M1 Macrophages Are Predominantly Recruited to the Major Pelvic Ganglion of the Rat Following Cavernous Nerve Injury

Abstract

Introduction Neurogenic erectile dysfunction (ED) is a common sequela of radical prostatectomy. The etiology involves injury to the autonomic cavernous nerves (CNs), which arise from the major pelvic ganglion (MPG), and subsequent neuroinflammation, which leads to recruitment of macrophages to the injury site. Currently, two macrophage phenotypes are known: neurotoxic M1 macrophages and neuroprotective M2 macrophages. Aim To examine whether bilateral cavernous nerve injury (BCNI) in a rat model of ED would increase recruitment of neurotoxic M1 macrophages to the MPG. Methods Male Sprague Dawley rats underwent bilateral cavernous nerve injury (BCNI) and the major pelvic ganglion (MPG) was harvested at various time points post-nerve injury. The corpora cavernosa was used to evaluate tissue myography responses to electrical field stimulation ex vivo. qPCR was used to examine the gene expression of global macrophage markers, M1 macrophage markers, M2 macrophage markers and cytokines/chemokines in the MPG. Mathematical calculation of M1/M2 index was used to quantify macrophage changes temporally. Western blot of MPG tissues was used to evaluate the protein amount of M1 and M2 macrophages markers quantitatively. Immunohistochemistry staining of MPGs for CD68, CD86 and CD206 was used to characterize M1 and M2 macrophage infiltration. Maine Outcome Measures Corpora cavernosa responsiveness ex vivo; gene (pPCR) and protein (Western Blot) expression of M1 and M2 markers, cytokines/chemokines; immunohistochemical localization of M1 and M2 macrophages. Results BCNI impaired corporal parasympathetic-mediated relaxation response to EFS and enhanced contraction response to EFS. Gene expression of pro-inflammatory (Il1b, Il16, Tnfa, Tgfb, Ccl2, Ccr2) and anti-inflammatory (Il10) cytokines was upregulated in the MPG 48 hours following injury. M1 markers (CD86, iNOS, IL1b) and M2 markers (CD206, ARG1, IL-10) was increased following BCNI in the MPG with the M1/M2 index above 1.0 indicating that more M1 macrophages are recruited to the MPG than M2 macrophages. Protein expression of the M1 macrophage markers (iNOS) was increased in MPGs following BCNI. However, the protein amount of M2 macrophage markers (Arginase-1) remained unchanged. Immunohistochemical characterization demonstrated predominant increases in M1 (CD68+CD86+) macrophages in the MPG following BCNI. Conclusion These results suggest that an increase in M1 macrophage infiltration of the MPG following BCNI is associated with impaired neurogenic mediated erectile tissue physiology ex vivo and thus has significant implications for CN axonal repair. Future studies are needed to demonstrate that inhibition of M1 macrophage recruitment prevents ED following CN-injury.

M2b macrophage polarization and its roles in diseases

“…32 However, abundant studies have shown that CD86 is expressed in and used as a marker to identify M1 macrophages. [55][56][57][58][59][60][61] These studies indicate that CD86 should be expressed in both M1 and M2b macrophages. Although many review articles have shown that CD86 is the marker for M1 and M2b, 5,6,8,41,62,63 CD86 is a suitable marker for discriminating M2b from the other subtypes of M2 macrophages, but not from M1 macrophages.…”

“…Among the subpopulations of macrophages, CD86 is expressed in M2b macrophages and considered as the marker for this subtype . However, abundant studies have shown that CD86 is expressed in and used as a marker to identify M1 macrophages . These studies indicate that CD86 should be expressed in both M1 and M2b macrophages.…”

Three-dimensional organoid culture reveals involvement of Wnt/β-catenin pathway in proliferation of bladder cancer cells

“…Total RNA was extracted from organoids using the RNeasy system (Qiagen, Hilden, Germany). Real-time qPCR was performed using the StepOnePlus system (Applied Biosystems, Foster City, CA, USA) as previously described [ 51 ]. TaqMan gene expression assays for AXIN2, KRT20 (CK20), KRT18 (CK18), KRT14 (CK14), KRT5 (CK5), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were used.…”

European Society for Sexual Medicine Consensus Statement on the Use of the Cavernous Nerve Injury Rodent Model to Study Postradical Prostatectomy Erectile Dysfunction

“…In nerve-sparing surgery, erectile dysfunction (ED) occurs as a consequence of iatrogenic damage to the periprostatic neurovascular bundle, which results in neuropraxia of the cavernous nerves (CNs). 5 , 6 , 7 , 8 , 9 Neurapraxia, as per the Seddon classification, is a temporary failure of nerve conduction because of a damage to the myelin sheath. 10 , 11 The Seddon classification of nerve injury separates injury based on the scale from microscopic to macroscopic along with severity of tissue injury, prognosis, and time for recovery.…”