Nano-adjuvanted dry powder vaccine for the mucosal immunization against airways pathogens

Nasal vaccination has been shown to provide optimal protection against respiratory pathogens. However, mucosal vaccination requires the implementation of specific immunization strategies to improve its effectiveness. Nanotechnology appears a key approach to improve the effectiveness of mucosal vaccines, since several nanomaterials provide mucoadhesion, enhance mucosal permeability, control antigen release and possess adjuvant properties. Mycoplasma hyopneumoniae is the main causative agent of enzootic pneumonia in pigs, a respiratory disease responsible for considerable economic losses in the pig farming worldwide. The present work developed, characterized, and tested in vivo an innovative dry powder nasal vaccine, obtained from the deposition on a solid carrier of an inactivated antigen and a chitosan-coated nanoemulsion, as an adjuvant. The nanoemulsion was obtained through a low-energy emulsification technique, a method that allowed to achieve nano droplets in the order of 200 nm. The oil phase selected was alpha-tocopherol, sunflower oil, and poly(ethylene glycol) hydroxystearate used as non-ionic tensioactive. The aqueous phase contained chitosan, which provides a positive charge to the emulsion, conferring mucoadhesive properties and favoring interactions with inactivated M. hyopneumoniae. Finally, the nanoemulsion was layered with a mild and scalable process onto a suitable solid carrier (i.e., lactose, mannitol, or calcium carbonate) to be transformed into a solid dosage form for administration as dry powder. In the experimental study, the nasal vaccine formulation with calcium carbonate was administered to piglets and compared to intramuscular administration of a commercial vaccine and of the dry powder without antigen, aimed at evaluating the ability of IN vaccination to elicit an in vivo local immune response and a systemic immune response. Intranasal vaccination was characterized by a significantly higher immune response in the nasal mucosa at 7 days post-vaccination, elicited comparable levels of Mycoplasma-specific IFN-gamma secreting cells and comparable, if not higher, responsiveness of B cells expressing IgA and IgG in peripheral blood mononuclear cells, with those detected upon a conventional intramuscular immunization. In conclusion, this study illustrates a simple and effective strategy for the development of a dry powder vaccine formulation for nasal administration which could be used as alternative to current parenteral commercial vaccines

Detecting early signs of skin damage using non-invasive biophysical parameters

Pressure ulcers (PUs) and incontinence-associated dermatitis (IAD) are types of skin damage that result from prolonged exposure to external insults including pressure, shear, friction, and moisture. The clinical symptoms, which often lead to chronic wounds, can involve erythema, skin swelling, oedema and skin breakdown. It is imperative to detect early signs of these conditions prior to chronic damage. However, subjective visual and tactile skin assessments typically used by clinicians lack predictive capability. Objective biophysical approaches have been suggested to provide the means of monitoring the early signs of damage, although further evidence is required to establish their performance in a range of clinically relevant situations. The goal of the doctoral research is to utilise an array of non-invasive biophysical sensors to understand the pathophysiological changes in the skin following different insults associated with PUs and IAD. This thesis presents the results from a series of complimentary retrospective and prospective studies. These involved the analysis of established data from the host lab, a survey of skin damage resulting from the application of personal protective equipment (PPE), and two prospective lab-based studies. The former involved the recruitment of able-bodied volunteers and healthcare workers who were exposed to moisture and mechanical loads through incontinence pads and lying postures, and respirator protective equipment (RPE), respectively. These findings were used to inform a cohort study of hospitalised patients with stage I pressure ulcers.Results revealed that prolonged exposure of the skin to mechanical loads and moisture can disrupt its barrier function, as evidenced by enhanced values of trans-epidermal water loss and stratum corneum hydration. Furthermore, among the biophysical skin parameters evaluated, TEWL and hydration were able to distinguish between compromised and healthy adjacent skin sites, but not between different insults. During a COVID-19 study of adverse reactions to PPE, cohorts of healthcare workers revealed that they had indentation marks, pressure damage, itchiness, rashes, and spots. The subsequent study of biophysical changes in skin health identified distinct changes in skin barrier function and hydration over vulnerable bony landmarks on the face (bridge of the nose) associated with RPE application. Findings from this study showed that the outputs of these parameters were influenced by the subject extrinsic and intrinsic factors, namely body mass index and daily working hours. The final clinical study revealed highly localised changes (&lt;5mm) in skin barrier function over the pressure ulcer site. In contrast to the lab-based studies, skin hydration did not differentiate between healthy and damaged skin. Complementary analyses also revealed the important role of inflammatory cytokines in the early detection of skin compromise, as well as the potential of epidermal corneocytes as novel biomarkers reflective of changes in skin health. The use of biophysical tools to monitor local changes in skin health could represent an important adjunct to clinical practice, which currently relies on subjective skin assessment. However, to support its wider translation in different care settings, future research is required to analyse time-dependent changes in these parameters, in both acute and long-term care facilities, to assess the prognostic value when determining skin status (healing or progressing to wounds). In addition, engagement with healthcare workers to analyse the barriers and facilitators to adoption along with the cost-effectiveness of introducing these tools is needed prior to wider adoption in practice. <br/

Nano-adjuvanted dry powder vaccine for the mucosal immunization against airways pathogens

Nasal vaccination has been shown to provide optimal protection against respiratory pathogens. However, mucosal vaccination requires the implementation of specific immunization strategies to improve its effectiveness. Nanotechnology appears a key approach to improve the effectiveness of mucosal vaccines, since several nanomaterials provide mucoadhesion, enhance mucosal permeability, control antigen release and possess adjuvant properties. Mycoplasma hyopneumoniae is the main causative agent of enzootic pneumonia in pigs, a respiratory disease responsible for considerable economic losses in the pig farming worldwide. The present work developed, characterized, and tested in vivo an innovative dry powder nasal vaccine, obtained from the deposition on a solid carrier of an inactivated antigen and a chitosan-coated nanoemulsion, as an adjuvant. The nanoemulsion was obtained through a low-energy emulsification technique, a method that allowed to achieve nano droplets in the order of 200 nm. The oil phase selected was alpha-tocopherol, sunflower oil, and poly(ethylene glycol) hydroxystearate used as non-ionic tensioactive. The aqueous phase contained chitosan, which provides a positive charge to the emulsion, conferring mucoadhesive properties and favoring interactions with inactivated M. hyopneumoniae. Finally, the nanoemulsion was layered with a mild and scalable process onto a suitable solid carrier (i.e., lactose, mannitol, or calcium carbonate) to be transformed into a solid dosage form for administration as dry powder. In the experimental study, the nasal vaccine formulation with calcium carbonate was administered to piglets and compared to intramuscular administration of a commercial vaccine and of the dry powder without antigen, aimed at evaluating the ability of IN vaccination to elicit an in vivo local immune response and a systemic immune response. Intranasal vaccination was characterized by a significantly higher immune response in the nasal mucosa at 7 days post-vaccination, elicited comparable levels of Mycoplasma-specific IFN-γ secreting cells and comparable, if not higher, responsiveness of B cells expressing IgA and IgG in peripheral blood mononuclear cells, with those detected upon a conventional intramuscular immunization. In conclusion, this study illustrates a simple and effective strategy for the development of a dry powder vaccine formulation for nasal administration which could be used as alternative to current parenteral commercial vaccines