Mechanism by Which Testicular Macrophages Initiate Immune Responses to Bacteria, while Maintaining Testicular Immune Privilege. (2011)

Testicular macrophages (TM) represent the largest population of immune cells in the testis of mammals constituting up to 25% of interstitial cells. They play an important role in the balance between defense against invading microorganisms and "testicular immune privilege" which serves to protect the neo-antigens of the meiotic and haploid germ cells appearing during puberty after establishment of self-tolerance. The testicular immune response, therefore, requires a specific regulatory mechanism which safeguards developing male germ cells, while permitting qualitatively normal inflammatory responses and maintaining protection against infection. Ascending bacterial infections of the male genitourinary tract by Escherichia coli are an important cause of male infertility. Thus understanding mechanisms by which immunocompetent cells such as testicular macrophages (TM) respond to infection and how bacterial pathogens manipulate defense pathways is of importance. Whole genome expression profiling of TM and peritoneal macrophages (PM) infected with uropathogenic E. coli (UPEC) revealed that PM responded with a much larger number of differentially regulated genes than TM (PM: 1710 genes, TM: 400 genes; total of both times points 30 Min and 60 Min, FDR < 0.05). Only a small number of genes (n=125) showed similar regulation in both macrophage populations. Hence, after infection with UPEC PM mobilized 5.3% of the genome (represented by the whole genome microarray), while TM mobilized only 1.2% of the genome. Interestingly, a multitude of genes implicated in calcium signaling pathways was a common feature which indicated a role of calcium-dependent nuclear factor of activated T cells (NFAT) signaling. UPEC-dependent NFAT activation was confirmed in both cultured TM and in TM in an in vivo orchitis model. Elevated expression of NFATC2-regulated anti-inflammatory cytokines was found in TM (IL-4, IL-13) and PM (IL-3, IL-4, IL-13). NFATC2 is activated by rapid influx of calcium, an activity delineated to the pore forming toxin alpha-hemolysin by mutant analysis. Alpha-hemolysin suppressed IL-6 and TNF-alpha cytokine release from PM and caused differential activation of MAP kinase and AP-1 signaling pathways in TM and PM leading to reciprocal expression of key pro-inflammatory cytokines in PM (IL-1alpha, IL-1beta, IL-6 downregulated) and TM (IL-1beta, IL-6 upregulated). In addition, unlike PM, LPS-treated TM were refractory to NFkB activation shown by the absence of degradation of IkBalpha and lack of pro-inflammatory cytokine secretion (IL-6, TNF-alpha). Although the immune response of TM compared to PM is reduced by blockage of NFKB activation, cells maintain a general responsiveness by activation of MAP kinase and AP-1 signaling pathways following LPS and alpha-hemolysin stimulation. In summation, these features enable TM to fulfill apparently paradox tasks, i.e. protection against microbes, while at the same time maintaining the immune privileged status of the testis to protect the developing germ cells. This research was funded by the Deutsche Forschungsgemeinschaft (DFG) Program grant KFO 181. (platform)