Investigating the pro-inflammatory differentiation of macrophages with bacterial ghosts in potential infection control.

In the complex realm of bacterial infections, particularly those caused by Staphylococcus aureus (S. aureus), macrophages play a pivotal role in orchestrating the immune response. During the initial stages of infection, the monocytes give rise to macrophages with a pro-inflammatory (M1 type) behaviour, engulfing and neutralizing the invading pathogens. However, under the sustained influence of S. aureus infection, monocytes can undergo a transition into an anti-inflammatory M2 state (pro-infection) rather than the M1 state (anti-infection), thereby compromising effective infection control. Therefore, it is necessary to develop a strategy that would preserve the pro-inflammatory functions of macrophages, in a safe and controlled manner. For this, we focused on harnessing the potential of S. aureus-derived ghost cells (GCs) which are non-live empty envelopes of bacterial cells, but with the antigenic determinants intact. Through a unique Lugol's-iodine treatment, we generated GCs and characterization of these GCs using gel electrophoresis, FTIR, flow cytometry, TEM, and SEM confirmed their structural integrity. Following this, we assessed the extend of cellular association of the GCs with RAW267.4 macrophages, and observed an immediate interaction between the two, as evident from the flowcytometry and microscopy studies. We then performed macrophage polarisation on a human monocyte-macrophage model cell line, THP-1. Our findings revealed that GCs effectively activated macrophages, and promoted a pro-inflammatory polarisation with the expression of M1 differentiation markers (CD86, TNFα, IL-1ß, IL-6, IL-12) evaluated through both qPCR and ELISA. Interestingly an intermediary expression of M2 markers viz., CD206 and IL-10 was also observed, but was overruled by the enhanced expression of M1 markers at a later time point. Overall, our study introduces a novel approach utilizing GCs to guide naïve macrophages towards M1 subtypes, thereby potentiating immune responses during microbial infections. This innovative strategy can modulate macrophage function, ultimately improving outcomes in S. aureus infections and beyond.

Correlation of desmoglein 1 and 3 immunohistochemistry with autoantibody levels and clinical severity in pemphigus.

BACKGROUND: Pemphigus is a chronic potentially fatal autoimmune bullous disorder. Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are the two common subtypes. PV is the most common and aggressive type characterized by oral mucosal erosions and cutaneous lesions. PF presents with blisters on the scalp, face, and upper trunk, and spares the mucosae. Direct immunofluorescence (DIF) is the gold standard for diagnosis. Immunohistochemistry (IHC) is an emerging alternate diagnostic tool. In this study, our objectives were to identify the staining patterns of desmoglein 1 (dsg 1) and desmoglein 3 (dsg 3) IHC and to correlate the same with autoantibody levels and clinical severity in patients with PV and PF. METHODS: Forty-nine clinically, histologically, and DIF-confirmed cases of pemphigus were included in the study. The IHC patterns were scored from 0 to 3+ with 3+ dsg 1 IHC exhibiting intense membranous staining in the upper layers of the epidermis and 3+ dsg 3 IHC showing intense basal layer staining. Enzyme-linked immunosorbent assay (ELISA) for anti-dsg 1 and 3 antibodies was performed in 38 cases where serum samples were available. The pemphigus disease activity index system was utilized for clinical scoring. RESULTS: A 0 to 1+ score was observed for dsg 1 IHC in 100% of PF cases. A score of 0 to 1+ was observed for dsg 3 IHC in 97.3% of PV cases. One hundred percent of cases with PF and 83.9% of patients with PV tested positive for ELISA anti-dsg 1 and 3 antibody titers, respectively. Anti-dsg 1 and 3 ELISA titers significantly correlated with the dsg 1 and dsg 3 IHC scores. The mucosal scores showed a significant association with both dsg 1 and 3 IHC (p < 0.001). The cutaneous scores showed a significant association with the dsg 3 IHC (p < 0.001). CONCLUSION: The IHC patterns for dsg 1 and 3 proved reliable in giving concordant results with the ELISA antibody titers and clinical severity.