Cryptococcus neoformans rapidly invades the murine brain by sequential breaching of airway and endothelial tissues barriers, followed by engulfment by microglia.

Cryptococcus neoformans causes lethal meningitis and accounts for approximately 10%-15% of AIDS-associated deaths worldwide. There are major gaps in our understanding of how this fungus invades the mammalian brain. To investigate the dynamics of C. neoformans tissue invasion, we mapped fungal localization and host cell interactions in infected brain, lung, and upper airways using mouse models of systemic and airway infection. To enable this, we developed an in situ imaging pipeline capable of measuring large volumes of tissue while preserving anatomical and cellular information by combining thick tissue sections, tissue clarification, and confocal imaging. We confirm high fungal burden in mouse upper airway after nasal inoculation. Yeast in turbinates were frequently titan cells, with faster kinetics than reported in mouse lungs. Importantly, we observed one instance of fungal cells enmeshed in lamina propria of the upper airways, suggesting penetration of airway mucosa as a possible route of tissue invasion and dissemination to the bloodstream. We extend previous literature positing bloodstream dissemination of C. neoformans, by finding viable fungi in the bloodstream of mice a few days after intranasal infection. As early as 24 h post systemic infection, the majority of C. neoformans cells traversed the blood-brain barrier, and were engulfed or in close proximity to microglia. Our work presents a new method for investigating microbial invasion, establishes that C. neoformans can breach multiple tissue barriers within the first days of infection, and demonstrates microglia as the first cells responding to C. neoformans invasion of the brain.IMPORTANCECryptococcal meningitis causes 10%-15% of AIDS-associated deaths globally. Still, brain-specific immunity to cryptococci is a conundrum. By employing innovative imaging, this study reveals what occurs during the first days of infection in brain and in airways. We found that titan cells predominate in upper airways and that cryptococci breach the upper airway mucosa, which implies that, at least in mice, the upper airways are a site for fungal dissemination. This would signify that mucosal immunity of the upper airway needs to be better understood. Importantly, we also show that microglia, the brain-resident macrophages, are the first responders to infection, and microglia clusters are formed surrounding cryptococci. This study opens the field to detailed molecular investigations on airway immune response, how fungus traverses the blood-brain barrier, how microglia respond to infection, and ultimately how microglia monitor the blood-brain barrier to preserve brain function.

Cryptococcus neoformans rapidly invades the murine brain by sequential breaching of airway and endothelial tissues barriers, followed by engulfment by microglia.

The fungus Cryptococcus neoformans causes lethal meningitis in humans with weakened immune systems and is estimated to account for 10-15% of AIDS-associated deaths worldwide. There are major gaps in our understanding of how this environmental fungus evades the immune system and invades the mammalian brain before the onset of overt symptoms. To investigate the dynamics of C. neoformans tissue invasion, we mapped early fungal localisation and host cell interactions at early times in infected brain, lung, and upper airways using mouse models of systemic and airway infection. To enable this, we developed an in situ imaging pipeline capable of measuring large volumes of tissue while preserving anatomical and cellular information by combining thick tissue sections, tissue clarification, and confocal imaging. Made possible by these techniques, we confirm high fungal burden in mouse upper airway turbinates after nasal inoculation. Surprisingly, most yeasts in turbinates were titan cells, indicating this microenvironment enables titan cell formation with faster kinetics than reported in mouse lungs. Importantly, we observed one instance of fungal cells enmeshed in lamina propria of upper airways, suggesting penetration of airway mucosa as a possible route of tissue invasion and dissemination to the bloodstream. We extend previous literature positing bloodstream dissemination of C. neoformans, via imaging C. neoformans within blood vessels of mouse lungs and finding viable fungi in the bloodstream of mice a few days after intranasal infection, suggesting that bloodstream access can occur via lung alveoli. In a model of systemic cryptococcosis, we show that as early as 24 h post infection, majority of C. neoformans cells traversed the blood-brain barrier, and are engulfed or in close proximity to microglia. Our work establishes that C. neoformans can breach multiple tissue barriers within the first days of infection. This work presents a new method for investigating cryptococcal invasion mechanisms and demonstrates microglia as the primary cells responding to C. neoformans invasion.

Challenges in the care of individuals with severe primary insulin-like growth factor-I deficiency (SPIGFD): an international, multi-stakeholder perspective.

BACKGROUND: Severe primary insulin-like growth factor-I (IGF-I) deficiency (SPIGFD) is a rare growth disorder characterized by short stature (standard deviation score [SDS] ≤ 3.0), low circulating concentrations of IGF-I (SDS ≤ 3.0), and normal or elevated concentrations of growth hormone (GH). Laron syndrome is the best characterized form of SPIGFD, caused by a defect in the GH receptor (GHR) gene. However, awareness of SPIGFD remains low, and individuals living with SPIGFD continue to face challenges associated with diagnosis, treatment and care. OBJECTIVE: To gather perspectives on the key challenges for individuals and families living with SPIGFD through a multi-stakeholder approach. By highlighting critical gaps in the awareness, diagnosis, and management of SPIGFD, this report aims to provide recommendations to improve care for people affected by SPIGFD globally. METHODS: An international group of clinical experts, researchers, and patient and caregiver representatives from the SPIGFD community participated in a virtual, half-day meeting to discuss key unmet needs and opportunities to improve the care of people living with SPIGFD. RESULTS: As a rare disorder, limited awareness and understanding of SPIGFD amongst healthcare professionals (HCPs) poses significant challenges in the diagnosis and treatment of those affected. Patients often face difficulties associated with receiving a formal diagnosis, delayed treatment initiation and limited access to appropriate therapy. This has a considerable impact on the physical health and quality of life for patients, highlighting a need for more education and clearer guidance for HCPs. Support from patient advocacy groups is valuable in helping patients and their families to find appropriate care. However, there remains a need to better understand the burden that SPIGFD has on individuals beyond height, including the impact on physical, emotional, and social wellbeing. CONCLUSIONS: To address the challenges faced by individuals and families affected by SPIGFD, greater awareness of SPIGFD is needed within the healthcare community, and a consensus on best practice in the care of individuals affected by this condition. Continued efforts are also needed at a global level to challenge existing perceptions around SPIGFD, and identify solutions that promote equitable access to appropriate care. Medical writing support was industry-sponsored.

Development of A Minimum Dataset for the Monitoring of Recombinant Human Growth Hormone (rhGH) Therapy Use in Children with Growth Hormone Deficiency (GHD) - A GloBE-Reg Initiative.

Introduction Although there are some recommendations in the literature on the assessments that should be performed in children on recombinant human growth hormone (rhGH) therapy, the level of consensus on these measurements is not clear. The objective of the current study was to identify the minimum dataset (MDS) that could be measured in a routine clinical setting across the world, aiming to minimise burden on clinicians and improve quality of data collection. Methods This study was undertaken by the GH Scientific Study Group (SSG) in GloBE-Reg, a new project that has developed a common registry platform that can support long-term safety and effectiveness studies of drugs. Twelve clinical experts from 7 international endocrine organisations identified by the GloBE-Reg Steering Committee, 2 patient representatives and representatives from 2 pharmaceutical companies with previous GH registry expertise collaborated to develop this recommendation. A comprehensive list of data fields routinely collected by each of the clinical and industry experts for children with GHD was compiled. Each member was asked to determine the: (1) Importance of the data field and (2) Ease of data collection. Data fields that achieved 70% consensus in terms of importance qualified for the MDS, provided <50% deemed the item difficult to collect. Results A total of 246 items were compiled and 27 removed due to redundancies, with 219 items subjected to the grading system. Of the 219 items, 111 achieved at least 70% consensus as important data to collect when monitoring children with GH deficiency (GHD) on rhGH treatment. Sixty-nine of the 219 items were deemed easy to collect. Combining the criteria of importance and ease of data collection, 63 met the criteria for the MDS. Several anomalies to the MDS rule were identified and highlighted for discussion, including whether the patients were involved in current or previous clinical trials, need for HbA1c monitoring, other past medical history, and adherence, enabling formulation of the final MDS recommendation of 43 items; 20 to be completed once, 14 every 6 months and 9 every 12 months. Conclusion In summary, this exercise performed through the GloBE-Reg initiative provides a recommendation of the minimum dataset requirement, collected through real-world data, for the monitoring of safety and effectiveness of rhGH in children with GHD, both for the current daily preparations and the newer long-acting growth hormone.

The effect of age on the quality of life of patients living with stomas: a pilot study.

BACKGROUND: The formation of a stoma is associated with important effects on quality of life (QOL). The objective of this prospective pilot study was to assess the effect of age on QOL after stoma formation. METHODS: Forty-nine patients requiring stoma formation had their QOL assessed by completion of the SF-36 v2 (SF-36v-1999 QualityMetric, Inc.) QOL questionnaire at three time points (preoperatively, at 6 months and at 12 months postoperatively). RESULTS: There was evidence of a difference in mental function according to age (P = 0.05) with younger patients continuing to improve in QOL to 12 months, whereas older patients started higher but any improvement in their QOL over time was too small to be shown. Physical function of QOL improved over time in both younger and older patients, but the pattern of change differed. Older patients started higher but reached their maximum QOL by 6 months, whereas younger patients continued to improve. There was also evidence of a difference related to presentation with acute presentations starting lower in terms of QOL than elective presentations and showing an increase in QOL at both 6 and 12 months. No difference in change in QOL could be shown for type of stoma, sex or reversal. CONCLUSIONS: There was evidence that age and presentation are factors that influence changes in the QOL of stoma patients. Studies have shown that a stoma nurse specialist may help to increase QOL. Therefore, stoma therapy should be targeted at older patients who require more physical and mental support in adjusting to life with a stoma.