Human papillomavirus types distribution in eastern Sicilian females with cervical lesions. A correlation with colposcopic and histological findings.

To determine human papillomavirus (HPV) types distribution in cervical lesions in a Southern Italian female population in Messina and their relationship between HPV type and grade of colposcopic and histopathological abnormality, a total of 253 women aged 17-68 years, with previous cytological abnormalities, were included in this study. HPV-DNA testing, colposcopy and biopsy were performed. For each sample, cervical cells were collected by centrifugation and DNA was extracted, followed by a PCR-based HPV-DNA assay and reverse dot blot genotyping. HPV-16 was found the most common type (46.6 %) followed by HPV-31 (26.9 %), -6 (18.6 %), -58 (8.8 %), -18 (6.7 %), -66 (5.7 %), -52 and -53 (4.7 %). Out of 62 women with abnormal transformation zone (ATZ) area compatible with squamous intraepithelial lesion (SIL) or cervical cancer (CC), 64.5 % was found high risk (HR) HPV-positive. Moreover the severity of the colposcopic diagnosis was positively correlated with the higher HPV oncogenicity risk (HPV-16 P = 0.023; and HPV-53 P = 0.047). The HPV-16 was found the most prevalent type within each histological category: 66.7 %, 31.2 %, 44 % and 37.2 % of CC, high grade (H)SIL, low grade (L)SIL and chronic cervicitis respectively; followed by HPV-31 present in 25 %, 8 %, and 13.3 % of HSIL, LSIL and chronic cervicitis respectively. A higher HPV incidence than the rest of Italy was found, in agreement with that detected by other authors for the South of the country. These data provide further information about the types prevalence in women with cervical lesions living in Eastern Sicily, suggesting the introduction of new targeted vaccines against a wider spectrum of HPV.

Pacemakers in the upper urinary tract.

BACKGROUND: Pacemakers in upper urinary tract (UUT) are still under study. AIM: We reviewed the role of some cells that seem to be involved in the propulsion of urinary bolus from UUT to the bladder. MATERIALS & METHODS: We focuses on evaluating studies on the mechanisms by which the UUT propels urine to the bladder via pacemaker cells. RESULTS: Electric active pacemaker cells generate pyeloureteric autorhythmicity driving adjacent smooth muscle cells (SMCs); it emphasizes the role of the interstitial cells of Cajal-like cells (ICC-LCs) localized in the UUT. Interstitial cells of Cajal (ICCs) are now thought to cooperate in conducting and amplifying pacemaker activity in the UUT. These cells produce electrical slow-wave potentials and determine the propagation of peristaltic activity. Identification of ICC-LCs is facilitated by use of c-kit antibodies. Contraction waves arising from the UUT and the propagation of these waves may require the direct involvement of ICC-LCs, as c-kit immunoreactivity appears developmentally at the same time as coordinated unidirectional peristaltic contraction. ICC-LCs observed in the UUT have morphological features similar to those of c-kitpositive ICCs in the gastrointestinal tract. In addition to gastrointestinal motility, ICCs may also play a significant role in the propagation, coordination, and modulation of ureteropelvic peristalsis. DISCUSSION: Alterations in ICC-LCs are closely associated with a variety of motility disorders and many congenital urological diseases of the UUT such as primary obstructive megaureter, congenital ureteropelvic junction obstruction, and vesicoureteral reflux. CONCLUSION: These observations open the way for further investigations of this cell type.

Polydeoxyribonucleotide administration improves the intra-testicular vascularization in rat experimental varicocele.

OBJECTIVE: To study the effect of PDRN on angiogenesis in a model of varicocele in rats. DESIGN: After the creation of experimental varicocele, rats were randomized to one of the four treatments: vehicle, PDRN, DMPX, and PDRN plus DMPX. Twenty-one days after randomization, all animals were euthanized and the left testis was harvested. SETTING: Academic hospital. ANIMAL(S): Male Sprague-Dawley rats were used. INTERVENTION(S): A clamp was passed behind the left renal vein distally to the spermatic vein insertion. A silk ligature was placed around the left renal vein at this site and was tied over the top of a probe. The latter was then withdrawn and the vein was allowed to expand. In shams, a suture was placed but it was not tied. MAIN OUTCOME MEASURE(S): To assess testicular microvascular density using CD34 immunostaining. RESULT(S): Microvascular density in the varicocele plus PDRN group was significantly higher than in other groups. CONCLUSION(S): PDRN could represent a novel therapeutic strategy for varicocele treatment in subfertile patients, improving the innate pathophysiologic mechanism of neoangiogenesis, through compensatory oxygen and metabolite supply to tubular and extratubular testicular compartments.

Melanocortin 4 receptor activation protects against testicular ischemia-reperfusion injury by triggering the cholinergic antiinflammatory pathway.

Melanocortins (MC) trigger a vagus nerve-mediated cholinergic-antiinflammatory pathway projecting to the testis. We tested whether pharmacological activation of brain MC receptors might protect the testis from the damage induced by ischemia-reperfusion. Adult male rats were subjected to 1-h testicular ischemia, followed by 24-h reperfusion [testicular ischemia-reperfusion (TI/R)]. Before TI/R, groups of animals were subjected to bilateral cervical vagotomy, or pretreated with the nicotinic acetylcholine receptor antagonist chlorisondamine or the selective MC(4) receptor antagonist HS024. Immediately after reperfusion, rats were ip treated with saline or the MC analog [Nle(4),D-Phe(7)]α-melanocyte-stimulating hormone (NDP-α-MSH) (340 µg/kg). We evaluated testicular IL-6 and TNF-α by Western blot analysis and organ damage by light microscopy. Some experimental groups were prepared for neural efferent activity recording along the vagus nerve starting 30 min after treatment with NDP-α-MSH or saline, and for a 30-min period. Additional groups of TI/R rats were treated for 30 d with saline, NDP-α-MSH, chlorisondamine plus NDP-α-MSH, or HS024 plus NDP-α-MSH to evaluate spermatogenesis, organ damage, and the apoptosis machinery. After a 24-h reperfusion, in TI/R saline-treated rats, there was an increase in IL-6 and TNF-α expression and a marked damage in both testes. NDP-α-MSH inhibited IL-6 and TNF-α expression, decreased histological damage, and increased neural efferent activity. Furthermore, NDP-α-MSH administration for 30 d greatly improved spermatogenesis, reduced organ damage, and inhibited apoptosis. All positive NDP-α-MSH effects were abrogated by vagotomy, chlorisondamine, or HS024. Our data suggest that selective MC(4) receptor agonists might be therapeutic candidates for the management of testicular torsion.

Ileocystoplasty in rats: metabolic, renal and enteropatch changes in a mid- and long-term follow-up.

INTRODUCTION: Bladder augmentation predisposes humans to many metabolic, renal and enteropatch changes. Our aim was to evaluate in a rat model of ileocystoplasty mid- and long-term urinary, metabolic, renal and graft changes. MATERIALS AND METHODS: We performed an ileocystoplasty and a sham operation in 30 rats. Seven augmented rats and 3 sham-operated animals were euthanized after 1, 3 and 6 months. We performed urinalysis, urine culture and blood sampling for serum electrolytes and urea. Histopathological changes of the ileal patch and kidneys were also evaluated. RESULTS: The urine cultures were positive in 14.3, 57 and 71%, respectively, 1, 3 and 6 months after surgery. Urinary pH, serum chloride and urea of the augmented group were significantly higher. Bladder calculi were formed in 23.8% of ileocystoplasty. Histopathological examination showed urothelialization of the graft with hyperplastic/metaplastic changes. The kidneys showed glomerular depletion and a marked distal tubular ectasia. CONCLUSIONS: Our data showed that, in a mid- and long-term follow-up, rat bladders subjected to ileocystoplasty displayed hyperchloremic metabolic acidosis, electrolyte imbalance, enhanced serum blood urea levels and glomerular/tubular changes. Hyperplastic and/or metaplastic changes at the junctional zone were observed. Our experimental results suggest that frequent monitoring of renal function and surveillance of humans who have undergone ileocystoplasty are recommended.

Melanocortin 4 receptor stimulation decreases pancreatitis severity in rats by activation of the cholinergic anti-inflammatory pathway.

OBJECTIVE: Acute pancreatitis is an inflammatory condition that may lead to multisystemic organ failure. Melanocortin peptides have been successfully used in experimental models of organ failure and shock, and their protective effect occurs through the activation of a vagus nerve-mediated cholinergic anti-inflammatory pathway by acting at brain melanocortin 4 receptors. In the light of these observations, we studied the effects of the selective melanocortin 4 receptor agonist RO27-3225 in an experimental model of cerulein-induced pancreatitis. DESIGN: Randomized experiment. SETTING: Research laboratory at a university hospital. SUBJECT: Experimental pancreatitis in rats. INTERVENTIONS: Acute pancreatitis was induced in male Sprague-Dawley rats by intraperitoneal injections of cerulein (80 µg/kg, four injections at hourly intervals). Before pancreatitis induction, groups of animals were subjected to bilateral cervical vagotomy, pretreated with the nicotinic acetylcholine receptor antagonist chlorisondamine or the selective melanocortin 4 receptor antagonist HS024, or not pretreated. Thirty minutes after the first cerulein injection, rats were intraperitoneally treated with a nanomolar dose of RO27-3225 or vehicle. Some experimental groups were prepared for neural efferent activity recording along the vagus nerve starting 30 mins after treatment with RO27-3225 or vehicle, and for a 30-min period. MEASUREMENTS AND MAIN RESULTS: Serum lipase and amylase activity, tumor necrosis factor-α and interleukin-6 expression, pancreatic myeloperoxidase activity, and histologic damage were evaluated; neural efferent activity of vagal fibers was also assessed. RO27-3225 reduced cerulein-induced serum lipase and amylase activity, blunted the expression of tumor necrosis factor-α and interleukin-6, abated the increase in pancreatic myeloperoxidase activity, and protected against histologic damage. Furthermore, RO27-3225 markedly increased neural efferent activity along the vagus nerve. Vagotomy, chlorisondamine, and HS024 abated these protective effects of RO27-3225. CONCLUSIONS: Our data show that melanocortin 4 receptor agonists reduce pancreatitis severity through the activation of the cholinergic anti-inflammatory pathway. These findings could be of particular interest in the clinical setting.

Activation of adenosine A2A receptors by polydeoxyribonucleotide increases vascular endothelial growth factor and protects against testicular damage induced by experimental varicocele in rats.

In rat experimental varicocele, polydeoxyribonucleotide (PDRN) induces vascular endothelial growth factor (VEGF) production, thereby enhancing testicular function. This may point to a new therapeutic approach in human varicocele.

Genistein aglycone: a new therapeutic approach to reduce endometrial hyperplasia.

OBJECTIVE: Endometrial hyperplasia without cytological atypia is commonly treated with progestins, but other treatment regimes may be available with equivalent efficacy and low side effects. DESIGN: A randomized double-blind, placebo and progesterone-controlled clinical trial to evaluate the effects of genistein aglycone in reducing endometrial hyperplasia. PATIENTS: A group of 56 premenopausal women with non-atypical endometrial hyperplasia were enrolled and received: genistein aglycone (n=19; 54 mg/day); norethisterone acetate (n=19; 10 mg/day on days 16-25 of the menstrual cycle) or placebo (n=18) for 6 months. MEASUREMENTS: Hysteroscopy was performed with biopsies and symptomology assessed at baseline, 3 and 6 months of administration. The effect on estrogen (ER) and progesterone receptors (PR) expression in uterine biopsies were assessed after 3 and 6 months. For each treatment follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), sex hormone-binding globulin (SHBG) and progesterone (PG) levels were also evaluated. RESULTS: After 6 months, 42% of genistein aglycone-administered subjects had a significant improvement of symptoms (histologically confirmed in the 29%) compared to 47% of norethisterone acetate subjects (histologically confirmed in the 31%), but only 12% in the placebo group with 19% exhibiting worsening symptoms and increased endometrial thickness. No significant differences were noted for hormone levels for any treatment, but immunohistochemical analysis revealed significantly reduced staining for ER-alpha and PR and enhanced ER-beta1 staining in genistein-administered subjects associated with a complete regression of bleeding. CONCLUSIONS: These results suggest that genistein aglycone might be useful for the management of endometrial hyperplasia without atypia in women that cannot be treated with progestin.

Peroxisome proliferator activated receptor beta/delta activation prevents extracellular regulated kinase 1/2 phosphorylation and protects the testis from ischemia and reperfusion injury.

PURPOSE: Testicular torsion is a medical emergency that requires immediate diagnosis and treatment to avoid subsequent testicular injury and infertility. PPARs are a family of nuclear hormone receptors belonging to the steroid receptor superfamily. Three PPAR isotypes (alpha, beta/delta and gamma) encoded by separate genes and showing different tissue distribution patterns have been identified. PPARbeta/delta is expressed in testis and its role is largely unknown. We tested whether pharmacological activation of PPARbeta/delta might protect the testis from ischemia and reperfusion injury. MATERIALS AND METHODS: Adult male Sprague-Dawley rats were subjected to 1-hour testicular ischemia, followed by 24 hours of reperfusion. Sham testicular ischemia-reperfusion rats served as controls. The animals were randomized to receive immediately after detorsion 1) L-165,041 (4 mg/kg intraperitoneally), a potent agonist of PPARbeta/delta, 2) GW9662 (Calbiochem(R)) (4 mg/kg intraperitoneally), an antagonist of PPAR, 3) L-165,041 (4 mg/kg intraperitoneally) plus GW9662 (4 mg/kg intraperitoneally) concomitantly or 4) vehicle (1 ml/kg 10% dimethyl sulfoxide/NaCl solution). We evaluated testicular extracellular signal regulated kinase, tumor necrosis factor-alpha and interleukin-6 by Western blot. We also investigated PPARbeta/delta activation by Western blot, mRNA expression and organ damage. RESULTS: Testicular ischemia-reperfusion injury caused a significant increase in extracellular signal regulated kinase, tumor necrosis factor-alpha and interleukin-6 expression in each testis. Furthermore, histological examination revealed marked damage. L-165,041 administration increased the PPARbeta/delta message and protein, inhibited extracellular signal regulated kinase, tumor necrosis factor-alpha and interleukin-6 expression, and decreased histological damage. Concomitant administration of GW9662 reversed the protection exerted by PPARbeta/delta agonist. CONCLUSIONS: These findings indicate that PPARbeta/delta agonists might be an attractive therapeutic candidate for managing testicular torsion.

Mitogen-activated protein kinase 3/mitogen-activated protein kinase 1 activates apoptosis during testicular ischemia-reperfusion injury in a nuclear factor-kappaB-independent manner.

Nuclear factor kappa-B (NF-kappaB), mitogen-activated protein kinase3/MAPK1 and MAPK8 are involved in testicular ischemia reperfusion injury (testicular-I/R). NF-kappaB knock-out mice (KO) subjected to testicular-I/R have a reduced testicular damage, blunted MAPK8 activation and enhanced MAPK3/MAPK1 activity. To better understand the role of MAPK3/MAPK1 up-regulation during testicular-I/R, we investigated the effects of PD98059, an inhibitor of MAPK3/MAPK1, in KO mice during testicular-I/R. KO and wild-type (WT) animals underwent 1 h testicular ischemia followed by 24 h reperfusion or a sham testicular-I/R. Animals received either PD98059 (5 mg/kg/ip) or its vehicle. MAPK3/MAPK1, BAX, caspase-3 and -9 and TNF-alpha expression were assessed along with histological examination and an immunostaining for protein of apoptosis. Testicular-I/R caused a greater increase in MAPK3/MAPK1 in KO than in WT animals in both testes. KO mice had a lower expression of the apoptotic proteins and TNF-alpha as well as reduced histological damage compared to WT. Immunostaining confirmed the lower expression of BAX in the Leydig cells of KO mice. Administration of PD98059, abrogated MAPK3/MAPK1 expression and slightly reduced TNF-alpha but did not improve or reverse the histological damage in KO. PD98059 significantly reduced the histological damage in WT mice and markedly reduced the apoptotic proteins in KO and WT mice. These results suggest that testicular-I/R triggers also a pathway of organ damage involving MAPK3/MAPK1, TNF-alpha, BAX, caspase-3 and -9 that activates an apoptotic machinery in an NF-kappaB independent manner. These findings should contribute to better understand testicular torsion-induced damage.

Involvement of mitogen-activated protein kinases (MAPKs) during testicular ischemia-reperfusion injury in nuclear factor-kappaB knock-out mice.

Nuclear factor kappa-B (NF-kappaB), extracellular regulated kinase (ERK 1/2) and c-jun-N terminal kinase (JNK) play an important role in testicular ischemia. We investigated the patterns of ERK1/2, JNK and p38 activation in NF-kappaB knockout (KO) mice subjected to testicular torsion. KO and normal littermate wild-type (WT) animals underwent at 1 h testicular ischemia followed by 24 h reperfusion (TI/R). Sham testicular ischemia-reperfusion mice served as controls. ERK 1/2, JNK and p38 expression by western blot analysis, tumor necrosis factor-alpha (TNF-alpha) expression (RT-PCR and western blot analysis) and a complete histological examination were carried out. TI/R caused a greater increase in phosphorylated form of ERK 1/2 in KO mice than in WT animals in either the ischemic testis and the contralateral one. By contrary, active form of JNK and p38 were completely abrogated in both testes of KO mice, while WT animals showed a significant activation of those kinases in both testes. TNF-alpha expression was markedly reduced in KO mice when compared to WT mice either at the mRNA and the protein level. Finally TI/R-induced histological damage was markedly reduced in KO mice. Our data indicate that NF-kappaB plays a pivotal role in the development of testicular ischemia-reperfusion injury and suggest that, in the absence of the transcriptional factor, the up-stream signal JNK and p38 may be abrogated while ERK 1/2 activity is enhanced.

Lipid peroxidation activates mitogen-activated protein kinases in testicular ischemia-reperfusion injury.

PURPOSE: Testicular damage after torsion has been attributed to many mechanisms, of which one is lipid peroxidation of the plasma membrane, which could cause the activation of the mitogen-activated protein kinase family. These proteins are of vital importance for signal transduction pathways and 2 of them, extracellular signal-regulated kinase and c-jun N-terminal kinase, participate in the pathogenesis of testicular ischemia. We investigated whether lipid peroxidation may trigger mitogen-activated protein kinase activation in testicular ischemia-reperfusion. MATERIALS AND METHODS: Adult male Sprague-Dawley rats were subjected to 1-hour testicular ischemia, followed by 24 hours of reperfusion. Sham testicular ischemia-reperfusion rats served as controls. Animals were randomized to receive raxofelast, an inhibitor of lipid peroxidation (20 mg/kg intraperitoneally administered 15 minutes before detorsion and 15 minutes after detorsion) or vehicle (1 ml/kg 10% dimethyl sulfoxide/NaCl solution). A group of animals was sacrificed 0, 10, 15, 20, 25 and 30 minutes, and 1, 2 and 3 hours, respectively, after detorsion to evaluate testicular c-jun N-terminal kinase, extracellular signal-regulated kinase and tumor necrosis factor-alpha activation by Western blot analysis, and mRNA expression and conjugated dienes using a spectrophotometer technique. Another group was sacrificed 24 hours after detorsion to evaluate histological alterations. RESULTS: Testicular ischemia-reperfusion injury caused a significant increase in the conjugated diene levels, extracellular signal-regulated kinase c-jun N-terminal kinase activity and tumor necrosis factor-alpha expression in both testes. Furthermore, histological examination revealed marked damage. Raxofelast inhibited these parameters and decreased histological damage. CONCLUSIONS: These data suggest that lipid peroxidation triggers extracellular signal-regulated kinase and c-jun N-terminal kinase activation. Furthermore, mitogen-activated protein kinase blockade might represent a potential therapeutic approach to treatment in patients with unilateral testicular torsion.

Prenatal testicular torsion: ultrasonographic features, management and histopathological findings.

AIM: To highlight the ultrasonographic features of prenatal torsion of the testis in utero (IUTT) at presentation, the neonatal management and the histological findings postorchiectomy or biopsy. METHODS: Seven newborns underwent emergency exploration for IUTT. All patients underwent a sonography and real-time color Doppler ultrasound study of the scrotum before any surgical procedure. A histological examination was performed in the removed specimens. RESULTS: Sonography of the scrotum revealed enlarged, heterogeneous testes. In all cases the color and power Doppler did not reveal any flow signal on the affected side. Four newborn with unilateral testicular torsion underwent orchiectomy and contralateral orchidopexy. In one neonate after detorsion and with the absence of gangrenous changes and a reassuring biopsy, a twisted testis could be treated conservatively with orchidopexy. In another case, the parents, acknowledging the inviability of the affected testis, gave consent only for a biopsy of the testis. In the neonate with bilateral IUTT, bilateral testicular biopsies were performed. Histology of the removed testes variably showed interstitial red cell extravasion and coagulation or hemorrhagic necrosis. Light microscopy of the preserved testis highlighted surviving seminiferous tubules, with gonocytes, spermatogonia and fetal Sertoli cells. CONCLUSIONS: An early diagnosis and treatment in IUTT is essential. Surgical exploration should be always performed through the inguinal route. In bilateral IUTT testes should be left to try to assure, as long as possible, a residual Leydig cell function.

Evidence for a role of mitogen-activated protein kinase 3/mitogen-activated protein kinase in the development of testicular ischemia-reperfusion injury.

Mitogen-activated protein kinase (MAPK) 3/MAPK1 (also known as ERK1/ERK2) plays an important role in the signal transduction pathways. To our knowledge, however, its role in the development of testicular ischemia-reperfusion injury has not yet been investigated. Therefore, we studied the pattern of MAPK3/MAPK1 activation in a experimental model of testicular ischemia-reperfusion injury. We also investigated MAPK8 to understand whether an association exists between these two MAPKs. Adult male Sprague-Dawley rats were subjected to 1 h of testicular ischemia followed by 24 h of reperfusion or to a sham testicular ischemia-reperfusion. Animals were randomized to receive PD98059, which is an inhibitor of MAPK3/MAPK1 (10 mg/kg i.p. administered immediately after detorsion), or its vehicle. The time course of MAPK3/MAPK1, MAPK8, and tumor necrosis factor (TNF; also known as TNF alpha) expression and a histological examination in both the ischemic-reperfused testis and the contralateral one were performed. In both testes, MAPK3/MAPK1 and MAPK8 expression appeared following 10 min of reperfusion and reached their highest activation after 30 min. The MAPK levels slowly decreased, and no significant expression of either kinase was observed following 2 h of reperfusion. Expression of TNF was evident after 1 h of reperfusion and reached its maximum increase after 3 h. PD98059 blunted MAPK3/MAPK1 and MAPK8, reduced TNF expression, and improved the testicular damage caused by ischemia-reperfusion injury in both testes. These data emphasize that MAPK3/MAPK1 has a role in testicular damage and that its blockade might have a future therapeutic role for the management of patients with unilateral testicular torsion.

Immunohistology of aquaporin-1 and stem cell factor-receptor in human undescended testes.

OBJECTIVES: Both aquaporin (AQP) 1 and the stem-cell factor/C-kit system seem to have a definite role in testis function, but very few studies have been reported in humans, especially in the paediatric age group. With the present study we wanted to investigate the expression of these proteins to better delineate their role in normal and pathologic testes. METHODS: Immunohistology using AQP 1 and C-kit antibodies was performed on paraffin sections of open-testicular biopsies from 32 undescended testes. The testes of cryptorchid patients, with ages ranging from 2 to 15 years, were biopsied during an orchidopexy operation, after obtaining informed consent. Control biopsies, from 8 patients of matched age, were obtained during operations for inguinal hernia or hydrocele, always after obtaining informed consent. Positive results were recorded as diffuse or focal patterns and scored as weak, moderate or strong immunostaining. RESULTS: AQP 1 antibody strongly depicted microvessel endothelial cells, but was unlabeled in endotubular and interstitial cell lines, in both control and undescended testes. The C-kit immunostaining in normal testes revealed a diffuse, strong staining in the cytoplasm of spermatogonia and primary spermatocytes. However, in the undescended testes a focal C-kit immunolabelling was weakly recognized in both spermatogonial and immature Sertoli cells. CONCLUSIONS: These results indicate a direct involvement of AQP 1 in the regulation of fluid transport across the endothelial cell membranes of testicular microvessels. A role of the C-kit receptor protein is also substantiated by its strong expression in the maturing spermatogonia of the normal testes, but was minimally or not recognizable in undescended prepubertal testes.