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From the * Department of Urology, Affiliated Drum
Tower Hospital, Nanjing University, School of Medicine, Nanjing, Jiangsu,
China; and the Department of Urology,
University of Texas Medical School at Houston and MD Anderson Cancer Center,
Houston, Texas.
| Correspondence to: Yutian Dai, Department of Urology, Gulou Hospital, Nanjing, Jiangsu 210008, China (e-mail: ytdai{at}hotmail.com), and Run Wang, Department of Urology, University of Texas Medical School at Houston and MD Anderson Cancer Center, Houston, TX 77030 (e-mail: run.wang{at}uth.tmc.edu). |
| Received for publication December 16, 2007; accepted for publication April 9, 2008. |
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Abstract |
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 Top Abstract References |
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Key words: Diabetes mellitus, erectile dysfunction, NO/cGMP pathway
Pathophysiological mechanisms underlying diabetes-associated ED are largely due to endothelial dysfunction, which could lead to the inability of the endothelium to produce vasorelaxing messengers and to maintain vasodilation and vascular homeostasis (Musicki and Burnett, 2007). Nitric oxide (NO) is one of the most important vasorelaxing messengers in the process of erection. It mediates the formation of cyclic guanosine monophosphate (cGMP), which is degraded by phosphodiesterase-5 (PDE5; Bredt and Snyder, 1994). The increased cGMP level will lead to the activation of protein kinases A and G, which subsequently phosphorylate the actin-myosin system proteins and Ca2+ channels located within the cellular membrane and sarcoplasmatic reticulum membrane. These processes will cause a reduction of free cytoplasmatic Ca2+ and finally the relaxation of smooth muscle, thereby triggering an erection. Therefore, the NO-cGMP pathway plays a key role in penile erection. Previous studies have demonstrated that in diabetes mellitus, the NO-cGMP pathway is adversely affected, which correspondingly impairs the relaxation of corpus cavernosum (Saenz de Tejada et al, 1989). One acceptable therapeutic strategy for the diabetic ED is to preserve or enhance the NO-cGMP pathway. Now, the most effective oral drugs are the inhibitors of PDE5, which inhibit the breakdown of intracellular cGMP and subsequently help achieve and maintain erection.
Caffeine is a common component in our everyday diet. It is a nonselective inhibitor of phosphodiesterases (Corbin and Francis, 2002), which can elevate the level of intracellular cGMP and cyclic adenosine monophosphate (cAMP). Caffeine could also decrease the influx of Ca2+ and reduce the Ca2+ sensitivity of contractile apparatus in some smooth muscles (Watanabe et al, 1992). These pharmacological effects lead to the relaxation of cavernous smooth muscles and decrease the contractile effects of cavernous muscle strips induced by high K+ solution, noradrenaline, and transmural electrical stimulation (Adebiyi and Adaikan, 2004). Recent studies suggest that long-term consumption of caffeine could improve the glucose homeostasis and the endothelial function in diabetic patients (Lopez-Garcia et al, 2006; Park et al, 2007). Thus, caffeine might enhance erectile function in these patients. However, current available epidemiology surveys were inconclusive regarding the effect of caffeine on erectile function. Some studies showed that caffeine had a positive effect on erectile function (Diokno et al, 1990; Akkus et al, 2002) and others indicated that caffeine had no effect in the general population (Berrada et al, 2003; Nicolosi et al, 2003). No epidemiology study is available to evaluate the effect of caffeine on ED in diabetic patients. To our knowledge, the effect of caffeine on ED in experimentally induced diabetic rats also has not been reported.
This study was designed to investigate the effect of caffeine on erectile function and to explore the effect of caffeine on cavernosus NO/cGMP pathway in diabetic rats.
Methods
Animal and Treatment—
Forty-eight male Sprague-Dawley rats weighing 200–250 g were obtained
from Shanghai Slac Laboratory Animal Co Ltd (Shanghai, China). Forty of them
were selected randomly and injected intraperitoneally with freshly prepared
streptozotocin (STZ; 65 mg/kg). The remainder were treated with vehicle (0.1
mol/L citrate-phosphate buffer, pH 4.5) as normal controls (group A). All rats
were kept in a temperature-controlled, air conditioned animal house with a
12-h light-dark cycle and were given free access to food and water. Procedures
were performed according to the recommendations of the institutional animal
care committee.
Seventy-two hours after the rats were injected with STZ, the blood glucose level in each rat was monitored at regular intervals throughout the study and immediately before sacrifice. Of the 40 STZ-induced rats, those having blood glucose levels higher than 300 mg/dL (16.6 mmol/L) were selected for the study. They were randomly divided into 3 groups and given different treatment regimens intragastrically for 8 weeks: group B (n = 8), diabetic controls, diabetic rats treated with normal saline; group C (n = 9), low–caffeine dose group, diabetic rats given caffeine with 10 mg/kg per day; group D (n = 9), moderate–caffeine dose group, diabetic rats given caffeine with 20 mg/kg per day.
After 8 weeks of treatment, intracavernous pressure (ICP), mean systemic arterial pressure (MAP), and the ratio of peak ICP to MAP (ICP/MAP ratio) were measured. The cavernosus cGMP was assessed by radioimmunoassay.
Erectile Function Evaluation— The measurement of ICP, calculation of the ICP/MAP ratio, and method of cavernous nerve stimulation were described previously (El-Sakka et al, 2001; Chen et al, 2007). Under anesthesia with urethane (1.6 g/kg intraperitoneally), each rat was placed on a heating pad to maintain body temperature. A PE-50 tube was inserted into the left carotid artery via a midline neck incision to record the MAP. Through a lower abdominal midline incision, the major pelvic ganglions, posterolateral to the prostate, were located. The pelvic nerves and the cavernous nerves were exposed and identified. The skin overlying the penis was incised and both penile crura were exposed by removing part of the overlying ischiocavernous muscle. A 23G needle connected to a PE-50 tube with heparinized saline (250 IU/mL) was carefully inserted into the right crus. The other end of the PE-50 tube was connected to a pressure sensor for measuring the intracavernous pressure. Electrostimulation was performed with a stainless steel bipolar hook electrode (each pole 0.2 mm in diameter; the 2 poles separated by 1 mm). The stimulus parameters were: current 5 mA, frequency 20 Hz, pulse width 0.2 ms, and duration 50 s. Pressure was measured and recorded with a Windows computer programmed multiplying channel physiograph and analyzed with RM6240B/C multi-channel biosignal collection processing system (Chengdu Implement Company, Chengdu, China).
Determine the Level of Cavernous cGMP—
The rat penis was amputated at the scarification. The corpora cavernosa
weighing 
50 mg were immediately frozen in liquid nitrogen after the
corpus spongiosum was removed. The frozen sample was ground into powder and
homogenized in 2 ml of cold sodium acetate buffer (50 mM, pH 4.75). cGMP was
extracted by the following method. Each homogenized penis sample was mixed
with 2 mL dehydrated alcohol followed by centrifugation at 3000 x
g for 15 min at 4°C, and the supernatant was collected. The
remaining sediment was washed again with 75% ethanol and centrifuged, and the
ethanolic phase was kept. These 2 supernatants were mixed and dried at
60°C (Seidler et al,
2002). Following redissolution, aliquots of the samples were
assayed for cGMP by radioimmunoassay with an RIA kit (Shanghai University of
Traditional Chinese Medicine, Shanghai, China) as per the manufacturer's
manual. Each sample was tested in duplicate. The protein content of the
pellets remaining after centrifugation was measured with BCA Protein Assay Kit
(Pierce, Rockford, Illinois). The results were expressed in picomoles of cGMP
per microgram protein (mean ± SD).
Statistical Assay— All data were expressed as mean ± SD. To compare among multiple groups, 1-way analysis of variance (ANOVA) was used. The least significant difference test (LSD) was employed for comparison between each 2 groups. All statistical analyses were processed through the Statistical Package for the Social Sciences (SPSS version 13.0 for Windows; SPSS Inc, Chicago, Illinois). P < .05 was considered statistically significant.
Results
General Data—
Of 40 STZ-induced rats, 26 had blood glucose higher than 300 mg/dL with
significantly increased food and water intakes, hyperuresis, and weight loss
compared with the normal controls. During this period, 2 rats in group B and 1
in group D died of complications (anorexia and infection). Body weight and
blood glucose level of the rats are shown in
Table 1.
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ICP and ICP/MAP Ratio— The results of ICP and ICP/MAP ratio with and without the administration of caffeine are shown in the Table 2 and Figure 1. There were no statistical differences of the baseline levels of ICP among all groups. With electrostimulation, significantly lower peak ICP values were observed in group B compared with group A (35.4 ± 9.1 vs 82.1 ± 5.1 mm Hg, P < .01). The peak ICP/MAP ratio was also significantly lower in group B compared with group A (0.33 ± 0.09 vs 0.76 ± 0.04, P < .01).
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In caffeine-treated rats, the peak ICP values were partially restored (49.6 ± 10.6 mm Hg for group C and 50.1 ± 11.4 mm Hg for group D) compared with normal saline-treated rats (35.4 ± 9.1 mm Hg for group B, P < .01). However, peak ICP values in caffeine-treated rats did not return to the normal level. The peak ICP/MAP ratio was also greater in caffeine-treated groups (0.47 ± 0.09 in group C and 0.46 ± 0.10 in group D) compared with the normal saline-treated rats (0.33 ± 0.09, P < .01) but was still less than that in the normal control rats. There were no significant differences of ICP and ICP/MAP ratio identified between 10 and 20 mg/kg per day caffeine-treated groups (group C vs group D, P > .05).
Cavernous cGMP— Table 3 showed the results of cavernous cGMP in each group. The cavernous cGMP decreased significantly in diabetic rats (0.079 ± 0.014 pmol/mg protein for group B, 0.103 ± 0.013 pmol/mg protein for group C, and 0.122 ± 0.026 pmol/mg protein for group D) when compared with normal control rats (0.209 ± 0.023 pmol/mg protein for group A, P < .01). However, when diabetic rats were treated with caffeine (group C and group D), the cGMP levels increased significantly compared with the diabetic rats treated with normal saline (group B, P < .05), even though the cGMP in caffeine-treated groups did not return to the normal level. No statistical differences of cGMP level were seen between 10 and 20 mg/kg per day caffeine-treated groups (Figure 2).
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Discussion
It is well known that penile erection depends on decreased penile vascular
resistance, which subsequently results in increased penile blood flow. The
action consists of the relaxation of the penile helicine arteries and the
cavernous smooth muscle that lines the cavernosal spaces
(Andersson and Wagner, 1995).
Research has demonstrated that reduced vasorelaxation or increased
vasoconstriction under diabetic conditions accounts for the development of
ED.
The NO-cGMP pathway plays an important role in initiating and maintaining an erection (Kandeel et al, 2001; Seidler et al, 2002). Previous studies have shown significant impairment of vasorelaxant effect of the NO-cGMP pathway in the diabetic condition, resulting in reduced erectile capability (Saenz de Tejada et al, 1989). In the STZ-induced diabetic rat models, a reduction of the ICP responses to nerve stimulation was observed 2–3 months after the STZ treatment (Rehman et al, 1997). In this study, we confirmed the measurable reductions of the peak ICP, the ICP/MAP ratio, and the level of cavernous cGMP at the 8-week mark in STZ-induced diabetic rats. These findings are similar to previously published results (Park et al, 2004; Bivalacqua et al, 2005).
On the basis of knowledge about the impaired NO-cGMP pathway in diabetes mellitus, PDE-5 inhibitors (sildenafil, vardenafil, tadalafil, and udenafil) are used to treat diabetic ED. The question is whether caffeine, as a nonselective PDE inhibitor used in our daily diet, is beneficial to penile erection in the diabetic condition. A study has shown that caffeine can increase the intracellular level of cGMP and cAMP and inhibit Ca2+ signaling (Moreland et al, 2001). It exhibits relaxant activities on various smooth muscles, including cavernous smooth muscle (Watanabe et al, 1992; Apaydin et al, 1998; Lindaman et al, 2002). Unfortunately, the role of caffeine on erectile function in epidemiology studies is controversial (Shiri et al, 2004). There is no study to evaluate the effect of caffeine for ED in diabetic patients. Therefore, study of the caffeine effect on erectile function in diabetic animal model can provide valuable information.
In our study, diabetic rats were treated with 2 doses of caffeine according to the 2000 report published by the Australia New Zealand Food Authority (ANZFA): low dose (10 mg/kg per day) and moderate dose (20 mg/kg per day) (Smith et al, 2000). The 10 mg/kg per day dosage in rats equates to 250 mg/person per day (Corbin and Francis, 2002), which is the average intake of adults every day. The 20 mg/kg dose is at the upper end of the moderate dose in rats. Most literature suggests the positive effects of caffeine, such as increased energy, alertness, and motivation, with the low to moderate dosages. The high levels of caffeine consumption (>500 mg/kg per day) were reported to have negative effects, such as increasing anxiety and impaired sleep, which are risk factors of ED (Sasaki et al, 2005; Quek et al, 2007). Therefore, the high dose was not included in our study.
Our study demonstrated the beneficial effect of caffeine given at the low and moderate dosages on erectile function in diabetic rats. The ICP and ICP/MAP ratio are well accepted parameters to assess erectile function in animal studies. Our data showed that an 8-week administration of caffeine could partially restore the erectile function as demonstrated by increased ICP and ICP/MAP ratio in diabetic rats, however, the caffeine given at the low and moderate dosages did not completely restore normal erection. We did not find a dose-dependent effect at the given low and moderate caffeine dosages. These dosages were used according to ANZFA's report, and the gap between the given dosages might be too narrow to demonstrate the difference in this animal model. It appears to be a drawback of our study that we did not include a high-dose treatment arm. Future study will incorporate the high-dose arm to elucidate the dose-dependent effect of caffeine on erectile function in this animal model.
Studies in gallbladder and aortic smooth muscles (Watanabe et al, 1992; Lindaman et al, 2002) demonstrate that the relaxant effect of caffeine on smooth muscles relates to the increased cGMP level. Because cGMP plays an important role in penile erection (Bredt and Snyder, 1994), we assumed that the cavernous cGMP levels in the diabetic rats with and without caffeine treatment might be different. Therefore, we measured the cavernous cGMP levels of the rats with radioimmunoassay to clarify the relationship between caffeine intake and the NO/cGMP pathway. Our study showed a significant up-regulation of cGMP level in the caffeine-treated groups compared with the diabetic controls. This finding can explain the mechanism of the beneficial effect of caffeine on erectile function in diabetic rats. Caffeine can also increase the level of cAMP in gallbladder, aortic, and bladder smooth muscle tissues (Watanabe et al, 1992; Sekiguchi et al, 2002; Yi et al, 2006). cAMP also can trigger vasodilation, resulting in erection. This possibility was not included in this study. In theory, the elevated cavernous cAMP by caffeine might also contribute to the restoration of erectile function in diabetic rats.
In this study, we demonstrated the beneficial effect of caffeine to enhance erectile function in diabetic rats by up-regulating cGMP in cavernous tissue. The etiology of diabetic ED is multifactorial (Moore and Wang, 2006). Despite the discovery of effective oral medications for ED, diabetic ED remains the medical condition most difficult to treat. At present, no single treatment strategy is effective for all diabetic ED cases. Combination therapies to target the multimolecular levels involved in diabetic ED are the future of managing this difficult condition (Chen et al, 2008). Caffeine, as shown in our study, cannot completely reverse ED in diabetic rats. However, as a daily diet, it might potentiate the effect of other erectile-genic medications, such as NO donators in the treatment of diabetic ED.
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Acknowledgments |
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References |
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TopAbstractReferences |
|---|
Akkus E, Kadioglu A, Esen A, Doran S, Ergen A, Anafarta K, Hattat H. Prevalence and correlates of erectile dysfunction in Turkey: a population-based study. Eur Urol. 2002; 41: 298 –304.[CrossRef][Medline]
Andersson KE, Wagner G. Physiology of penile erection.
Physiol Rev. 1995; 75: 191
–236.
Apaydin S, Gonen C, Guven H. The probable role of nitric oxide on the relaxations obtained by caffeine and aminophylline in rat uterus. Pharmacol Res. 1998; 38: 387 –392.[CrossRef][Medline]
Berrada S, Kadri N, Mechakra-Tahiri S, Nejjari C. Prevalence of erectile dysfunction and its correlates: a population-based study in Morocco. Int J Impot Res. 2003; 15(suppl 1): S3 –S7.
Bivalacqua TJ, Usta MF, Kendirci M, Pradhan L, Alvarez X, Champion HC, Kadowitz PJ, Hellstrom WJ. Superoxide anion production in the rat penis impairs erectile function in diabetes: influence of in vivo extracellular superoxide dismutase gene therapy. J Sex Med. 2005; 2: 187 –197.[CrossRef][Medline]
Blanco R, Saenz de Tejada I, Goldstein I, Krane RJ, Wotiz HH, Cohen RA. Dysfunctional penile cholinergic nerves in diabetic impotent men. J Urol. 1990;144: 278 –280.[Medline]
Bredt DS, Snyder SH. Nitric oxide: a physiologic messenger molecule. Annu Rev Biochem. 1994; 63: 175 –195.[CrossRef][Medline]
Chen Y, Dai Y, Wang R. Treatment strategies for diabetic patients suffering from erectile dysfunction. Expert Opin Pharmacother. 2008;9: 257 –266.[CrossRef][Medline]
Chen Y, Li SX, Yao LS, Wang R, Dai YT. Valsartan treatment reverses erectile dysfunction in diabetic rats. Int J Impot Res. 2007;19: 366 –370.[CrossRef][Medline]
Corbin JD, Francis SH. Pharmacology of phosphodiesterase-5 inhibitors. Int J Clin Pract. 2002; 56: 453 –459.[Medline]
Diokno AC, Brown MB, Herzog AR. Sexual function in the elderly.
Arch Intern Med. 1990; 150: 197
–200.
El-Sakka A, Yen TS, Lin CS, Lue TF. Traumatic arteriogenic erectile dysfunction: a rat model. Int J Impot Res. 2001; 13: 162 –171.[CrossRef][Medline]
Kandeel FR, Koussa VK, Swerdloff RS. Male sexual function and its
disorders: physiology, pathophysiology, clinical investigation, and treatment.
Endocr Rev. 2001; 22: 342
–388.
Lindaman BA, Hinkhouse MM, Conklin JL, Cullen JJ. The effect of phosphodiesterase inhibition on gallbladder motility in vitro. J Surg Res. 2002;105: 102 –108.[CrossRef][Medline]
Lopez-Garcia E, van Dam RM, Qi L, Hu FB. Coffee consumption and
markers of inflammation and endothelial dysfunction in healthy and diabetic
women. Am J Clin Nutr. 2006; 84: 888
–893.
Moore CR, Wang R. Pathophysiology and treatment of diabetic erectile dysfunction. Asian J Androl. 2006; 8: 675 –684.[CrossRef][Medline]
Moreland RB, Hsieh G, Nakane M, Brioni JD. The biochemical and
neurologic basis for the treatment of male erectile dysfunction. J
Pharmacol Exp Ther. 2001;296: 225
–234.
Musicki B, Burnett AL. Endothelial dysfunction in diabetic erectile dysfunction. Int J Impot Res. 2007; 19: 129 –138.[CrossRef][Medline]
Nicolosi A, Moreira ED Jr, Shirai M, Bin Mohd Tambi MI, Glasser DB. Epidemiology of erectile dysfunction in four countries: cross-national study of the prevalence and correlates of erectile dysfunction. Urology. 2003;61: 201 –206.[CrossRef][Medline]
Park CS, Ryu SD, Hwang SY. Elevation of intracavernous pressure and NO-cGMP activity by a new herbal formula in penile tissues of aged and diabetic rats. J Ethnopharmacol. 2004; 94: 85 –92.[CrossRef][Medline]
Park S, Jang JS, Hong SM. Long-term consumption of caffeine improves glucose homeostasis by enhancing insulinotropic action through islet insulin/insulin-like growth factor 1 signaling in diabetic rats. Metabolism. 2007; 56: 599 –607.[CrossRef][Medline]
Quek KF, Sallam AA, Ng CH, Chua CB. Prevalence of sexual problems and Its association with social, psychological and physical factors among men in a Malaysian population: a cross-sectional study. J Sex Med. 2008; 5(1): 70 –76.[Medline]
Rathmann W, Giani G. Global prevalence of diabetes: estimates for
the year 2000 and projections for 2030. Diabetes Care. 2004; 27: 2568
–2569.
Rehman J, Chenven E, Brink P, Peterson B, Walcott B, Wen YP, Melman A, Christ G. Diminished neurogenic but not pharmacological erections in the 2- to 3-month experimentally diabetic F-344 rat. Am J Physiol. 1997;272: H1960 –H1971.[Medline]
Richardson D, Vinik A. Etiology and treatment of erectile failure in diabetes mellitus. Curr Diab Rep. 2002; 2: 501 –509.[Medline]
Saenz de Tejada I, Goldstein I, Azadzoi K, Krane RJ, Cohen RA. Impaired neurogenic and endothelium-mediated relaxation of penile smooth muscle from diabetic men with impotence. N Engl J Med. 1989; 320: 1025 –1030.[Abstract]
Sasaki H, Yamasaki H, Ogawa K, Nanjo K, Kawamori R, Iwamoto Y, Katayama S, Shirai M. Prevalence and risk factors for erectile dysfunction in Japanese diabetics. Diabetes Res Clin Pract. 2005; 70: 81 –89.[CrossRef][Medline]
Seidler M, Uckert S, Waldkirch E, Stief CG, Oelke M, Tsikas D, Sohn M, Jonas U. In vitro effects of a novel class of nitric oxide (NO) donating compounds on isolated human erectile tissue. Eur Urol. 2002; 42: 523 –528.[CrossRef][Medline]
Sekiguchi F, Miyake Y, Kashimoto T, Sunano S. Unaltered caffeine-induced relaxation in the aorta of stroke-prone spontaneously hypertensive rats (SHRSP). J Smooth Muscle Res. 2002; 38: 11 –22.[CrossRef][Medline]
Shiri R, Koskimaki J, Hakama M, Hakkinen J, Huhtala H, Tammela TL, Auvinen A. Effect of life-style factors on incidence of erectile dysfunction. Int J Impot Res. 2004; 16: 389 –394.[CrossRef][Medline]
Smith PF, Smith A, Miners J, McNeil JAP. Report From the Expert Working Group on the Safety Aspects of Dietary Caffeine. Canberra: Australian New Zealand Food Authority; 2000 .
Watanabe C, Yamamoto H, Hirano K, Kobayashi S, Kanaide H.
Mechanisms of caffeine-induced contraction and relaxation of rat aortic smooth
muscle. J Physiol. 1992; 456: 193
–213.
Yi CR, Wei ZQ, Deng XL, Sun ZY, Li XR, Tian CG. Effects of coffee and caffeine on bladder dysfunction in streptozotocin-induced diabetic rats. Acta Pharmacol Sin. 2006; 27: 1037 –1043.[CrossRef][Medline]
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