Whole genome microarray of the major pelvic ganglion after cavernous nerve injury: new insights into molecular profile changes after nerve injury

What's known on the subject? and What does the study add? With the present study, we aimed to provide a global picture of the molecular processes that are activated by CN injury. The present study used genomic expression profiling to identify candidate genes that might be useful targets in the CN recovery process and, thus, the ultimate preservation of penile erection. Regeneration of the CN and axonal outgrowth clearly involve changes in multiple biochemical pathways that have never been investigated by microarray analysis. We analyzed global gene expression in the major pelvic ganglion at early stages (48 h and 14 days) after CN injury and focused on the detection of changes in genes related to nervous tissue repair and proliferation. The findings of the present study provide important insight into the molecular systems affected by CN injury and identify candidate genes that may be utilized for novel molecular‐based therapies for the preservation and protection of the CN during RP.

OBJECTIVES

• ? To to examine the complexity of the many molecular systems involved in supporting cavernous nerve (CN) repair and regeneration in a rat model of bilateral crush injury utilizing a microarray analysis approach. Erectile dysfunction (ED) is a common clinical complication after prostate cancer treatment by radical prostatectomy, and recovery of erectile function can take as long as 2 years.
• ? There are gaps in our understanding of the autonomic pelvic innervation of the penis that still need to be addressed for the development of an adequate treatment strategy for post‐prostatectomy ED. The molecular mechanisms of the intrinsic ability of CN to regenerate after an injury have not been elucidated.

MATERIALS AND METHODS

• ? We analyzed global gene expression in the major pelvic ganglion 48 h and 14 days after CN injury. Overall, a comparative analysis showed that 325 genes changed at the 48‐h time point and 114 genes changed at 14 days. There were 60 changed genes in common with both time points.
• ? Using the Ingenuity Pathway Analysis® system (Ingenuity Systems, Inc., Redwood City, CA, USA), we were able to analyze the significantly changed genes that were unique and common to each time point by biological function.
• ? We focused on the detection of changes related to nervous tissue repair and proliferation, molecular networks of neurotrophic factors, stem cell regulation and synaptic transmission.

RESULTS

• ? There was strong evidence of the early mobilization of genes involved in repair and neuroprotection mechanisms (SERPINF1, IGF1, PLAU/PLAUR, ARG1).
• ? Genes related to nervous system development (ATF3 GJA1, PLAU, SERPINE1), nerve regeneration (SERPINE2, IGF1, ATF3, ARG1) and synaptic transmission (GJC1, GAL) were changed.
• ? Several genes related to proliferation as well as apoptosis (A2M, ATF3, C3, EGR4, FN1, GJA1, GAL) were also changed, possibly as part of a protective mechanism or the initiation of remodelling.

CONCLUSIONS

• ? The results obtained show that multiple biological processes are associated with injury and repair of the CN and provide a systematic genome‐wide screen for neurotrophic and/or inhibitory pathways of nerve regeneration.
• ? These data identify the candidate genes that may be utilized in novel molecular‐based therapies for the preservation and protection of the CN during radical prostatectomy.

     

盆神经节海绵体内移植对双侧海绵体神经损伤后大鼠阴茎勃起功能的影响

目的　研究双侧海绵体神经离断后 ,自体盆神经节海绵体内移植对大鼠阴茎勃起功能的影响。方法　将 2 6只雄性SD大鼠 (3～ 6个月 ,30 0～ 4 0 0g 只 )随机分为 2组 :(1)实验组 (n =16 )行双侧海绵体神经离断及左盆神经节移植入左侧阴茎脚内 ;(2 )对照组 (n =10 )仅离断双侧海绵体神经。1个月及 3个月后阿朴吗啡皮下注射检测勃起功能 ,电刺激左阴茎脚移植区域或右侧盆神经节比较海绵体内压的变化 ,NADPH染色检测海绵体内一氧化氮合酶 (NOS)阳性神经纤维 ,透射电镜观察盆神经节移植后的存活情况。结果　两组大鼠 1个月及 3个月后注射阿朴吗啡后均无勃起反应 ;电刺激左阴茎脚盆神经节移植区域后亦不能导致勃起。 1个月后电刺激两组左阴茎脚后的海绵体内压变化分别为 (9 4 1± 3 2 0 )、(4 16± 2 5 8)cmH2 O(t=4 76 9,P <0 0 5 ) ,3个月后分别为 (13 6 7± 4 18)、(5 0 9± 2 74 )cmH2 O(t=9 18,P <0 0 5 )。两组大鼠左阴茎脚区域的NOS阳性神经纤维数目 1个月后分别为 (2 18 7± 2 4 5 )、(18 0± 3 7) (t=14 77,P <0 0 5 ) ,3个月后分别为 (183 2± 19 7)、(19 0± 3 8) (t =12 18,P <0 0 5 ) ;透射电镜证实了神经节移植后的存活。结论　盆神经节海绵体内移植后可以存     

Recruitment of intracavernously injected adipose-derived stem cells to the major pelvic ganglion improves erectile function in a rat model of cavernous nerve injury

Background

Intracavernous (IC) injection of stem cells has been shown to ameliorate cavernous-nerve (CN) injury-induced erectile dysfunction (ED). However, the mechanisms of action of adipose-derived stem cells (ADSC) remain unclear.

Objectives

To investigate the mechanism of action and fate of IC injected ADSC in a rat model of CN crush injury.

Design, setting, and participants

Sprague-Dawley rats (n = 110) were randomly divided into five groups. Thirty-five rats underwent sham surgery and IC injection of ADSC (n = 25) or vehicle (n = 10). Another 75 rats underwent bilateral CN crush injury and were treated with vehicle or ADSC injected either IC or in the dorsal penile perineural space. At 1, 3, 7 (n = 5), and 28 d (n = 10) postsurgery, penile tissues and major pelvic ganglia (MPG) were harvested for histology. ADSC were labeled with 5-ethynyl-2-deoxyuridine (EdU) before treatment. Rats in the 28-d groups were examined for erectile function prior to tissue harvest.

Measurements

IC pressure recording on CN electrostimulation, immunohistochemistry of the penis and the MPG, and number of EdU-positive (EdU+) cells in the injection site and the MPG.

Results and limitations

IC, but not perineural, injection of ADSC resulted in significantly improved erectile function. Significantly more EdU+ ADSC appeared in the MPG of animals with CN injury and IC injection of ADSC compared with those injected perineurally and those in the sham group. One day after crush injury, stromal cell-derived factor-1 (SDF-1) was upregulated in the MPG, providing an incentive for ADSC recruitment toward the MPG. Neuroregeneration was observed in the group that underwent IC injection of ADSC, and IC ADSC treatment had beneficial effects on the smooth muscle/collagen ratio in the corpus cavernosum.

Conclusions

CN injury upregulates SDF-1 expression in the MPG and thereby attracts intracavernously injected ADSC. At the MPG, ADSC exert neuroregenerative effects on the cell bodies of injured nerves, resulting in enhanced erectile response.     

Erythropoietin promotes the recovery of erectile function following cavernous nerve injury

PURPOSE: We investigated the effects of recombinant human (rh) erythropoietin (EPO) on erectile function recovery in a rat model of cavernous nerve (CN) injury. MATERIALS AND METHODS: Male rats underwent unilateral CN transection and excision of a 5 mm segment of the contralateral CN. One group received rhEPO (5,000 U/kg) subcutaneously daily for 14 days, while another received rhEPO 1 day and 1 hour prior to nerve injury. An additional group of animals was pretreated with 1 dose of darbepoetin (25 microg/kg). At 14 days following CN injury rats underwent erection physiology studies. Axonal regeneration was evaluated by electron microscopy. EPO receptor expression in the penis and major pelvic ganglion was evaluated immunohistochemically and by real-time polymerase chain reaction. RESULTS: Daily rhEPO effectively recovered erections after CN injury compared with saline treatment. Maximal intracavernous pressure area under the curve normalized to mean arterial pressure was significantly greater in EPO treated vs saline treated animals (p < 0.05). rhEPO and darbepoetin pretreatment was as effective as continuous 14-day therapy. EPO receptor expression was localized to neuronal cell bodies of the major pelvic ganglion, penile nerves and endothelial cells in the penis. Electron microscopy revealed significant improvement in axonal regeneration in rhEPO treated animals 14 days following injury compared to controls. CONCLUSIONS: EPO receptors are expressed in local neuronal and vascular tissues. Exogenous administration of rhEPO or darbepoetin in the setting of CN injury promotes erectile function recovery. This occurs through axonal regeneration of the injured nerve and possible penile protection.     

Bilateral peroneal nerve palsy following emergent stabilization of a pelvic ring injury

External noninvasive compressive devices are becoming popular for emergent stabilization of pelvic ring disruptions. The ease of application utilizing available materials such as sheets, the noninvasive nature of such measures, and perceived absence of complications has made this a popular stabilization modality. The authors report a case of bilateral peroneal nerve palsy related to the use of external compressive wraps in a patient with pelvic ring injury.     

大鼠坐骨神经损伤后感觉神经元死亡数量的研究

目的 研究周围神经损伤后,脊神经节感觉神经元的死亡数量。方法 先计算正常SD大鼠两侧的脊神经节感觉神经元胞体数量是否对称,再切断并原位吻合右侧大鼠坐骨神经,左侧不作任何处理,于术后不同时间取L4-L6脊神经节作苏木素-伊红(HE)染色,计算脊神经节感觉神经元胞体数量的变化。结果 正常SD大鼠两侧的脊神经节感觉神经元胞体数量呈对称分布;右侧坐骨神经损伤后,其脊神经节感觉神经元胞体数量较左侧减少:30 d后,脊神经节细胞的死亡比率达30％,神经元死亡高峰在神经损伤后的第2周内。结论大鼠坐骨神经损伤后,脊神经节感觉神经元的胞体有死亡,其死亡具有一定的特征。     

Anorectal injury following pelvic radiotherapy

BACKGROUND: Current knowledge of the effects of radiation on the anorectum is based on a limited number of studies. Variability in delivery techniques, both currently and historically, combined with a paucity of prospective and randomized studies makes interpretation of the literature difficult. This review presents the existing evidence and identifies areas that require further work. METHODS: This review is based on a literature search (Medline and PubMed) and manual cross-referencing. RESULTS AND CONCLUSION: More than three-quarters of patients receiving pelvic radiotherapy experience acute anorectal symptoms and up to one-fifth suffer from late-phase radiation proctitis. About 5 per cent develop other chronic complications, such as fistula, stricture and disabling faecal incontinence. The risk of rectal cancer may be increased. Conservative treatment options are of limited value. Surgery may be considered if symptoms are severe, provided sphincter function is adequate and recurrent disease is excluded. Large prospective studies with accurate dosimetric data and long-term follow-up are needed to provide meaningful information on which to base new strategies to minimize the side-effects from radiotherapy.     

Localization and expression of R-cadherin in skeletal muscle following nerve injury

Rats in which the sciatic nerves were cut were divided into two groups: animals with nerve sutured and animals with nerve not sutured. In the unsutured group, the levels of R-cadherin expression increased and then decreased to values lower than those of controls. In the sutured group, the levels of R-cadherin expression increased and then decreased to almost control values. These results suggest that R-cadherin plays some role in cells of normal and regenerating muscles.     

Accuracy of trauma ultrasound in major pelvic injury

BACKGROUND: Trauma ultrasound (US) utilizing the focused assessment with sonography in trauma (FAST) is often performed to detect traumatic free peritoneal fluid (FPF). Yet its accuracy is unclear in certain trauma subgroups such as those with major pelvic fractures whose emergent diagnostic and therapeutic needs are unique. We hypothesized that in patients with major pelvic injury (MPI) trauma ultrasound would perform with lower accuracy than has previously been reported. METHODS: Retrospective analysis of adult trauma patients with pelvic fractures seen at an urban Level I emergency department and trauma center. Patients were identified from the institutional trauma registry and ultrasound database from 1999 to 2003. All patients aged >16 years with MPI (Tile classification A2, all type B and C pelvic fractures, and type C acetabular fractures determined by a blinded orthopedic traumatologist) and who had a trauma US performed during the initial emergency department evaluation were included. All ultrasounds were performed by emergency physicians or surgeons using the four-quadrant FAST evaluation. Results of US were compared with one of three reference standards: abdominal/pelvic computed tomography, diagnostic peritoneal tap, or exploratory laparotomy. Two-by-two tables were constructed for diagnostic indices. RESULTS: In all, 96 patients were eligible; 9 were excluded for indeterminate ultrasound results. Of the remaining 87 patients, the pelvic fracture types were distributed as follows: 9% type A2, 72% type B, 16% type C, and 3% type C acetabular fractures. Overall US sensitivity for detection of FPF was 80.8%, specificity was 86.9%, positive predictive value was 72.4%, and negative predictive value was 91.4%. Categorization of sensitivity according to pelvic ring fracture type is as follows: type A2 fractures: sensitivity and specificity, 75.0%; type B fractures: sensitivity, 73.3%, specificity, 85.1%; and type C fractures (pelvis and acetabulum): sensitivity and specificity, 100%. Of the true-positive US results, blood was the FPF in 16 of 21 (76%) and urine from intraperitoneal bladder rupture in 4 in 21 (19%) patients. CONCLUSION: US in the initial evaluation of traumatic peritoneal fluid in major pelvic injury patients has lower sensitivity and specificity than previously reported for blunt trauma patients. Additionally, uroperitoneum comprises a substantial proportion of traumatic free peritoneal fluid in patients with MPI.     

坐骨神经损伤及神经生长因子对背根神经节中TrkA及其mRNA表达的调控

目的 研究周围神经损伤、神经再生及外源性神经生长因子对脊神经背根神经节中TrkA及TrkA mRNA表达的影响。方法 将24只SD大鼠坐骨神经从神经中段切断后，外膜缝合修复神经。实验组动物的胫后肌内每日注射NGF200U，术后3、7、14、28d，采用免疫组织化学及原位杂交方法，检查大鼠坐骨神经损伤、神经再生及外源性NGF靶肌肉注射后脊神经背根神经节中TrkA及TrkA mRNA表达。结果 正常背根神经节中有大量的神经元表达TrkA及TrkA mRNA；坐骨神经损伤后，背根神经节中TrkA的表达下降，术后7d组最低，28d接近正常；NGF靶肌肉注射后，背根神经节中TrkA的表达明显增高。结论 神经损伤后背根神经节中TrkA的表达降低，靶肌肉注射外源性NGF背根神经节中TrkA的表达增加，表明靶肌肉注射可作为NGF的有效给药途径.     

Fibrosis of corpus cavernosum in animals following cavernous nerve ablation

Aim: To investigate alterations of smooth muscle celis and collagen fibers in corpus cavernosum following cavernous neurectomy and its relation to the expression of transforming growth factor-β1 (TGF-β1). Methods: Ten adult male SD rats (neurectomy group) were subject to a bilateral cavernous nerve (CN) resection aseptically under an operating microscope, with 6 sham-operated rats as the control. Fifteen weeks after the operation, the penile speci mens were collected and prepared for quantitative-analyzing of ratio of smooth muscle to collagen fibers in corpus cavernosum with confocal microscopy, and for detecting the expression of TGF-β1 by RT-PCR and western-blot. Resulte: Smooth muscle celis that show red color after fluorescent-labeling with tetramethylrhodamine isothiocyanate phalloidin and collagen fibers that produce green autofluorescence after paraformaldehyde fixation were clearly iden tified     

The effect of FK1706 on erectile function following bilateral cavernous nerve crush injury in a rat model

PURPOSE: We investigated the neurotrophic effect of FK1706 on erectile recovery following bilateral cavernous nerve crush injury in a rat model. MATERIALS AND METHODS: A total of 28 male Sprague-Dawley rats were randomly divided into 4 equal groups. Seven animals underwent sham operation and subcutaneous vehicle injection, whereas 21 underwent bilateral cavernous nerve crush injury followed by vehicle injection alone, or by low (0.1 mg/kg) or high (1.0 mg/kg) dose FK1706 treatment. Injections were continued 5 days weekly for 8 weeks. Erectile function was then assessed by cavernous nerve electrostimulation and penile tissue was evaluated immunohistochemically. RESULTS: No erectile dysfunction was identified in the sham treated group (mean maximal intracavernous pressure +/- SEM 106.8 +/- 6.4 cm H(2)O), whereas nerve injury significantly decreased ICP to 17.9 +/- 7.0 cm H(2)O. FK1706 facilitated neural and erectile recovery in a concentration dependent manner with a mean ICP in the high dose FK treatment group of 80.1 +/- 7.8 cm H(2)O compared with 44.1 +/- 12.9 cm H(2)O in the low dose group. Similar stepwise findings were observed using mean area under the curve data. Sham treated animals showed regular axon sizes and shapes with homogenous GAP-43 and neurofilament staining, whereas injured axons showed irregular shapes, sizes and staining patterns. FK1706 treatment restored axon shape and staining patterns. Injury significantly decreased nicotinamide adenine dinucleotide phosphate staining and FK1706 treatment showed a nonsignificant trend toward increased staining. CONCLUSIONS: Bilateral cavernous nerve crush causes reproducible erectile dysfunction, consistent with prior experiments. High dose subcutaneous FK1706 therapy promotes significant neuroregeneration and erectile function recovery.     

Long-term survival of autotransplanted major pelvic ganglion in the corpus cavernosum of adult rats

PURPOSE: Neurogenic impotence is a common complication after radical pelvic surgery, irradiation or perineal trauma. Neuronal transplantation is a new frontier for treating neurological disorders. We investigated whether the major pelvic ganglion can survive and become functional after being implanted into the corpus cavernosum in adult rats. MATERIALS AND METHODS: Adult male rats (13) were divided into 3 groups and sacrificed at 3 time points, namely 30 (4), 60 (5) and 90 (4) days. All rats underwent excision of the right major pelvic ganglion and left cavernous nerve. The right ganglion was implanted into the right crus of the penis. Electrostimulation was applied to the left major pelvic ganglion and cavernous nerve (1.5 mA.) and right crus (10 mA.) at sacrifice. The crural region and left ganglion were then excised for immunostaining of neuronal nitric oxide synthase (nNOS), protein gene product 9.5 and growth associated protein 43. Image analysis was used to calculate the area stained in pixels. Electron microscopy of the implanted area was performed to assess neuronal survival. RESULTS: Although the degree varied, all neuronal implants survived after transplantation. The response to electrostimulation was insufficient to produce erection. No difference was noted among the areas of nNOS staining when specimens from the 3 time points were compared. The area of expression of nNOS, protein gene product 9.5 and growth associated protein 43 was larger in the implanted area than in the surrounding cavernous tissue. Under electron microscopy most surviving implants showed normal ultrastructure, although areas of fibrotic replacement were seen in several implants. CONCLUSIONS: Our results show that the autotransplanted major pelvic ganglion expresses nNOS, protein gene product 9.5 and growth associated protein 43, and survived up to 90 days after implantation into the corpus cavernosum. Further studies with fetal neuronal tissue seem warranted.     

Prevention of pelvic sepsis in major open pelviperineal injury

Compound pelvic fractures are deemed to be one of the most severe orthopaedic injuries with an extremely high morbidity and mortality. After the initial resuscitation phase the prevention of pelvic sepsis is one of the main treatment goals for patients with an open pelvic fracture. If there is a suspicion of a rectal injury or if the wounds are in the perineal area, The Princess Alexandra Hospital's management plan includes early faecal diversion combined with vigorous soft tissue debridement, VAC^® therapy and (if indicated) external fixation of the pelvic fracture. We present our flowchart for the treatment of trauma patients with compound pelvic fractures illustrated by a case report describing a 32 year old patient who sustained an open pelvic ring injury in a workplace accident. The aim of this paper is to underline the importance of a safe, straightforward approach to compound pelvic fractures.     

大鼠阴茎海绵体nNOS神经纤维损伤后再生可能性的研究

目的 探讨海绵体神经损伤对阴茎海绵体ｎＮＯＳ神经纤维的影响。方法 采用免疫组织化学技术ＳＰ法测定大鼠海绵体神经损伤后阴茎勃起组织ｎＮＯＳ神经纤维数量。结果 双侧损伤组术后３周,阴茎海绵体ｎＮＯＳ神经纤维显著减少,６个月后仍很少;单侧损伤组,术后３周损伤侧ｎＮＯＳ神经纤维的改变类似于双侧损伤组,６个月后损伤侧ｎＮＯＳ神经纤维明显增多,接近于对侧水平。结论 阴茎海绵体ｎＮＯＳ神经纤维在单侧海绵体神经损