Dataset Information

Local Activation of the Alternative Pathway of Complement System in Mycotic Keratitis Patient Tear.

ABSTRACT: Aspergillus flavus and Fusarium solani are the predominant causative agents of mycotic keratitis in the tropical part of the world. Tear proteins play a major role in the innate immune response against these fungal infections as has been shown by the presence of complement proteins and neutrophil extracellular trap proteins in keratitis patients tear. In this study, we established the presence of the components of the alternate pathway of complement system and their functional state in the tear film of mycotic keratitis patients. The complement proteins namely, C3 and CFH were found only in the open-eye tear of patients but not in control individuals. In vitro analysis showed binding of purified C3b and CFH to fungal spores, which confirmed that the spores can provide a foreign surface for forming the complement complex. Analysis of spore bound tear proteins by mass spectrometry exhibited the presence of known proteins of the alternate pathway complement cascade in keratitis patient tear. Hemolytic assay using rabbit RBC confirmed the presence of a functional alternate pathway of complement cascade in the tear proteome of the patients. The presence of negative regulators, CFH and CFI, in the patient tear indicate that the complement activity is tightly regulated during fungal infection. Mass spectrometry data show vitronectin and clusterin, two known inhibitors of the membrane attack complex only in the patient tear. These data demonstrate the activation of the alternate pathway of complement cascade during the early stages of infection. Interestingly, the production of multiple negative regulators of complement cascade implies the pathogen can effectively evade the host complement system during infection.

SUBMITTER: Shait Mohammed MR

PROVIDER: S-EPMC7218074 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Local Activation of the Alternative Pathway of Complement System in Mycotic Keratitis Patient Tear.

Shait Mohammed Mohammed Razeeth MR Krishnan Sandhya S Amrathlal Rabbind Singh RS Jayapal Jeya Maheshwari JM Namperumalsamy Venkatesh Prajna VP Prajna Lalitha L Kuppamuthu Dharmalingam D

Frontiers in cellular and infection microbiology 20200506

<i>Aspergillus flavus and Fusarium solani</i> are the predominant causative agents of mycotic keratitis in the tropical part of the world. Tear proteins play a major role in the innate immune response against these fungal infections as has been shown by the presence of complement proteins and neutrophil extracellular trap proteins in keratitis patients tear. In this study, we established the presence of the components of the alternate pathway of complement system and their functional state in th ...[more]

PMID: 32435625

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Project description:The innate immune system plays a major role in the pathogenesis of nonalcoholic steatohepatitis (NASH). Recently we reported complement activation in human NASH. However, it remained unclear whether the alternative pathway of complement, which amplifies C3 activation and which is frequently associated with pathological complement activation leading to disease, was involved. Here, alternative pathway components were investigated in liver biopsies of obese subjects with healthy livers (n = 10) or with NASH (n = 12) using quantitative PCR, Western blotting, and immunofluorescence staining. Properdin accumulated in areas where neutrophils surrounded steatotic hepatocytes, and colocalized with the C3 activation product C3c. C3 activation status as expressed by the C3c/native C3 ratio was 2.6-fold higher (p<0.01) in subjects with NASH despite reduced native C3 concentrations (0.94±0.12 vs. 0.57±0.09; p<0.01). Hepatic properdin levels positively correlated with levels of C3c (rs = 0.69; p<0.05) and C3c/C3 activation ratio (rs = 0.59; p<0.05). C3c, C3 activation status (C3c/C3 ratio) and properdin levels increased with higher lobular inflammation scores as determined according to the Kleiner classification (C3c: p<0.01, C3c/C3 ratio: p<0.05, properdin: p<0.05). Hepatic mRNA expression of factor B and factor D did not differ between subjects with healthy livers and subjects with NASH (factor B: 1.00±0.19 vs. 0.71±0.07, p = 0.26; factor D: 1.00±0.21 vs. 0.66±0.14, p = 0.29;). Hepatic mRNA and protein levels of Decay Accelerating Factor tended to be increased in subjects with NASH (mRNA: 1.00±0.14 vs. 2.37±0.72; p = 0.22; protein: 0.51±0.11 vs. 1.97±0.67; p = 0.28). In contrast, factor H mRNA was downregulated in patients with NASH (1.00±0.09 vs. 0.71±0.06; p<0.05) and a similar trend was observed with hepatic protein levels (1.12±0.16 vs. 0.78±0.07; p = 0.08). Collectively, these data suggest a role for alternative pathway activation in driving hepatic inflammation in NASH. Therefore, alternative pathway factors may be considered attractive targets for treating NASH by inhibiting complement activation.

Project description:The complement system is an integral part of innate immunity that detects and eliminates invading pathogens through a cascade of reactions. The destructive effects of the complement activation on host cells are inhibited through versatile regulators that are present in plasma and bound to membranes. Impairment in the capacity of these regulators to function in the proper manner results in autoimmune diseases. To better understand the delicate balance between complement activation and regulation, we have developed a comprehensive quantitative model of the alternative pathway. Our model incorporates a system of ordinary differential equations that describes the dynamics of the four steps of the alternative pathway under physiological conditions: (i) initiation (fluid phase), (ii) amplification (surfaces), (iii) termination (pathogen), and (iv) regulation (host cell and fluid phase). We have examined complement activation and regulation on different surfaces, using the cellular dimensions of a characteristic bacterium (E. coli) and host cell (human erythrocyte). In addition, we have incorporated neutrophil-secreted properdin into the model highlighting the cross talk of neutrophils with the alternative pathway in coordinating innate immunity. Our study yields a series of time-dependent response data for all alternative pathway proteins, fragments, and complexes. We demonstrate the robustness of alternative pathway on the surface of pathogens in which complement components were able to saturate the entire region in about 54 minutes, while occupying less than one percent on host cells at the same time period. Our model reveals that tight regulation of complement starts in fluid phase in which propagation of the alternative pathway was inhibited through the dismantlement of fluid phase convertases. Our model also depicts the intricate role that properdin released from neutrophils plays in initiating and propagating the alternative pathway during bacterial infection.

Dataset Information

Local Activation of the Alternative Pathway of Complement System in Mycotic Keratitis Patient Tear.

Publications

Local Activation of the Alternative Pathway of Complement System in Mycotic Keratitis Patient Tear.

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