Chief Investigator: Miss Elena Cavallaro

Funding Amount: $34,874

Recipient: Flinders University

Overview:

Bronchiolitis and pneumonia are leading causes of infant hospitalisation in Australia. The exaggerated immunological insult stemming from these infections on developing lungs is associated with early-childhood asthma development. We have used a small protein, feG, to reduce the consequences of acute lung injury in various adult animal models, by dampening the immune response. We aim to assess feG efficacy in reducing the short and long-term effects of bronchiolitis and pneumonia during infancy, where feG may be an ideal candidate for returning the balance of the immune response in the airways.

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Research Outcomes:

Researchers: Dr Elena Cavallaro, A/Prof Dani-Louise Dixon

Research Completed: 2021

Research Findings:

Bronchiolitis and pneumonia are major causes of infant hospitalisation in Australia. Illness extends beyond the acute phase of infection, with various studies identifying a link between bronchiolitis and pneumonia in infancy and the development of early-childhood chronic wheeze and asthma. Although the mechanism behind this has not been completely elucidated, previous studies indicate an association with the exaggerated immunological insult stemming from these infections on developing lungs as a contributing factor. We have used a small protein, feG, to reduce the consequences of acute lung injury in various adult animal models, by dampening the immune response. We aim to assess feG efficacy in reducing the short and long-term effects of bronchiolitis and pneumonia during infancy, where feG may be an ideal candidate for returning the balance of the immune response in the airways. Respiratory syncytial virus (RSV)-induced bronchiolitis continues to be the leading cause of infant hospitalisation in Australia. Various factors have attributed to this, including: the lack of understanding behind the mechanism of the short- and long-term consequences of the disease; the continued lack of development of an effective therapeutic or vaccination for the disease; and, the absence of small animal model which accurately depicts the disease process we observe in human infants. The lack of an effective animal model hinders the ability to investigate appropriately the effectiveness of potential therapeutics targeting the exaggerated immunological response. This grant has supported the investigation of an effective animal model of RSV bronchiolitis, the differential response to RSV infection in an adult and infant animal model of RSV infection, and the capacity to investigate feG as a potential therapeutic in RSV infection.

Key Outcomes:

Results and Discussion:

RSV in vivo model

Inoculation of the Sprague Dawley rat via intratracheal (I.T) intubation models a neutrophilic response at 96 hours-post instillation, with strong evidence of interstitial peri-bronchiolar neutrophil infiltration and bronchial epithelial cell infection. Of the outcomes analyzed, it appears that an infant Sprague dawley rat model of I.T. RSV infection demonstrates high variability in responses. Despite attempting to capture the immunological immaturity of the human infant in a weanling rat, inherent differences in fetal and postnatal environments between litters may not be accounted for at this age resulting in an unstable model. The adult model of RSV infection demonstrates greater reproducibility at each time-point, with a clear alveolar infiltration of immune cells and accumulation of neutrophil associated cytokines (CINC-1) at 96-hours. In addition to immunological changes, physiological changes in lung function are also observed in the adult model of RSV infection.

Assessing feG efficacy in an RSV in vivo model

A time-point of 96 hours post inoculation was selected to model RSV infection in vivo, with feG administrated at 72 hours post RSV inoculation due to first evidence of disease outcomes (attempting to replicate what would occur in a hospital setting). For LPS, a time point of 2 hours post LPS instillation was selected for feG administration due to previous studies from our laboratory indicating an uptrend in inflammatory mediators at this timepoint.

In the adult RSV in vivo model, feG administered IV 72 hours post inoculation appears to increase the number of cells entering the lungs at 96 hours post inoculation. Despite an increase in immune cells, feG administration does appear to improve lung function as measured through respiratory mechanics. This may indicate a refractory cellular infiltrate occurring between feG administration and outcomes analysis. Due to the variability of the inflammatory response in the infant RSV in vitro model, feG was not assessed in this age group.

Conclusion:

Overall, RSV appears to differentially effect adult and infant Sprague Dawley rats. Further investigation is currently underway to understand the mechanisms behind these differences, and the contribution to the variability observed in the immune response in the infant model. We aim to further investigate the mechanisms behind the differential responses observed in the adult and infant animals models, in the hope that this will help to elucidate potential pathways associated with the differential responses to RSV infection observed in humans. In the adult RSV model, at the timings selected, feG appears to worsen immune cell influx into the lungs. Despite this, feG administration is associated with improved respiratory function. Further investigation into alternative timing and additional dosing of feG will next be assessed to investigate the mechanisms behind this differential response, with an aim to determine optimal dosing to maintain the improvement in respiratory physiological outcomes whilst preventing a potential second wave of immune cell influx and the associated damage. In vivo human cell models are currently being used to assess potential changes in neutrophil function with feG administration to further clarify what has been observed in the in vivo model.

Research Papers:

Papers (all in preparation):

• Assessing the Temporal Response of Respiratory Syncytial Virus Inoculation in a Rat in vivo Bronchiolitis Model: A Comparative Analysis of Immunological and Physiological Responses by Age. E.C Cavallaro, J.H.Mc Evoy-May, K.D. Forsyth, D-L. Dixon
• Developing a Paediatric in vivo Model of Lipopolysaccharide-lnduced Acute Lung Injury: A Comparative Analysis of Immunological and Physiological Responses by Age. E.C Cavallaro, J. H. McEvoy-May, D-L. Dixon
• The role of tripeptide feG in Ameliorating the Acute Immunological and Physiological Changes in an in vivo Model of RSV Bronchiolitis. E.C Cavallaro, J. H. Mc Evoy-May, D-L. Dixon

Conference Presentations (presented – copy provided at end of file):

• Cavallaro EC, McEvoy JH, Dixon DL. Immunological and physiological characterization of an in vivo adult and infant model of respiratory syncytial virus bronchiolitis: modelling the neutrophilic response. ReSViNET 5th Expert Meeting {2019}

Cavallaro EC, McEvoy JH, Dixon DL. Immunological and physiological characterization of an in vivo adult and infant model of respiratory syncytial virus bronchiolitis: modelling the neutrophilic response. Australian and New Zealand Society for Immunology {2019}

Related Publications:

Future Outcomes: