Res Sq [Preprint]. 2024 Nov 21:rs.3.rs-5479740. doi: 10.21203/rs.3.rs-5479740/v1.

ABSTRACT

Traditionally, the thermostability of a protein is defined by a melting temperature, at which half of the protein is unfolded. However, this definition cannot indicate the structural origin of a heat-induced unfolding pathway. Here, the thermoring structures were studied on the ATP-dependent heat-induced unfolding of the first nucleotide-binding domain from the human cystic fibrosis transmembrane conductance regulator. The results showed that initial theoretical and experimental melting thresholds aligned well after three structural perturbations including the F508del mutation, the most common cause of cystic fibrosis. This alignment further demonstrated that the heat-induced unfolding process began with the disruption of the least-stable noncovalent interaction within the biggest thermoring along the single peptide chain. The C-terminal region, which was related to the least-stable noncovalent interaction and the ATP-dependent dimerization of two nucleotide-binding domains, emerged as a crucial determinant of the thermal stability of the isolated protein and a potential interfacial drug target to alleviate the thermal defect caused by the F508del mutation. This groundbreaking discovery significantly advances our understanding of protein activity, thermal stability, and molecular pathology.

PMID:39606474 | PMC:PMC11601864 | DOI:10.21203/rs.3.rs-5479740/v1

Res Sq [Preprint]. 2024 Nov 12:rs.3.rs-5391023. doi: 10.21203/rs.3.rs-5391023/v1.

ABSTRACT

Antibiotic resistance is one of the most pressing threats to human health, yet recent work highlights how loss of resistance may also drive pathogenesis in some bacteria. In two recent studies, we found that β-lactam antibiotic and nutrient stresses faced during infection selected for the genetic inactivation of the Pseudomonas aeruginosa (Pa) antibiotic efflux pump mexEFoprN. Unexpectedly, efflux pump mutations increased Pa virulence during infection; however, neither the prevalence of efflux pump inactivating mutations in real human infections, nor the mechanisms driving increased virulence of efflux pump mutants are known. We hypothesized that human infection would select for efflux pump mutations that drive increased virulence in Pa clinical isolates. Using genome sequencing of hundreds of Pa clinical isolates, we show that mexEFoprN efflux pump inactivating mutations are enriched in Pa cystic fibrosis isolates relative to Pa intensive care unit clinical isolates. Combining RNA-seq, metabolomics, genetic approaches, and infection models we show that efflux pump mutants have elevated expression of two key Pa virulence factors, elastase and rhamnolipids, which increased Pa virulence and lung damage during both acute and chronic infections. Increased virulence factor production was driven by higher Pseudomonas quinolone signal levels in the efflux pump mutants. Finally, genetic restoration of the efflux pump in a representative ICU clinical isolate and the notorious CF Pa Liverpool epidemic strain reduced their virulence. Together, our findings suggest that mutations inactivating antibiotic resistance mechanisms could lead to greater patient mortality and morbidity.

PMID:39606469 | PMC:PMC11601840 | DOI:10.21203/rs.3.rs-5391023/v1

Res Sq [Preprint]. 2024 Nov 13:rs.3.rs-5195002. doi: 10.21203/rs.3.rs-5195002/v1.

ABSTRACT

Background Clinical trials in cystic fibrosis (CF) disproportionately over-represent non-Hispanic, White individuals. Barriers and facilitators to enrolling racially and ethnically minoritized individuals with CF are not well understood. This study explored research coordinator (RC) perspectives on recruitment and enrollment of minoritized people with CF (PwCF) into clinical trials. Methods A cross-sectional survey was administered to RCs in the CF Therapeutics Development Network (CF TDN), eliciting perceived barriers and facilitators to inclusion of minoritized PwCF in clinical research. Respondents were categorized based on their self-reported experience discussing and successfully enrolling minoritized PwCF into clinical trials. Results Among 48 respondents, the majority (n = 31, 64%) had little to no experience discussing CF clinical trials with minoritized PwCF. Respondents who had a moderate or great deal of experience identified that having a trusted clinical team member first introduce the study to PwCF as the most common strategy for recruitment. Experienced respondents also identified the importance of having team members who speak the same language or are the same culture as the minoritized PwCF. Respondents who had little to no experience successfully enrolling minoritized PwCF into clinical trials cited low numbers of minoritized patients at their study center as an important consideration. All respondents emphasized language barriers in enrollment including need for adequate translation services and printed materials in the PwCF's primary language. Conclusion Our study identified modifiable barriers that may be addressed at the level of trial design and study center.

PMID:39606463 | PMC:PMC11601850 | DOI:10.21203/rs.3.rs-5195002/v1

Front Immunol. 2024 Nov 13;15:1397886. doi: 10.3389/fimmu.2024.1397886. eCollection 2024.

ABSTRACT

INTRODUCTION: Cystic fibrosis (CF) is a hereditary autosomal recessive disease driven by deleterious variants of the CFTR gene, leading, among other symptoms, to increased lung infection susceptibility. Mucus accumulation in the CF lung is, as of yet, considered as one important factor contributing to its colonization by opportunistic pathogens such as Pseudomonas aeruginosa. However, in recent years evidence was provided that alveolar macrophages, which form the first line of defense against airborne pathogens, seem to be intrinsically defective with regard to bactericidal functionality in the CF lung. To assess the impact of CFTR deficiency in human macrophages only insufficient systems are available.

METHODS: To address this problem and to evaluate the role of CFTR in human macrophages, we successfully differentiated human induced pluripotent stem cells (iPSC) from a CF p.Phe508del homozygous individual and a healthy donor into primitive macrophages (iMacΔF508 and iMacWT), respectively, and compared the bactericidal functionality in the relevant cell type.

RESULTS: iMacΔF508 showed impaired P. aeruginosa clearance and intracellular killing capacity in comparison to iMacWT. Furthermore, iMacΔF508 exhibited a less acidic lysosomal pH, and upon P. aeruginosa infection, there were signs of mitochondrial fragmentation and autophagosome formation together with a hyperinflammatory phenotype and deficient type I interferon response.

CONCLUSION: In summary, we present a defective phenotype in iMacΔF508 upon P. aeruginosa infection, which will constitute an ideal platform to further study the role of macrophages in the context of CF.

PMID:39606224 | PMC:PMC11601075 | DOI:10.3389/fimmu.2024.1397886

eGastroenterology. 2024 Nov;2(4):e100102. doi: 10.1136/egastro-2024-100102. Epub 2024 Nov 10.

ABSTRACT

BACKGROUND: Cystic fibrosis (CF) is an autosomal recessive genetic disorder caused by loss-of-function mutations in the CF transmembrane conductance regulator gene. CF-related pancreatic lesions are known to cause exocrine dysfunctions such as pancreatic insufficiency, and endocrine dysfunctions, including CF related diabetes. In a previous study, we generated CF rabbits using CRISPR/Cas9-mediated gene editing.

METHODS: CF rabbits were subjected to histological analysis with a focus on CF associated pancreatic lesions. Endocrine function related assays were conducted to evaluate CF related pancreatic endocrine disorders in these animals.

RESULTS: We report that CF rabbits develop spontaneous pancreatic lesions at a young age, characterised by pancreatic inflammation and fibrosis, vacuolar degeneration, epithelium mucus-secretory cell metaplasia, and pancreatic duct dilation. The size of the pancreatic islets in the CF rabbits is significantly smaller than that of the wild type animals. Consistent with these pathological findings, young CF rabbits exhibited signs of pancreatic endocrine related disorders such as lower insulin levels and impaired glucose metabolism.

CONCLUSIONS: Our results suggest that the CF rabbit could serve as a valuable model for translational research on CF related pancreatic endocrine dysfunction.

PMID:39605883 | PMC:PMC11594368 | DOI:10.1136/egastro-2024-100102

bioRxiv [Preprint]. 2024 Nov 21:2024.11.20.624197. doi: 10.1101/2024.11.20.624197.

ABSTRACT

Trikafta is well-known for correcting the thermal and gating defects caused by the most common cystic fibrosis mutation F508del in the human cystic fibrosis transmembrane conductance regulator even at physiological temperature. However, the exact pathway is still unclear. Here, the noncovalent interactions among two transmembrane domains (TMD 1 and TMD2), the regulatory (R) domain and two nucleotide binding domains (NBD1 and NBD2), along with the thermoring structures of NBD1, were analyzed around the active gating center. The results demonstrated that Trikafta binding to TMD1 and TMD2 rearranged their interactions with the R domain, releasing the C-terminal region from NBD1 for its tight ATP-dependent dimerization with NBD2, stabilizing NBD1. Taken together, although the deletion of F508 induces the primary thermal defect of NBD1 and then the gating defect at the TMD1-TMD2 interface, Trikafta rescued them in a reverse manner allosterically. Thus, the thermoring structure can be used to uncover the pathway of a drug to correct the thermal defect of health-related protein.

SIGNIFICANCE: Trikafta modulators have been approved by the FDA to treat the most common cystic fibrosis- causing mutation F508del CFTR. However, the molecular action mechanisms of these modulators are still unknown. Following the identification of the gating center in CFTR, this study further revealed that the specific noncovalent interactions of the phosphorylated S813 site with cytoplasmic loops 1 and 4 and N-/C- terminal tails of TMD1 upon Trikafta-triggered tight TMD1- TMD2 interactions at the gating center play a pivotal role in rescuing the primary gating defect and then the thermal defect of F508del CFTR.

HIGHLIGHTS: Trikafta strengthened TMD1-TMD2 interactions at the gating center of ΔF508-CFTR Tight TMD1-TMD2 interactions allowed specific interactions of the R domain with the ICL1- ICL4 interface and the N-/C- terminal tails of TMD1 Subsequently, the C-terminal region was released from NBD1 for tight ATP-dependent NBD1-NBD2 dimerization, stabilizing NBD1 of ΔF508-CFTR.

PMID:39605627 | PMC:PMC11601583 | DOI:10.1101/2024.11.20.624197

bioRxiv [Preprint]. 2024 Nov 13:2024.11.12.622518. doi: 10.1101/2024.11.12.622518.

ABSTRACT

Transplantation of airway basal stem cells could achieve a durable cure for genetic diseases of the airway, such as cystic fibrosis and primary ciliary dyskinesia. Recent work demonstrated the potential of primary- and pluripotent stem cell (PSC)-derived basal cells to efficiently engrai into the mouse trachea aier injury. However, there are many hurdles to overcome in translating these approaches to humans including developing safe and efficient methods for delivery in larger animal models. We propose a model which targets preconditioning and cell-delivery to the intrapulmonary airways utilizing a micro- bronchoscope for delivery. The detergent polidocanol was adapted for distal lung pre-conditioning, inducing intrapulmonary airway epithelial denudation by 5 and 24-hours post-delivery. While initial re- epithelialization of airways occurred later than tracheas, complete repair was observed within 7-days. Both PSC-derived and primary basal cells delivered via micro-bronchoscope post-polidocanol injury engraied in tracheas and intrapulmonary airways, respectively. Transplanted cells differentiated into ciliated and secretory lineages while maintaining a population of basal cells. These findings demonstrate the utility of bronchoscopically targeted pre-conditioning and cell delivery to the conducting intra- pulmonary airways, providing an important framework for pre-clinical translation of approaches for engineered airway epithelial regeneration.

PMID:39605510 | PMC:PMC11601482 | DOI:10.1101/2024.11.12.622518

bioRxiv [Preprint]. 2024 Nov 16:2024.11.16.623925. doi: 10.1101/2024.11.16.623925.

ABSTRACT

Chronic bacterial infections are often polymicrobial, comprising multiple bacterial species or variants of the same species. Because chronic infections may last for decades, they have the potential to generate high levels of intraspecific variation through within-host diversification over time, and the potential for superinfections to occur through the introduction of multiple pathogen populations to the ongoing infection. Traditional methods for identifying infective agents generally involve isolating one single colony from a given sample, usually after selecting for a specific pathogen or antibiotic resistance profile. Isolating a recognized virulent or difficult to treat pathogen is an important part of informing clinical treatment and correlative research; however, these reductive methods alone, do not provide researchers or healthcare providers with the potentially important perspective on the true pathogen population structure and dynamics over time. To begin to address this limitation, in this study, we compare findings on Staphylococcus aureus single colonies versus and pools of colonies taken from fresh sputum samples from three patients with cystic fibrosis to isolates collected from the same sputum samples and processed by the clinical microbiology laboratory. Phenotypic and genotypic analysis of isolated S. aureus populations revealed coexisting lineages in two of three sputum samples as well as population structures that were not reflected in the single colony isolates. Altogether, our observations presented here demonstrate that clinically relevant diversity can be missed with standard sampling methods when assessing chronic infections. More broadly, this work outlines the potential impact that comprehensive population-level sampling may have for both research efforts and more effective treatment practices.

DATA SUMMARY: The authors confirm all supporting data, code and protocols have been provided within the article.

PMID:39605409 | PMC:PMC11601312 | DOI:10.1101/2024.11.16.623925

bioRxiv [Preprint]. 2024 Nov 20:2024.11.18.623945. doi: 10.1101/2024.11.18.623945.

ABSTRACT

INTRODUCTION: Over 1 million people have chronic pulmonary aspergillosis (CPA) secondary to pulmonary tuberculosis. Additionally, Aspergillus fumigatus ( Af ) has been reported as one of the most common pathogens associated with mycobacteria in patients with cystic fibrosis. Mycobacterial virulence factors, like lipoarabinomannan, have been shown to interfere with host's intracellular pathways required for an effective immune response, however, the immunological basis for mycobacterial-fungal coinfection is still unknown. We therefore investigated the effect of lipoarabinomannan on macrophage responses against Af .

METHODS: Bone marrow-derived macrophages (BMDMs) were stimulated with non-mannose-capped lipoarabinomannan (LAM) from Mycobacterium smegmatis or mannose-capped lipoarabinomannan (ManLAM) from Mycobacterium tuberculosis for 2 hours and then infected with swollen Af conidia. Cell death was assessed by lactate dehydrogenase release. Cytokine release was measured in supernatant using Enzyme Linked Immuno-Sorbent Assay (ELISA). Colony forming units counting and time-lapse fluorescence microscopy was performed for studying conidia killing by macrophages.

RESULTS: BMDMs stimulated with LAM showed increased cell death and inflammatory cytokine release in a dose-dependent manner, characterised by a significant increase of IL-1β release. Time-lapse fluorescence microscopy and CFUs revealed that both LAM and ManLAM significantly decrease the capacity of macrophages to kill Af conidia within the first 6 hours of infection.

CONCLUSIONS: The mycobacterial virulence factor, lipoarabinomannan, disrupts macrophage capacity to efficiently clear Af at early stages of infection in-vitro .

PMID:39605324 | PMC:PMC11601501 | DOI:10.1101/2024.11.18.623945

Protein Sci. 2024 Dec;33(12):e5227. doi: 10.1002/pro.5227.

ABSTRACT

Human neutrophil elastase (hNE), a serine protease released by neutrophils during inflammation, plays a major role in the pathophysiology of several conditions especially in inflammatory lung diseases. Its inhibition constitutes, therefore, a promising therapeutic strategy to combat these diseases. In this work, we characterized the in vitro properties of a VHH (i.e., the antigen binding domain of camelid heavy chain-only antibodies), referred to as NbE201. This VHH is able to inhibit tightly, selectively and competitively both human and murine elastases with the inhibition constants (Ki) of 4.1 ± 0.9 nM and 36.8 ± 3.9 nM, respectively. The IC50 for the inhibition of the hydrolysis of elastin is in the same range to that of alpha-1 antitrypsin (i.e., the main endogenous inhibitor of hNE also used in the clinic) and 14 times better than that of Sivelestat (i.e., the 2nd clinically approved hNE inhibitor). The X-ray crystal structure of the NbE201-hNE complex reveals that the Complementarity Determining Regions CDR1 and CDR3 of the VHH bind into the substrate binding pocket of hNE and prevent the access to small or macromolecular substrates. They do not, however, bind deep enough into the pocket to be hydrolyzed. NbE201 is highly stable towards oxidation, deamidation, and chemical or thermal denaturation. NbE201 is therefore likely to tolerate manufacturing processes during drug development. These results highlight the high potential of NbE201 as a (pre)clinical tool to diagnose and treat diseases associated with excessive hNE activity, and for fundamental research to better understand the role of hNE in these conditions.

PMID:39604162 | DOI:10.1002/pro.5227

Eur Respir J. 2024 Nov 27:2401574. doi: 10.1183/13993003.01574-2024. Online ahead of print.

ABSTRACT

BACKGROUND: Macrolide maintenance therapy (MMT) has demonstrated notable efficacy in reducing exacerbation in patients with bronchiectasis, which is a major risk factor for cardiovascular events. However, a comprehensive assessment of the cardiovascular benefits and safety profile of MMT in this population is lacking.

METHODS: This territory-wide cohort study analyzed patients diagnosed with bronchiectasis in Hong Kong between 2001 and 2018. Patients were classified as MMT receivers or macrolide non-receivers based on the administration of MMT. Propensity score (PS) matching was employed for confounding factors adjustment. The primary outcome of interest was major adverse cardiovascular events (MACE), a composite of cardiovascular death, myocardial infarction and stroke. The safety outcome was the occurrence of ventricular arrhythmias or sudden cardiac death. Cox proportional hazard regression analysis was utilized to compare the incidence of outcomes across the two groups.

RESULTS: A total of 22 895 patients with bronchiectasis were identified. Following 1:2 PS matching, the final cohort consisted of 3137 individuals, with 1123 MMT receivers and 2014 macrolide non-receivers. MMT administration was associated with a significant reduced risk of MACE (16.38 versus 24.11 events per 1000 person years; HR 0.68; 95% CI 0.52-0.90). Importantly, the use of MMT was not associated with elevated risk of ventricular arrhythmias or sudden cardiac death (7.17 versus 7.67 events per 1000 person years; HR 0.93; 95% CI 0.60-1.44).

CONCLUSIONS: The administration of MMT in patients with bronchiectasis was associated with a significant reduction in the risk of MACE, without any evidence suggesting an increased risk of severe arrhythmia-related adverse events.

PMID:39603670 | DOI:10.1183/13993003.01574-2024

Horm Res Paediatr. 2024 Nov 27:1-12. doi: 10.1159/000542786. Online ahead of print.

ABSTRACT

INTRODUCTION: Cystic fibrosis-related diabetes (CFRD) is linked to worsening pulmonary function and increased mortality in people with cystic fibrosis (pwCF). Because of this correlation, early recognition of CFRD is important. Current recommendations for detecting glucose abnormalities using an oral glucose tolerance test (OGTT) can be difficult to achieve in a busy clinical setting.

METHODS: We trialed a 10-day CGM (Dexcom G6Pro) session in patients seen in our pediatric CF clinic that could not do an OGTT (reasons include emesis with OGTT or fear of needles) or that had an abnormal OGTT (to gather additional data to make treatment decisions).

RESULTS: Of the 36 sensors placed, 34 (94%) were returned. Devices were worn for a median of 10 days (range 4-10 days). Of the 34 CGMs returned, 20 (59%) met the criterion for referral to a pediatric endocrinologist.

CONCLUSION: CGM placement is feasible in a busy CF clinic to detect glucose abnormalities in children.

PMID:39602887 | DOI:10.1159/000542786

Children (Basel). 2024 Nov 18;11(11):1396. doi: 10.3390/children11111396.

ABSTRACT

BACKGROUND/OBJECTIVES: Hope is a universal, multidimensional, and nuanced concept that may have specific meaning for young people living with chronic health conditions anticipated to last into adulthood. We previously identified definitions of hope for youth living with chronic health conditions derived from young people's and their caregivers' own words. Here, we aimed to develop a hope assessment tool to facilitate the future evaluation of interventions to support wellness and health for young people growing up with chronic health conditions; Methods: We developed Likert-type scale questions using the young people's and caregivers' definitions of hope and applied the think-aloud cognitive interview method to assess understanding and to inform sequential iteration. Interviews were recorded and insights from participant interviews were analyzed thematically.

RESULTS: In total, 11 youth (age 12-16 years) with various chronic health conditions completed surveys and interviews over three iteration cycles. Responses to the six-point Likert-scale questions ranged from 1 (none of the time) to 6 (all of the time) (median 5). All of the young people (n = 11) reported that they do things they enjoy, either all of the time or most of the time. In contrast, only 36% felt energetic, either all or most of the time. Three themes were identified: my body and hope; my identity, self-image, and hope; and my world and hope.

CONCLUSIONS: In addition to gaining important feedback that allowed us to improve item word choice to maximize assessment tool understanding, we gained valuable insights into the multidimensional construct of hope. Thematic analysis revealed the importance of physical symptoms and identity to the meaning of hope in the context of a young person's life. Our new hope assessment tool derived from the young people's own definition of hope has face and content validity for use in clinical and research settings to evaluate hope among pediatric patients living with chronic health conditions.

PMID:39594971 | PMC:PMC11593005 | DOI:10.3390/children11111396

Pediatr Pulmonol. 2024 Nov 27:e27424. doi: 10.1002/ppul.27424. Online ahead of print.

ABSTRACT

A 17-year-old female with cystic fibrosis was diagnosed with alpha-gal syndrome after 6 months of progressively worsening abdominal pain. Alpha-gal is an acquired allergy to mammalian products that can impact what individuals are able to consume. This syndrome may impact an individual's ability to take medications that are animal derived or contain animal derived excipients. Through dietary removal of mammalian products, she was able to see a reduction in gastrointestinal symptoms. With the success of dietary modifications and modification of her allergy medication formulation, maintenance medication changes did not need to be made. This allowed her to continue her pre-established medication treatment plan (including exogenous pancreatic enzymes). This case highlights potential complexities in medication management of alpha-gal syndrome in an adolescent with cystic fibrosis.

PMID:39601456 | DOI:10.1002/ppul.27424

Am J Physiol Lung Cell Mol Physiol. 2024 Nov 27. doi: 10.1152/ajplung.00137.2024. Online ahead of print.

ABSTRACT

The epithelial lining of luminal organs provides an immune barrier against external factors and regulates transport of nutrients, ions, and water into the body. Several conditions are associated with a breakdown or dysfunction of the epithelial lining. Short circuit current (Isc) measurement using a bulky, expensive, and hard to deploy system known as the Ussing chamber is the gold standard for evaluation of epithelial transport function but requires tissue excision. We demonstrated the ability of the Isc probe to measure Isc in normal wild type (WT) versus reduced CFTR function knockout (KO) rats as a relevant animal model for testing ion channel function.

PMID:39601216 | DOI:10.1152/ajplung.00137.2024

Nutrients. 2024 Nov 19;16(22):3957. doi: 10.3390/nu16223957.

ABSTRACT

INTRODUCTION: Cystic fibrosis transmembrane conductance regulator (CFTR) modulators have greatly improved outcomes in persons with CF (pwCF); however, there is still significant heterogeneity in clinical responses, particularly with regard to respiratory infection and inflammation. Exogenous administration of ketones has profound systemic anti-inflammatory effects and produces several nutrient-signaling and metabolic effects that may benefit multiple organ systems affected in pwCF. This pilot study was designed to determine the feasibility of administration of a ketone monoester (KME) to increase circulating D-beta hydroxybutyrate concentrations (D-βHB) and to improve subjective measures of CF-specific quality of life and markers of inflammation in serum and sputum in adults with CF.

METHODS: Fourteen participants receiving modulator therapy were randomized to receive either KME (n = 9) or placebo control (PC, n = 5) for 5-7 days during hospitalization for treatment of acute pulmonary exacerbation or as outpatients under standard care.

RESULTS: The KME was well tolerated, with only mild reports of gastrointestinal distress. D-βHB concentrations increased from 0.2 ± 0.1 mM to 1.6 ± 0.6 mM in the KME group compared to 0.2 ± 0.0 to 0.3 ± 0.1 in the PC group (p = 0.011) within 15 min following consumption and remained elevated, relative to baseline, for over 2 h. Pulmonary function was not altered after single- or short-term KME administration, but participants in the KME group self-reported higher subjective respiratory scores compared to PC in both cases (p = 0.031). Plasma inflammatory markers were not statistically different between groups following the short-term (5-7 d) intervention (p > 0.05). However, an exploratory analysis of plasma pre- and post-IL-6 concentrations was significant (p = 0.028) in the KME group but not PC. Sputum IFNγ (p = 0.057), IL-12p70 (p = 0.057), IL-1β (p = 0.100), IL-15 (p = 0.057), IL-1α (p = 0.114), and MPO (p = 0.133) were lower in the KME group compared to PC but did not achieve statistical significance.

CONCLUSIONS: With the emerging role of exogenous ketones as nutrient signaling molecules and mediators of metabolism, we showed that KME is well tolerated, increases circulating D-βHB concentrations, and produces outcomes that justify the need for large-scale clinical trials to investigate the role of KME on whole-body and tissue lipid accumulation and inflammation in pwCF.

PMID:39599743 | DOI:10.3390/nu16223957

Nutrients. 2024 Nov 11;16(22):3850. doi: 10.3390/nu16223850.

ABSTRACT

OBJECTIVES/BACKGROUND: Vitamin D-binding protein (VDBP) and free vitamin D are new markers that are being studied as a possible markers of vitamin D status. The main aim of our study was to analyze the VDBP genotype and quantify the levels of free vitamin D in a sample of cystic fibrosis (CF) patients.

METHODS: We conducted a multicenter, cross-sectional, and prospective study including patients with CF and exocrine pancreatic insufficiency who were clinically stable. We investigated vitamin D levels (total and free) and the different VDBP haplotypes. Free vitamin D levels were measured using an electro-chemiluminescence assay.

RESULTS: A sample of 48 patients was obtained (52% male; median age 13.8 years). The most common allele of VDBP was Gc1s (72%) > Gc2 (52%) > Gc1f (27%). The median calcidiol was 21.2 ng/mL (IR 15.3-26.9), and 81% had levels in the insufficiency range: 23 patients (48%) below 20 ng/mL, and 16 (33%) between 20 and 30 ng/mL. The median free vitamin D level was 4.2 pg/mL (IR 3.9-5.6). A positive correlation was observed between calcidiol and free vitamin D levels (r = 0.871; p < 0.0001). After adjustment for season, vitamin D supplementation, sex, and CF-related diabetes, patients with Gc1f polymorphism had a lower risk of vitamin D deficiency, OR 0.22 (95% CI 0.05-0.99), and p = 0.027. A negative linear trend was observed between the polymorphisms grouped into three categories (Gc1/Gc1, Gc1/Gc2, and Gc2/Gc2, in that order) and vitamin D and free vitamin D levels (p = 0.025 and p = 0.033, respectively).

CONCLUSION: In CF, as in the general population, the most common VDBP haplotype in the Caucasian race is Gc1s. VDBP polymorphisms influence serum vitamin D and free vitamin D levels in CF patients. There is a good correlation between free vitamin D and calcidiol levels, suggesting that measuring the latter in CF does not seem to provide any additional benefit.

PMID:39599636 | DOI:10.3390/nu16223850

Pharmaceutics. 2024 Nov 17;16(11):1465. doi: 10.3390/pharmaceutics16111465.

ABSTRACT

Background: Cystic fibrosis is a hereditary disease, which causes the accumulation of dense mucus in the lungs accompanied by frequent local inflammation. The non-steroidal anti-inflammatory drug ibuprofen (IBU) and the mucolytic mannitol (MAN) can treat these symptoms. Compared to per os administration, a lower dose of these drugs is sufficient to achieve the desired effect by delivering them in a pulmonary manner. However, it is still a challenge to administer high drug doses to the lungs. We aim to develop two inhaled powder formulations, a single-drug product of MAN and a combined formulation containing IBU and MAN. Methods: MAN was dissolved in an aqueous solution of Poloxamer-188 (POL). In the case of the combined formulation, a suspension was first prepared in a planetary mill via wet milling in POL medium. After the addition of leucine (LEU), the formulations were spray-dried. The prepared DPI samples were analyzed by using laser diffraction, scanning electron microscopy, powder X-ray diffraction, differential scanning calorimetry, density tests, in vitro aerodynamic studies (Andersen Cascade Impactor, Spraytec® device), in vitro dissolution tests in artificial lung fluid, and in silico tests with stochastic lung model. Results: The DPIs showed suitability for inhalation with low-density spherical particles of appropriate size. The LEU-containing systems were characterized by high lung deposition and adequate aerodynamic diameter. The amorphization during the procedures resulted in rapid drug release. Conclusions: We have successfully produced a single-drug formulation and an innovative combination formulation, which could provide complex treatment for patients with cystic fibrosis to improve their quality of life.

PMID:39598588 | DOI:10.3390/pharmaceutics16111465

Pharmaceutics. 2024 Oct 24;16(11):1361. doi: 10.3390/pharmaceutics16111361.

ABSTRACT

BACKGROUND/OBJECTIVES: Research on cyclodextrin-based metal-organic frameworks (CD-MOFs) is still in its infancy, but their potential for use in drug delivery-expressly in the lung-seems promising. We aimed to use the freeze-drying method to create a novel approach for preparing CD-MOFs. MOFs consisting of γ-cyclodextrin (γCD) and potassium cations (K+) were employed to encapsulate the poorly water-soluble model drug Ibuprofen (IBU) for the treatment of cystic fibrosis (CF).

METHODS: Using the LeanQbD® software (v2022), we designed the experiments based on the Quality by Design (QbD) concept. According to QbD, we identified the three most critical factors, which were the molar ratio of the IBU to the γCD, incubation time, and the percentage of the organic solvent. light-, scanning electron microscope (SEM) and laser diffraction were utilized to observe the morphology and particle size of the samples. In addition, the products were characterized by Differential Scanning Calorimetry (DSC), X-ray Powder Diffraction (XRPD), Fourier Transform Infrared Spectroscopy (FT-IR) and nuclear magnetic resonance spectroscopy (NMR).

RESULTS: Based on characterizations, we concluded that a γCD-MOF/IBU complex was also formed using the freeze-drying method. Using formulations with optimal aerodynamic properties, we achieved 38.10 ± 5.06 and 47.18 ± 4.18 Fine Particle Fraction% (FPF%) based on the Andersen Cascade Impactor measurement. With these formulations, we achieved a fast dissolution profile and increased IBU solubility.

CONCLUSIONS: This research successfully demonstrates the innovative use of freeze-drying to produce γCD-MOFs for inhalable IBU delivery. The method enabled to modify the particle size, which was crucial for successful pulmonary intake, emphasizing the need for further investigation of these formulations as effective delivery systems.

PMID:39598485 | DOI:10.3390/pharmaceutics16111361

Life (Basel). 2024 Nov 13;14(11):1474. doi: 10.3390/life14111474.

ABSTRACT

The gold standard for histological acute cellular rejection diagnosis is transbronchial forceps biopsy (FB), but in recent years, transbronchial cryobiopsy (CB) has been increasingly used. This study aims to compare the diagnostic rate and safety of FBs and CBs performed in two different periods. We retrospectively reviewed our case history for the two biopsy procedures: 251 FBs (223 for surveillance purposes and 28 for clinical indication) and 218 consecutive CBs (159 for surveillance purposes and 59 for clinical indication). All biopsies were scored according to the ISHLT criteria. Diagnostic yield was higher in the CB group for all the parameters considered: a grade of acute rejection (AR) was detected in 95.0% vs. 84.5% in the CB vs. FB groups (p < 0.001). The diagnostic rate of airway inflammation was 65.1% vs. 51.8% (p = 0.005), and 89.0% vs. 64.9% (p < 0.001) for chronic rejection. Pneumothorax requiring chest drainage occurred in 4% of the CB group and 3% of the FB group. Moderate and severe bleeding complicated CB and FB procedures in seven (3%) and three cases (1%), respectively. Transbronchial cryobiopsies improved the diagnostic yield in the monitoring of the lung allograft. The complication rate did not increase significantly in CBs vs. FBs.

PMID:39598272 | DOI:10.3390/life14111474