Interferon-γ inhibits group B Streptococcus survival within human endothelial cells

Endothelial dysfunction is a major component of the pathophysiology of septicaemic group B Streptococcus (GBS) infections. Although cytokines have been shown to activate human umbilical vein endothelial cells (HUVECs), the capacity of interferon (IFN)-γ to enhance the microbicidal activity of HUVECs against GBS has not been studied. We report that the viability of intracellular bacteria was reduced in HUVECs activated by IFN-γ. Enhanced fusion of lysosomes with bacteria-containing vacuoles was observed by acid phosphatase and the colocalisation of Rab-5, Rab-7 and lysosomal-associated membrane protein-1 with GBS in IFN-γ-activated HUVECs. IFN-γ resulted in an enhancement of the phagosome maturation process in HUVECs, improving the capacity to control the intracellular survival of GBS.

Papel de metaloproteases de Estreptococos do grupo B na interação, viabilidade celular e indução de apoptose e necrose em células endoteliais e epiteliais humanas / Role of metalloproteinases of Group B Streptococci in the interaction, cell viability and induction of apoptosis and necrosis in human epithelial and endothelial cells

Estreptococos do grupo B (EGB) é a principal causa de sepse e meningite neonatal e tem sido recentemente reconhecido como patógeno responsável por infecções invasivas em adultos imunocomprometidos (idosos ou portadores de doenças crônicas). Os EGB produzem inúmeras enzimas extracelulares, várias das quais interagem com o sistema imune do hospedeiro e são importantes durante a interação EGB-hospedeiro, bem como para o desenvolvimento da doença. Estudos anteriores mostraram que metaloproteases estão envolvidas em várias vias metabólicas em diferentes tipos celulares. Por esta razão, nós decidimos investigar o possível envolvimento de metaloproteases de EGB durante a interação celular e apoptose/necrose induzida pelo micro-organismo em células endoteliais da veia umbilical humana (HUVEC) e da linhagem de epitélio respiratório (A549). Tratamento de EGB com inibidores de metaloproteases (EDTA, EGTA e FEN) não induziu alterações no crescimento bacteriano, mas promoveu alterações na expressão de proteínas de superfície, capacidade adesiva e perfil de sobrevivência intracelular do patógeno. O EGB e o sobrenadante do crescimento bacteriano (meio condicionado; MC) promoveram a morte das células HUVEC e A549. Contudo, o tratamento com inibidores de metaloproteases restauraram a viabilidade celular induzida pelos EGB e o MC, sugerindo que metaloproteases bacteriana estão envolvidas no rompimento da barreira celular, promovendo a disseminação bacteriana. Este trabalho descreve pela primeira vez apoptose e necrose induzidas pelo EGB e MC em HUVEC e células A549 após 24h de incubação, respectivamente. Nós também observamos redução da pró-caspase-3 após infecção das HUVEC com EGB e MC, sugerindo ativação da caspase-3. Além disso, o aumento da expressão da proteína pró-apoptótica Bax e diminuição dos níveis da proteína anti-apoptótica Bcl-2 em HUVEC, demonstram o envolvimento do mecanismo apoptótico mitocondrial (via intrínseca). A melhor compreensão das bases molecular...

Group B streptococcus (GBS) is the leading cause of neonatal sepsis and meningitis and has recently been recognized as an increasingly common cause of invasive disease in immunocompromised adults (elderly or chronic diseases). GBS produces a number of extracellular enzymes, several of which interact with the host immune system and are important for the GBS- host interaction and for the development of disease. Previous studies showed that metalloproteases are involved in several metabolic pathways in different cellular types. For this reason, we decided to investigate the possible involvement of GBS metalloproteases during cell interaction and apoptosis/necrosis induced by microorganism in human umbilical vein endothelial cells (HUVEC) and epithelial respiratory cells line (A549). Treatment of GBS with metalloproteases inhibitors (EDTA, EGTA and PHEN) did not induce alteration on bacterial growth, but promoted changes in the expression of surface proteins, adhesive capacity and profile of intracellular survival of the pathogen. The GBS and supernatant of bacterial growth medium (conditioned medium; MC) promoted the death of HUVEC and A549 cells. However, the metalloproteases inhibitors treatment restored the cellular viability induced by GBS and MC, suggesting that GBS metalloproteases are involved in the disruption of cell barrier, promoting bacterial dissemination. This study describes for the first time apoptosis and necrosis induced by GBS and MC in HUVEC and A549 cells after 24h incubation, respectively. We also observe reduction of pro-caspase-3 after infection of HUVEC with GBS and MC, suggesting activation of caspase-3. Moreover, the over-expression of pro -apoptotic protein Bax and decrease of anti-apoptotic protein Bcl-2 levels in HUVEC show the involvement of mitochondrial apoptotic mechanism (intrinsic via). Enhanced understanding of the molecular basis of GBS pathogenesis may pinpoint novel bacterial and host molecules that can represent novel...