NGF, TrKA,p75NTR in Men With ED and Diabetes With or Without MS (ED)

Role and Clinical Relevance of Nerve Growth Factor (NGF) and Its Receptors (TrKA and p75NTR) in Patients With Erectile Dysfunction and Diabetes With or Wothout Metabolic Syndrome

Ester Illiano, MD, Mario Rende,Full Professor, Alessandra Psitilli, Researcher, Anna Stabile, Researcher

To evaluate the level of NGF,TrKA and p75NTR in patientd with erectile dysfunction and diabetes with or without metabolic syndrome

Penile erection is a complex phenomenon which implies a delicate and co-ordinated equilibrium among the neurological, vascular and the smooth muscle compartment. It includes arterial dilation, trabecular smooth muscle relaxation and activation of the corporeal veno-occlusive mechanism. Erectile Dysfunction (ED) is defined as the persistent inability to attain and maintain an erection sufficient to permit satisfactory sexual performance. Erectile Dysfunction may affect physical and psychosocial health and may have a significant impact on the quality of life (QoL) of sufferers and their partner's . There is increasing evidence that ED can be an early manifestation of coronary artery and peripheral vascular disease. Erectile Dysfunction should not be regarded only as a QoL issue, but also as a potential warning sign of cardiovascular disease (CVD) .The most important organic causes of ED are vascular, metabolic, neurogenic, hormonal and pharmacological. 35-90% of diabetic patients suffer from ED In the latter metabolic disease, Dai et al. have identified as a possible etiology of ED a lack of interaction between nerve growth factor (NGF) and its TrKA receptor, resulting in a noticeable increase in NGF levels in the corpora cavernosa11. Furthermore, Hou et al. have shown that the activation of the NGF / TrKA system in the corpora cavernosa (increased expression of NGF and TrKA mRNA and upregulation of c-raf, ERK1 / 2 and CREB1) improves erectile function in mice with ED and diabetes mellitus. The objectives of this study are: The primary outcome is to evaluate the systemic plasma concentration and the concentration in cavernous bodies of penis of NGF in patients with ED and diabetes with or without metabolic syndrome (a); (b) the expression of its TrKA and p75NTR receptors in the mononuclear cells taken from peripheral blood and in cavernous bodies of penis. The secondary outcome is to evaluate whether the levels of NGF and its receptors in the mononuclear cells can correlate with the clinical staging of ED. Prospective pilot study. 12 patients diagnosed with ED with diabetes with or without metabolic syndrome( Mets )aged between 18 and 65 will be recruited. Each patient will undergo an initial evaluation that includes: the pathological and pharmacological anamnesis, the local objective examination and the compilation of the validated questionnaire International Index of Erectile Function Questionnaire short form (IIEF5). Based on the IIEF5 score, patients will be divided into 4 groups (each of 10 patients): mild ED (score 17-21) mild to moderate ED (score 12-16) Moderate ED (score 8-11) Severe ED (score 5-7) In vivo study: Each patient will then be submitted in succession to: A collection of peripheral blood from the routinary cubital vein for hormone dosage Follicle-stimulating hormone (FSH), Luteinizing hormone (LH), TESTOSTERONE FREE AND TOTAL, ESTRADIOL, PROLACTIN) and metabolic evaluation (total HDL cholesterol, LDH, glycemia, lipidemic framework); and an aliquot will be used for in vitro study An introverted cavernous infiltration of Prostaglandin E1 to achieve erection (Aprostadil); A blood sample from both the corpus cavernosum and the cubital vein, once the erection is achieved Vitro study To verify a possible baseline interaction of prostaglandin E1 on the concentration of NGF released in the medium and on the expression of its receptors, an in vitro study will be performed using mononuclear cells isolated from peripheral blood (PBMC) obtained from buffy coat treated with prostaglandin E1 Evaluation of NGF and Cytokine levels A portion of each blood sample and medium conditioned by PBMC will be used for the enzyme immunoassay of NGF and pro and anti-inflammatory cytokine levels (TNF alfa and IL-10). Expression analysis of TrKA and p75NTR receptors and intracellular cytokines in PBMCs. The remaining part of each blood sample and PBMC of the in vitro experiment will be used for the cytofluorimetric evaluation of the expression of TrKA, p75NTR and the pro and anti-inflammatory cytokines and reactive oxygen species Evaluation of the levels of NGF and its receptors in the different degrees of severity of the EDThe levels of NGF and its receptors will be stratified into the 4 groups of clinical severity of the ED In vitro study An in vitro study will be performed using PBMC obtained from buffy coat of healthy subjects treated with prostaglandin E1. Statistical analysis of the results The data obtained during the study will be treated with appropriate statistical tests. The results will be expressed in terms of mean ± standard error and compared by appropriate statistical tests (ANOVA or non-parametric where appropriate). Expected results From the present study we expect to obtain more information on the impact and relevance of the NGF / TrKA / p75NTR pathway on DE in patients with Mets. Furthermore, we expect a correlation between the plasma levels of NGF and its receptors in relation to the clinical severity of the ED.

A peripheral blood sample from the cubital vein A collection of peripheral blood from the routinary cubital vein for hormone dosage and metabolic evaluation An introverted cavernous infiltration of Prostaglandin E1 to achieve erection (Aprostadil); A blood sample from both the corpus cavernosum and the cubital vein, once the erection is achieved Baseline interaction of prostaglandin E1 on the concentration of NGF released in the medium and on the expression of its receptors Evaluation of NGF and Cytokine levels Expression analysis of TrKA and p75NTR receptors and intracellular cytokines in PBMCs

Evaluation of NGF and Cytokine levels A portion of each blood sample and medium conditioned by PBMC will be used for the enzyme immunoassay of NGF and pro and anti-inflammatory cytokine levels (TNF alfa and IL-10). Expression analysis of TrKA and p75NTR receptors and intracellular cytokines in PBMCs The remaining part of each blood sample and PBMC of the in vitro experiment will be used for the cytofluorimetric evaluation of the expression of TrKA, p75NTR and the pro and anti-inflammatory cytokines and reactive oxygen species.

to evaluate any differences between the data obtained from the systemic levy versus the intracavernous sampling.

is to evaluate if the levels of NGF and its receptors in the mononuclear cells can correlate with the clinical staging of ED.

Inclusion Criteria: men with DE and metabolic syndrome Exclusion Criteria: men with DE without metabolic syndrome

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