Development of multi-generation lower respiratory tract model and insights into the transport and deposition characteristics of inhalable particles.

Airborne particles can spread quickly and enter human respiratory system via inhalation, causing chronic diseases, even cancer. Although recent studies have informed of toxicity of various pollutants, understanding the transport and deposition characteristics of particles in lower respiratory tract is still challenging. The current study proposes a novel model to simulate flow field change from the entrance of lower respiratory tract to pulmonary acinus, while studying particle transport and deposition characteristics. This model for lower respiratory tract with several bronchial extensions containing virtual pulmonary acinus is calculated using computational fluid dynamics and dynamics mesh. The results showed that in the first 10 generations of the lower respiratory tract, vortices and gravity interfered with particles' trajectory, affecting particle deposition distribution. For the first to the tenth-generation respiratory tract, coarse particles were deposited throughout almost the whole respiratory tract model. In contrast, ultrafine particles did not deposit in the higher-generation respiratory tract. The particle enrichment ability of various lobes was uneven with three particle deposition fraction variation patterns. Virtual pulmonary acinus influenced particle deposition and distribution because of vortex ring's trapped ability during expansion and contraction. This new attempt to build a virtual pulmonary acinus model to simulate particle deposition effects in human respiratory system may provide a reference for studying the toxicities of inhalable particles in the exposed human body.

Acinar micromechanics in health and lung injury: what we have learned from quantitative morphology.

Within the pulmonary acini ventilation and blood perfusion are brought together on a huge surface area separated by a very thin blood-gas barrier of tissue components to allow efficient gas exchange. During ventilation pulmonary acini are cyclically subjected to deformations which become manifest in changes of the dimensions of both alveolar and ductal airspaces as well as the interalveolar septa, composed of a dense capillary network and the delicate tissue layer forming the blood-gas barrier. These ventilation-related changes are referred to as micromechanics. In lung diseases, abnormalities in acinar micromechanics can be linked with injurious stresses and strains acting on the blood-gas barrier. The mechanisms by which interalveolar septa and the blood-gas barrier adapt to an increase in alveolar volume have been suggested to include unfolding, stretching, or changes in shape other than stretching and unfolding. Folding results in the formation of pleats in which alveolar epithelium is not exposed to air and parts of the blood-gas barrier are folded on each other. The opening of a collapsed alveolus (recruitment) can be considered as an extreme variant of septal wall unfolding. Alveolar recruitment can be detected with imaging techniques which achieve light microscopic resolution. Unfolding of pleats and stretching of the blood-gas barrier, however, require electron microscopic resolution to identify the basement membrane. While stretching results in an increase of the area of the basement membrane, unfolding of pleats and shape changes do not. Real time visualization of these processes, however, is currently not possible. In this review we provide an overview of septal wall micromechanics with focus on unfolding/folding as well as stretching. At the same time we provide a state-of-the-art design-based stereology methodology to quantify microarchitecture of alveoli and interalveolar septa based on different imaging techniques and design-based stereology.

Fluid-structure interaction analysis of airflow, structural mechanics and aerosol dynamics in a four-generation acinar model.

Elucidating the aerosol dynamics in the pulmonary acinar region is imperative for both health risk assessment and inhalation therapy, especially nowadays with the occurrence of the global COVID-19 pandemic. During respiration, the chest's outward elastic recoil and the lungs' inward elastic recoil lead to a change of transmural pressure, which drives the lungs to expand and contract to inhale and expel airflow and aerosol. In contrast to research using predefined wall motion, we developed a four-generation acinar model and applied an oscillatory pressure on the model outface to generate structure deformation and airflow. With such tools at hand, we performed a computational simulation that addressed both the airflow characteristic, structural mechanics, and aerosol dynamics in the human pulmonary acinar region. Our results showed that there is no recirculating flow in the sac. The structural displacement and stress were found to be positively related to the change of model volume and peaked at the end of inspiration. It was noteworthy that the stress distribution on the acinar wall was significantly heterogeneous, and obvious concentrations of stress were found at the junction of the alveoli and the ducts or the junction of the alveoli and alveoli in the sac. Our result demonstrated the effect of breathing cycles and aerosol diameter on deposition fraction and location of aerosols in the size range of 0.1-5 µm. Multiple respiratory cycles were found necessary for adequate deposition or escape of submicron particles while having a negligible influence on the transport of large particles, which were dominated by gravity. Our study can provide new insights into the further investigation of airflow, structural mechanics, and aerosol dynamics in the acinar depth.

Reassessing the Abundance of miRNAs in the Human Pancreas and Rodent Cell Lines and Its Implication.

miRNAs are critical for pancreas development and function. However, we found that there are discrepancies regarding pancreatic miRNA abundance in published datasets. To obtain a more relevant profile that is closer to the true profile, we profiled small RNAs from human islets cells, acini, and four rodent pancreatic cell lines routinely used in diabetes and pancreatic research using a bias reduction protocol for small RNA sequencing. In contrast to the previous notion that miR-375-3p is the most abundant pancreatic miRNA, we found that miR-148a-3p and miR-7-5p were also abundant in islets. In silico studies using predicted and validated targets of these three miRNAs revealed that they may work cooperatively in endocrine and exocrine cells. Our results also suggest, compared to the most-studied miR-375, that both miR-148a-3p and miR-7-5p may play more critical roles in the human pancreas. Moreover, according to in silico-predicted targets, we found that miR-375-3p had a much broader target spectrum by targeting the coding sequence and the 5' untranslated region, rather than the conventional 3' untranslated region, suggesting additional unexplored roles of miR-375-3p beyond the pancreas. Our study provides a valuable new resource for studying miRNAs in pancreata.

Revisiting the Problem of Finding Effective Sepsis Treatment Solutions.

We studied the effect of a preparation containing ultralow doses of formic aldehyde on the course of experimental sepsis caused by intraperitoneal injection of two different strains of Pseudomonas aeruginosa (1623 and 5266) to C57BL/6 male mice. Microscopy and quantitative bacteriological tests in the dynamics of the infectious process demonstrated a positive effect of the drug: 100% survival of animals, preserved histological structure of the studied organs (lungs, liver, kidneys, spleen, and adrenal glands), a sharp decrease in the level of contamination of the blood and organ homogenates during the first hours after infection, and complete absence of bacteria in inoculates on day 7 after infection. These findings suggest the effectiveness of ultralow doses of formic acid aldehyde in the composition of the medicinal product in the treatment of experimental sepsis caused by P. aeruginosa strains 1623 and 5266 in mice.

Neuronal-epithelial cross-talk drives acinar specification via NRG1-ERBB3-mTORC2 signaling.

Acinar cells are the principal secretory units of multiple exocrine organs. A single-cell, layered, lumenized acinus forms from a large cohort of epithelial progenitors that must initiate and coordinate three cellular programs of acinar specification, namely, lineage progression, secretion, and polarization. Despite this well-known outcome, the mechanism(s) that regulate these complex programs are unknown. Here, we demonstrate that neuronal-epithelial cross-talk drives acinar specification through neuregulin (NRG1)-ERBB3-mTORC2 signaling. Using single-cell and global RNA sequencing of developing murine salivary glands, we identified NRG1-ERBB3 to precisely overlap with acinar specification during gland development. Genetic deletion of Erbb3 prevented cell lineage progression and the establishment of lumenized, secretory acini. Conversely, NRG1 treatment of isolated epithelia was sufficient to recapitulate the development of secretory acini. Mechanistically, we found that NRG1-ERBB3 regulates each developmental program through an mTORC2 signaling pathway. Thus, we reveal that a neuronal-epithelial (NRG1/ERBB3/mTORC2) mechanism orchestrates the creation of functional acini.

In situ Dephosphorylation Assay with Recombinant Nil Phosphatase.

The activity of numerous autophagy-related proteins depends on their phosphorylation status, which places importance on understanding the responsible kinases and phosphatases. Great progress has been made in identifying kinases regulating autophagy, but much less is known about the phosphatases counteracting their function. Genetic screens and modern proteomic approaches provide powerful tools to identify candidate phosphatases, but further experiments are required to assign direct roles for candidates. We have devised a novel protocol to test the role of purified phosphatases in dephosphorylating specific targets in situ . This approach has the potential to visualize context-specific differences in target dephosphorylation that are not easily detected by lysate-based approaches such as Western blots. Graphical abstract.

Liver Acinus Dynamic Chip for Assessment of Drug-Induced Zonal Hepatotoxicity.

Zonation along the liver acinus is considered a key feature of liver physiology. Here, we developed a liver acinus dynamic (LADY) chip that recapitulates a key functional structure of the liver acinus and hepatic zonation. Corresponding to the blood flow from portal triads to the central vein in vivo, gradual flow of oxygen and glucose-carrying culture medium into the HepG2 cell chamber of the LADY chip generated zonal protein expression patterns in periportal (PP) zone 1 and perivenous (PV) zone 3. Higher levels of albumin secretion and urea production were obtained in a HepG2/HUVECs co-culture LADY chip than in HepG2 mono-culture one. Zonal expression of PEPCK as a PP marker and CYP2E1 as a PV marker was successfully generated. Cell death rate of the PV cells was higher than that of the PP cells since zonal factors responsible for metabolic activation of acetaminophen (APAP) were highly expressed in the PV region. We also found the co-culture enhanced metabolic capacity to process APAP, thus improving resistance to APAP toxicity, in comparison with HepG2 mono-culture. These results indicate that our LADY chip successfully represents liver zonation and could be useful in drug development studies as a drug-induced zonal hepatotoxicity testing platform.

Mathematical modeling of pulmonary acinus structure: Verification of acinar shape effects on pathway structure using rat lungs.

The pulmonary acinus is the gas exchange unit in the lung and has a very complex microstructure. The structure model is essential to understand the relationship between structural heterogeneity and mechanical phenomena at the acinus level with computational approaches. We propose an acinus structure model represented by a cluster of truncated octahedra in conical, double-conical, inverted conical, or chestnut-like conical confinement to accommodate recent experimental information of rodent acinar shapes. The basis of the model is the combined use of Voronoi and Delaunay tessellations and the optimization of the ductal tree assuming the number of alveoli and the mean path length as quantities related to gas exchange. Before applying the Voronoi tessellation, controlling the seed coordinates enables us to model acinus with arbitrary shapes. Depending on the acinar shape, the distribution of path length varies. The lengths are more widely spread for the cone acinus, with a bias toward higher values, while most of the lengths for the inverted cone acinus primarily take a similar value. Longer pathways have smaller tortuosity and more generations, and duct length per generation is almost constant irrespective of generation, which agrees well with available experimental data. The pathway structure of cone and chestnut-like cone acini is similar to the surface acini's features reported in experiments. According to space-filling requirements in the lung, other conical acini may also be acceptable. The mathematical acinus structure model with various conical shapes can be a platform for computational studies on regional differences in lung functions along the lung surface, underlying respiratory physiology and pathophysiology.

A model of the pulmonary acinar circulatory system with gas exchange function to explore the influence of alveolar diameter.

Lung diseases such as acute respiratory distress syndrome affect the patient's lung compliance, which in turn affects the ability of gas exchange. Changes in alveolar diameter relate to local lung compliance. How alveolar diameter affects gas exchange, particularly oxygen concentrations in alveolar capillaries, is a topic of concern for researchers, and can be studied using mathematical models. The level of small-scale mathematical models of the pulmonary circulatory system was the alveolar capillaries, but existing models do not consider the gas-exchange function and fail to reflect the influence of alveolar diameter. Therefore, we proposed a pulmonary acinar capillary model with gas exchange function, and most importantly, introduced alveolar diameter into the model, to analyze the effect of alveolar diameter on the gas exchange function of the pulmonary acini. The model was tested by three respiratory function simulation experiments. According to the simulation results of changing diameter, we found that the alveolar diameter mainly affects the alveolar gas exchange function of lung acinar inlets and the middle section compared with the peripheral section.

Micrometer aerosol deposition in normal and emphysematous subacinar models.

Emphysema is a chronic respiratory disease characterized by interalveolar septa destruction and enlarged air sacs. How the inhalation dosimetry in the pulmonary acini varies in the time course of emphysema is still unclear. The aim of this study is to numerically evaluate the impact of septal destructions on particle deposition in a pyramid-shape subacinar model that is composed of 496 alveoli. Four emphysematous models were generated by progressively removing the inter-alveolar septa from the normal geometry. Spatial distribution and temporal evolution of particle deposition were quantified in expanding/contracting subacinar models on both total and regional basis using a well-validated discrete-phase Lagrangian model. Airflow fields in the subacinar cavities are sensitive to the septal raptures, with regular, radial streamlines in the proximal alveoli in the normal geometry in contrast to unsymmetrical and recirculating flows in the emphysematous subacini. Intensified collateral ventilation and significantly increased doses in the outer wall and base are observed in disease than heath. The deposition rate of small particles (1-1.5 µm) is more sensitive to the level of septal rapture than large particles (2.5-3 µm). Unexpectedly, more particles per unit area deposit on the outer wall and at the base of the subacinus than on the inner septal walls. The subacinus-averaged doses increase with progressing septal destructions, suggesting an escalating risk factor to the acinar health at the late stages of emphysema.

[Numerical simulation on the deposition characteristics of inhaled particles in human pulmonary acinus region under the influence of multi-factors].

Research on the deposition of inhaled particles in human pulmonary acinus region is important to the pathogenesis investigation, prevention and treatment of lung diseases. Most of the current research focus on the final deposition fraction of inhaled particles in human acinar region, but little is involved in their dynamic deposition characteristics. In this paper, five multi-alveolar models, G3-G7, were built. The evaluation parameter 1/4 deposition time was introduced to study the particle deposition speed. The deposition characteristics of particles in the diameter ranging 0.1-5 µm were numerically simulated and summarized under the influence of factors such as the generation and structure of model, particle diameter and respiratory mode, shedding some new light on the further research of transport of inhaled particles. The results showed that the generation and structure of model had a significance effect on the deposition of particles. 0.1 µm particles were dominated by Brownian diffusion, which experienced a high deposition fraction, a fast deposition speed and a logarithmic deposition curve, while 5 µm particles were dominated by gravitational sedimentation, with a high deposition fraction, a fast deposition speed and an S-shaped deposition curve. The deposition of 0.3-1 µm particles were influenced greatly by convention and varied with the change of respiratory mode. The research methods and results in this paper can provide theoretical basis and data support for the further exploration of the mechanism, prevention and treatment of lung diseases.

Efficient Therapeutic Delivery by a Novel Cell-Penetrating Peptide Derived from Acinus.

In this study, we have identified a novel cell-penetrating sequence, termed hAP10, from the C-terminus of the human protein Acinus. hAP10 was able to efficiently enter various normal and cancerous cells, likely through an endocytosis pathway, and to deliver an EGFP cargo to the cell interior. Cell penetration of a peptide, hAP10DR, derived from hAP10 by mutation of an aspartic acid residue to an arginine was dramatically increased. Interestingly, a peptide containing a portion of the heptad leucine repeat region domain of the survival protein AAC-11 (residues 377-399) fused to either hAP10 or hAP10DR was able to induce tumor cells, but not normal cells, death both ex vivo on Sézary patients' circulating cells and to inhibit tumor growth in vivo in a sub-cutaneous xenograft mouse model for the Sézary syndrome. Combined, our results indicate that hAP10 and hAP10DR may represent promising vehicles for the in vitro or in vivo delivery of bioactive cargos, with potential use in clinical settings.

Alveolar size effects on nanoparticle deposition in rhythmically expanding-contracting terminal alveolar models.

Significant differences in alveolar size exist in humans of different ages, gender, health, and among different species. The effects of alveolar sizes, as well as the accompanying breathing frequencies, on regional and local dosimetry of inhaled nanoparticles have not been sufficiently studied. Despite a well-accepted qualitative understanding of the advection-diffusion-sedimentation mechanism in the acinar region, a quantitative picture of the interactions among these factors remains inchoate. The objective of this study is to quantify the effects of alveolar size on the regional and local deposition of inhaled nanoparticles in alveolar models of varying complexities and to understand the dynamic interactions among different deposition mechanisms. Three different models were considered that retained 1, 4, and 45 alveoli, respectively. For each model, the baseline geometry was scaled by », ½, 2, 4, and 8 times by volume. Temporal evolution and spatial distribution of particle deposition were tracked using a discrete-phase Lagrangian model. Lower retentions of inhaled nanoparticles were observed in the larger alveoli under the same respiration frequency, while similar retentions were found among different geometrical scales if breathing frequencies allometrically matched the alveolar size. Dimensional analysis reveals a manifold deposition mechanism with tantamount contributions from advection, diffusion, and gravitational sedimentation, each of which can become dominant depending on the location in the alveoli. Results of this study indicate that empirical correlations obtained from one sub-population cannot be directly applied to others, nor can they be simply scaled as a function of the alveolar size or respiration frequency due to the regime-transiting deposition mechanism that is both localized and dynamic.

Ultrastructural studies on the pancreatic acini in duck (Anas boscas) and pigeon (Columba livia).

The aim of this research work was to study the histological structure of the pancreatic acini by transmission electron microscope in two avian species, duck and pigeon. The specimens were collected and processed for electron microscopic study. The results showed that the acini of the two avian species were two types; the first one was an electron dense and the second one an electron lucent. The light acinar cells were larger in size than the dark cells. These cells contained centrally located ovoid nuclei with prominent nucleoli and abundant euchromatin. The cytoplasm was electron lucent, with many rough endoplasmic reticulum, polymorphic mitochondria. Numerous zymogen granules were distributed in the basal part and around the nucleus, so these cells considered active cells. The dark acinar cells were characterized by an electron dense cytoplasm. The most prominent cell organelle in these cells were the zymogen granules that appeared in different sizes while other organelles as mitochondria, and rough endoplasmic reticulum were inconspicuous or few, so these cells were considered as inactive cells. The nucleus with indented nuclear membrane located centrally with prominent nucleoli and abundant heterochromatin. Prominent intercellular spaces between the individual acinar cells, as well as well-developed basement membrane separating the electron dense cells and the lumen contained the secretion between acinar cells. It could be concluded that the acinar cells in ducks and pigeons were divided into two types, that is, light and dark acinar cells which mainly attributed to the activity of these cells.

Yorkie Growth-Promoting Activity Is Limited by Atg1-Mediated Phosphorylation.

The expression of multiple growth-promoting genes is coordinated by the transcriptional co-activator Yorkie with its major regulatory input provided by the Hippo-Warts kinase cascade. Here, we identify Atg1/ULK1-mediated phosphorylation of Yorkie as an additional inhibitory input independent of the Hippo-Warts pathway. Two serine residues in Yorkie, S74 and S97, are Atg1/ULK1 consensus target sites and are phosphorylated by ULK1 in vitro, thereby preventing its binding to Scalloped. In vivo, gain of function of Atg1, or its activator Acinus, caused elevated Yorkie phosphorylation and inhibited Yorkie's growth-promoting activity. Loss of function of Atg1 or Acinus raised expression of Yorkie target genes and increased tissue size. Unlike Atg1's role in autophagy, Atg1-mediated phosphorylation of Yorkie does not require Atg13. Atg1 is activated by starvation and other cellular stressors and therefore can impose temporary stress-induced constraints on the growth-promoting gene networks under the control of Hippo-Yorkie signaling.

Quantitative Proteomics Reveals a Role for SERINE/ARGININE-Rich 45 in Regulating RNA Metabolism and Modulating Transcriptional Suppression via the ASAP Complex in Arabidopsis thaliana.

Pre-mRNA alternative splicing is a conserved mechanism for eukaryotic cells to leverage existing genetic resources to create a diverse pool of protein products. It is regulated in coordination with other events in RNA metabolism such as transcription, polyadenylation, RNA transport, and nonsense-mediated decay via protein networks. SERINE/ARGININE-RICH 45 (SR45) is thought to be a neutral splicing regulator. It is orthologous to a component of the apoptosis and splicing-associated protein (ASAP) complex functioning to regulate RNA metabolism at multiple levels. Within this context, we try to understand why the sr45-1 mutant Arabidopsis has malformed flowers, delayed flowering time, and increased disease resistance. Prior studies revealed increased expression for some disease resistance genes and the flowering suppressor Flowering Locus C (FLC) in sr45-1 mutants and a physical association between SR45 and reproductive process-related RNAs. Here, we used Tandem Mass Tag-based quantitative mass spectrometry to compare the protein abundance from inflorescence between Arabidopsis wild-type (Col-0) and sr45-1 mutant plants. A total of 7,206 proteins were quantified, of which 227 proteins exhibited significantly different accumulation. Only a small percentage of these proteins overlapped with the dataset of RNAs with altered expression. The proteomics results revealed that the sr45-1 mutant had increased amounts of enzymes for glucosinolate biosynthesis which are important for disease resistance. Furthermore, the mutant inflorescence had a drastically reduced amount of the Sin3-associated protein 18 (SAP18), a second ASAP complex component, despite no significant reduction in SAP18 RNA. The third ASAP component protein, ACINUS, also had lower abundance without significant RNA changes in the sr45-1 mutant. To test the effect of SR45 on SAP18, a SAP18-GFP fusion protein was overproduced in transgenic Arabidopsis Col-0 and sr45-1 plants. SAP18-GFP has less accumulation in the nucleus, the site of activity for the ASAP complex, without SR45. Furthermore, transgenic sr45-1 mutants overproducing SAP18-GFP expressed even more FLC and had a more severe flowering delay than non-transgenic sr45-1 mutants. These results suggest that SR45 is required to maintain the wild-type level of SAP18 protein accumulation in the nucleus and that FLC-regulated flowering time is regulated by the correct expression and localization of the ASAP complex.

Targeting inhaled fibers to the pulmonary acinus: Opportunities for augmented delivery from in silico simulations.

Non-spherical particles, and fibers in particular, are potentially attractive airborne carriers for pulmonary drug delivery. Not only do they exhibit a high surface-to-volume ratio relative to spherical aerosols, but their aerodynamic properties also enable them to reach deep into the lungs. Until present, however, our understanding of the deposition characteristics of inhaled aerosols in the distal acinar lung regions has been mostly limited to spheres. To shed light on the fate of elongated aerosols in the pulmonary depths, we explore through in silico numerical simulations the deposition and dispersion characteristics of ellipsoid-shaped fibers in a physiologically-realistic acinar geometry under oscillatory breathing flow conditions mimicking various inhalation maneuvers. The transient translation and rotational movement of micron-sized elongated particles under drag, lift, and gravitational forces are simulated as a function of size (dp) and aspect ratio (AR). Our findings underscore how acinar deposition characteristics are intimately linked to the geometrical combination of dp and AR under oscillatory flow conditions. Surprisingly, the elongation of the traditionally recommended size range of spherical particles (i.e., 2-3 µm) for acinar deposition may lead to a decrease in deposition efficiency and dispersion. Instead, our findings advocate how elongating particles (i.e., high AR) in the larger size range of 4-6 µm might be leveraged for improved targeted deposition to the acinar regions. Together, these results point to new windows of opportunities in selecting the shape and size of micron-sized fibers for targeted pulmonary deposition. Such in silico efforts represent an essential stepping stone in further exploring aerosol drug carrier designs for inhalation therapy to the deep lungs.

[Progress on numerical simulation of the deposition of inhaled particles in human pulmonary acinus region].

The inhalation and deposition of particles in human pulmonary acinus region can cause lung diseases. Numerical simulation of the deposition of inhaled particles in the pulmonary acinus region has offered an effective gateway to the prevention and clinical treatment of these diseases. Based on some important affecting factors such as pulmonary acinar models, model motion, breathing patterns, particulate characteristics, lung diseases and ages, the present research results of numerical simulation in human pulmonary acinus region were summarized and analyzed, and the future development directions were put forward in this paper, providing new insights into the further research and application of the numerical simulation in the pulmonary acinus region.

The arginine and serine-rich domains of Acinus modulate splicing.

Apoptotic chromatin condensation inducer in the nucleus (Acinus) is an RNA-binding protein that has a functional role in inducing apoptotic chromatin condensation and regulating messenger RNA (mRNA) processing. Acinus interacts with the spliceosomal machinery and is a member of the ASAP (apoptosis and splicing-associated protein complex) as well as the EJC (exon junction complex), which gets deposited onto mRNA during splicing. In this study, we have used in vivo splicing assays to characterize the function of Acinus in pre-mRNA splicing more closely. We show that full-length Acinus-S', an isoform of Acinus, does not have a role in modulating splice site selection in human immunodeficiency virus 1 minigene reporter system. In contrast, we observed that the tethering of arginine/serine (RS) and RNPS1-SAP18-binding (RSB) domains of Acinus could regulate the selection of alternative splice sites, thereby revealing the potential of Acinus in stimulating alternative splicing. Altogether, our data suggest that the RS and RSB domains play a critical role in regulating splicing activity via selection of distinct splice sites during pre-mRNA splicing.