膀胱組織的發炎，更會進一步使膀胱組織產生兩種不同路徑之計畫性細胞死亡。

Para-phenylenediamine (PPD) is a chemical substance that is widely used as a permanent hair dye. PPD via its toxic metabolites may induce some urinary bladder lesions and allergic contact dermatitis. To determine the occurrence and severity of urinary bladder dysfunction possibly caused by PPD, we used intraperitoneal injection of PPD for 4 weeks for
induction of chronic bladder injury in the female Wistar rats. I measured bladder cystometrogram by saline infusion from the bladder dome in urethane anesthetized rats. I also measured the reactive oxygen species (ROS) in vitro and in vivo for oxidative stress and observed pathological changes in the formalin-fixed bladder sections. In addition, We explored the protein expression of apoptosis、autophagy and pyroptosis by western blot. The results showed that PPD treatment increased the frequency of micturition. PPD treatment also increased the level of ROS in blood and urinary bladder (H2O2 related oxidative stress). We also found that PPD induced the number of leukocyte infiltration including neutrophils and mast cells in the damaged bladder sections. In addition, the result of western blot indicated that PPD treatment markedly induced autophagy, apoptosis and but not caspase-1 mediated pyroptosis. Our study results indicate that PPD induces oxidative stress and inflammatory leukocytosis in the urinary bladder, which subsequent causes bladder dysfunction, possibly involving autophagy and apoptosis menchanism.

Abrams P, Cardozo L, Fall M et al. The standardisation of terminology of lower urinary tract function: report from the Standardisation sub-committee of the International Continence Society. Neurourol Urodyn 21, 67-78, 2002.

Adams JM, Cory S. The Bcl-2 protein family: arbiters of cell survival. Science 281, 1322-6, 1998.

Antonsson B, Montessuit S, Lauper S et al. Bax oligomerization is required for channel-forming activity in liposomes and to trigger cytochrome c release from mitochondria. Biochem J 345, 271-8, 2000.

Antonsson B, Conti F, Ciavatta A et al. Inhibition of Bax channel-forming activity by Bcl-2. Science 277, 370-372, 1997.

Andrew AS, Schned AR, Heaney JA et al. Bladder cancer risk and personal hair dye use. Int J Cancer 109, 1-6, 2004.

Aikawa K, Leggett RE, Levin RM. Effect of age on hydrogen peroxide mediated contraction damage in the male rat bladder. J Urol 170, 2082-5, 2003.

Baron JA. Smoking and estrogen-related disease. Am J Epidemiol 119, 9-22, 1984.
Bump RC, McClish DM. Cigarette smoking and pure genuine stress incontinence of urine: a comparison of risk factors and determinants between smokers and nonsmokers. Am J Obstet Gynecol 170, 79-82, 1994.

Brodsky IE, Medzhitov R. Pyroptosis: macrophage suicide exposes hidden invaders. Curr Biol. 21, 72-75, 2011.

Bump RC, Sugerman HJ, Fantl JA et al. Obesity and lower urinary-tract function in women – effect of surgically induced weight-loss. Am J Obstet Gynecol 167, 2-9, 1992.

Bulmer P, Yang Q, Abrams P. Does cigarette smoking cause detrusor instability in women? J Obstet Gynaecol 21, 8-9, 2001.

Bourquia A, Jabrane AJ, Ramdani B et al. Systemic to toxicity of paraphenylenediamine. Presse Medicale 17, 798, 1988.

Chien CT, Yu HJ, Lin TB et al. Neural mechanisms of impaired micturition reflex in rats with acute partial bladder outlet obstruction. Neuroscience 96, 221-30, 2000.

Chien CT, Shyue SK, Lai MK. Bcl-xL augmentation potentially reduces ischemia/reperfusion induced proximal and distal tubular apoptosis and autophagy. Transplantation 84, 1183-90, 2007.

Chien CT, Yu HJ, Lin TB et al. Substance P via NK1 receptor facilitates hyperactive bladder afferent signaling via action of ROS. Am J Physiol Renal Physiol 284, F840-51, 2003.

Chien WC, Hayakawa S, Shimizu K et al. Catechins prevents substance P-induced hyperactive bladder in rats via the downregulation of ICAM and ROS. Neurosci Lett 367, 213-7, 2004.

Chen Y, Gibson SB: Is mitochondrial generation of reactive oxygen species a trigger for autophagy? Autophagy 4, 246-8, 2008.

Chung SD, Lai TY, Chien CT et al. Activating Nrf-2 Signaling Depresses Unilateral Ureteral Obstruction-Evoked Mitochondrial Stress-Related Autophagy, Apoptosis and Pyroptosis in Kidney. PLoSone 7, e47299, 2012.

Comhair SA, Erzurum SC. Antioxidant responses to oxidant-mediated lung diseases. Am J Physiol Lung Cell Mol Physiol 283, L246-55, 2002.

Caterina MJ, Schumacher MA, Tominaga M et al. The capsaicin receptor: a heat-activated ion channel in the pain pathway. Nature 389, 816-24, 1997.

Dallosso HM, McGrother CW, Matthews RJ et al. The association of diet and other lifestyle factors with overactive bladder and stress incontinence: a longitudinal study in women. Public Health Nutrition 7, 885-891, 2004.

de Jongh R, Haenen GR, van Koeveringe GA et al. Oxidative stress reduces the muscarinic receptor function in the urinary bladder. Neurourol Urodyn 26, 302-8, 2007.

de Groat WC. A neurologic basis for the overactive bladder. Urology 50, 36-52, 1997.

Dibbert B, Weber M, Wilfried H et al. Cytokinemediated Bax deficiency and consequent delayed neutrophil apoptosis: A general mechanism to accumulate effector cells in inflammation. Proc Natl Acad Sci USA 96, 13330-13335, 1999.

El-Ansay EH, Ahmed MEK, Clague HW. Systemic toxicity of paraphenylenediamine. Lancet 1, 1983.

Elbadawi A, Yalla SV, Resnick NM. Structural basis of geriatric voiding dysfunction. III. Detrusor overactivity. Urology 150, 1668-1680, 1993.

Erdem E, Leggett R, Dicks B et al. Effects of bladder ischaemia/
reperfusion on superoxide dismutase activity and contraction. BJU Int 96, 169-74, 2005.

Franco I. Overactive Bladder in Children. Part 1: Pathophysiology. Urology 178, 761-768, 2007.

Fridovich I. Superoxide Dismutases. Ann Biochem. 44, 147-159, 1975.

Fowler CJ, Griffiths D, de Groat WC. The neural control of micturition. Nat Rev Neurosci. 9(6), 453-66, 2008.

Lowry O H, Rosebrough N J, Farr A L. Protein measurement
with the Folin phenol reagent. J. Biol. Chem. 193, 265, 1951.

Gabow PA, Kachny WD, Kelleher SP. The spectrum of rhabdomyolysis. Medicine 61, 14-52, 1982.

Gross A, Jockel J, Wei MC. Enforced dimerization of BAX results in its translocation, mitochondrial dysfunction and apoptosis. EMBO J 17, 3878-85, 1998.

Ha JH, Noh HS, Shin IW et al. Mitigation of H2O2-induced autophagic cell death by propofol in H9c2 cardiomyocytes. Cell Biol Toxicol 28, 19-29, 2012.

Haferkamp A, Elbadawi A. Ultrastructural changes in the aging bladder. Der Urology 43, 527-534, 2004.

Herzog AR, Diokno AC, Brown MB. Two-year incidence, remission, and change patterns of urinary incontinence in non-institutionalized older adults. J Gerontol 4,67-74, 1990.

Hisayama T, Shinkai M, Takayanagi I et al. Mechanism of action of nicotine in isolated urinary-bladder of guinea-pig. Br J Pharmacol 95, 65-72, 1988.

Isaka Y, Suzuki C, Abe T et al. Bcl-2 protects tubular epithelial cells from ischemia/reperfusion injury by dual mechanisms. Transplant Proc 41, 52-4, 2009.

Juszczak K, Królczyk G, Filipek M et al. Animal models of overactive bladder: cyclophosphamide (CYP)-induced cystitis in rats. Folia Med Cracov 48, 13-23, 2007.

Kathiria AS, Butcher LD, Feagins LA et al. Prohibitin 1 modulates mitochondrial stress-related autophagy in human colonic epithelial cells. PLoS One 7, e31231, 2012.

Kaplan SA, Roehrborn CG, Rovner ES et al. Tolterodine and Tamsulosin for Treatment of Men With Lower Urinary Tract Symptoms and Overactive Bladder. JAMA 296, 2319-2328, 2006.

Kong S, Davison AJ. The relative effectiveness of .OH, H2O2, O2-, and reducing free radicals in causing damage to biomembranes. A study of radiation damage to erythrocyte ghosts using selective free radical scavengers. Biochim Biophys Acta. 640(1), 313-25, 1981.

Koskimaki J, Hakama M, Huhtala H et al. Association of dietary elements and lower urinary tract symptoms. Scand J Urol Nephrol 34, 46-50, 2000.

Lin BR, Yu CJ, Chen WC et al. Green tea extract supplement reduces D-galactosamine-induced acute liver injury by inhibition of apoptotic and proinflammatory signaling. Biomed Sci. 16, 35, 2009.

Lin AT, Yang CH, Chen KK et al. Detrusor mitochondrial lipid peroxidation and superoxide dismutase activity in partial bladder outlet obstruction of rabbits. Neurourol Urodyn 24, 282-7, 2005.

Lynes WL, Flynn SD, Shortliffe LD et al. Mast cell involvement in interstitial cystitis. J Urol.138, 746-52,1987.

MacPhee DG, Podger DM. Hair dyes (Correspondence). Med
J Aust 2, 33, 1975.

Maggi CA, Giuliani S, Conte B et al. Prostanoids modulate reflex micturition by acting through capsaicin-sensitive afferents. Eur J Pharmacol 145, 105-12, 1998.
Masuda H, Kihara K, Saito K et al. Reactive oxygen species mediate detrusor overactivity via sensitization of afferent pathway in the bladder of anaesthetized rats. BJU International 101, 775-780, 2007.

Møller LA, Lose G, Jørgensen T. Incidence and remission rates of lower urinary tract symptoms at one year in women aged 40–60: longitudinal study. Br Med J 320, 29-32, 2000.

Miao EA, Rajan JV, Aderem A. Caspase-1-induced pyroptotic cell death. Immunol Rev 243, 206-14, 2011.

Miller AF. Superoxide dismutases: active sites that save,but a protein that kills. Curr Opin Chem. Biol. 8, 162-168, 2004.

Milsom I, Abrams P, Cardozo L et al. How widespread are the symptoms of an overactive bladder and how are they managed? A population-based prevalence study. BJU International 87, 760-766, 2001.

Nott HW. Systemic poisoning by hair dye. The Br Med J 1, 1-2, 1924.

Orrenius S, Nicotera P, Zhivotovsky B. Cell death mechanisms and their implications in toxicology. Toxicol Sci 119, 3-19, 2011.

Pascual C, Castillo MD, Romay C. A New Luminol Sensitized Chemiluminescence Method for Determination of Superoxide Dismutase. Analytical Letters 25(5), 837-849, 2006.

Parekh MH, Lobel RO, O'Connor LJ et al. Protective effect of Vitamin E on the response of the rabbit bladder to partial outlet obstruction. J Urol 166, 341-6, 2001.

Parkin J, Shea C, Sant GR. Intravesical dimethyl sulfoxide (DMSO) for interstitial cystitis--a practical approach. Urology 49, 105-7, 1997.

Perry S, Shaw C, Assassa P et al. An epidemiological study to establish the prevalence of urinary symptoms and felt need in the community: the Leicestershire MRC Incontinence Study. J Public Health Med 22, 27-34, 2000.

Qian H, Yang Y, Wang X. Curcumin enhanced adriamycin-induced human liver-derived Hepatoma G2 cell death through activation of mitochondria-mediated apoptosis and autophagy. Eur J Pharm Sci 43, 125-131, 2011.

Ruan T, Lin YS, Lin KS et al. Sensory transduction of pulmonary reactive oxygen species by capsaicin-sensitive vagal afferents fibers in rats. J Physiol 565, 563-78, 2005.

Sant GR, Kempuraj D, Marchand JE et al. The Mast Cell in Interstitial Cystitis: Role in Pathophysiology and Pathogenesis. Urology 69, 34-40, 2007.

Scandalio JG. Oxygen stress and superoxide dismutases. Plant Physiol 101, 7-12, 1993.

Schroder K, Tschopp J. The Inflammasomes. Cell 140, 821-832, 2010.

Shafik A, Shafik IA. Overactive bladder inhibition in response to pelvic floor muscle exercises. World Urology 20, 374-377, 2003.

Simon HU, Haj-Yehia A, Levi-Schaffer F. Role of reactive oxygen species (ROS) in apoptosis induction. Apoptosis 5, 415-418, 2000.

Simon HU, Yousefi S, Schranz C et al. Direct demonstration of delayed eosinophil apoptosis as a mechanism causing tissue eosinophilia. J Immunol 158, 3902-3908, 1997.

Singla S, Miglani S, Lai AK et al. Para-phenylenediamine (PPD) Poisoning. JIACM 6, 236-8, 2005.

Smith JA. Neutrophils, host defense, and inflammation: A double-edged sword. J Leukoc Biol 56, 672-686, 1994.

Sontag JM. Carcinogenicity of substituted-benzenediamines
(phenylenediamines) in rats and mice. J. National Cancer Institute,
66, 591-602, 1981.

Sood AK, Yadav SP, Sood S. Hair dye poisoning. Jassoc Physicians Ind 44, 69, 1996.

Stewart WF, Van Rooyen JB, Cundiff GW et al. Prevalence and burden of overactive bladder in the United States. World Journal of Urology 20, 327-336, 2003.

Subak LL, Johnson C, Whitcomb E et al. Does weight loss improve incontinence in moderately obese women? Int Urogynecol J Pelvic Floor Dysfunct 13, 0-3, 2002.

Suliman SM, Homeida M, Aboud OI. Paraphenylenediamine induced acute tubular necrosis following hair dye ingestion. Human Toxicol 2, 3-5, 1983.

Thun MJ, Altekruse SF, Namboodiri MM et al. Hair dye use and risk of fatal cancers in U.S. women. J Natl Cancer Inst. 86(3), 210-5, 1994.

Yu HJ, Chien CT, Lai YJ et al. Hypoxia preconditioning attenuates bladder overdistension-induced oxidative injury by up-regulation of Bcl-2 in the rat. Physiol. 554, 815-828, 2004.

Yagi H, Hind AM, Khalil SI. Acute poisoning from hair dye. East Africa Med J. 68, 4-11, 1991.

Yeh CH, Chiang HS, Lai TY et al. Unilateral ureteral obstruction evokes renal tubular apoptosis via the enhanced oxidative stress and endoplasmic reticulum stress in the rat. Neurourol Urodyn 30, 472-9, 2011.