Alveolar Regeneration Research Articles

Fluid Dynamic Transcrestal Sinus Floor Elevation Using a New Surgical Instrument, Flusilift and Hyaluronic Acid as Only Biomaterial: A Case Report

Despite validated surgical techniques and the development of biomaterials, the procedures aimed at increasing the maxillary bone volume by sinus floor elevation have complications with various degrees of relevance. The perforation of the Schneiderian membrane is one of the most frequent events while performing the detachment of the membrane and it can increase the risk of iatrogenic sinusitis, impairment of functional homeostasis, dispersion of the graft material in the antral cavity as well as its bacterial colonization with a subsequential failure of the procedure. This report presents a case where transcrestal sinus lift was performed using Flusilift (Sweden & Martina, Due Carrare PD), a new instrument that allows fluid dynamic elevation of the sinus floor using saline solution to detach the Schneider’s membrane in an atraumatic way without using a sinus elevator and obtain an adequate alveolar ridge regeneration using hyaluronic acid in gel formulation to support an implant placement. Hyaluronic acid seems to play a key role in wound healing and contributes to a faster bone neoformation in bone regeneration procedures.

Alveolar Regeneration in COVID-19 Patients: A Network Perspective.

A viral infection involves entry and replication of viral nucleic acid in a host organism, subsequently leading to biochemical and structural alterations in the host cell. In the case of SARS-CoV-2 viral infection, over-activation of the host immune system may lead to lung damage. Albeit the regeneration and fibrotic repair processes being the two protective host responses, prolonged injury may lead to excessive fibrosis, a pathological state that can result in lung collapse. In this review, we discuss regeneration and fibrosis processes in response to SARS-CoV-2 and provide our viewpoint on the triggering of alveolar regeneration in coronavirus disease 2019 (COVID-19) patients.

Science, Medicine, and the Anesthesiologist

Science, Medicine, and the Anesthesiologist

Effect of Systemic Treatment by Nanoparticles on The inferior Alveolar Nerve Regeneration and Brain after Crush Injury in Rabbit.

Effect of Systemic Treatment by Nanoparticles on The inferior Alveolar Nerve Regeneration and Brain after Crush Injury in Rabbit.

Release of Notch activity coordinated by IL-1β signalling confers differentiation plasticity of airway progenitors via Fosl2 during alveolar regeneration

While the acquisition of cellular plasticity in adult stem cells is essential for rapid regeneration after tissue injury, little is known about the underlying mechanisms governing this process. Our data reveal the coordination of airway progenitor differentiation plasticity by inflammatory signals during alveolar regeneration. Upon damage, IL-1β signalling-dependent modulation of Jag1/2 expression in ciliated cells results in the inhibition of Notch signalling in secretory cells, which drives reprogramming and acquisition of differentiation plasticity. We identify a transcription factor Fosl2/Fra2 for secretory cell fate conversion to alveolar type 2 (AT2) cells retaining the distinct genetic and epigenetic signatures of secretory lineages. We furthermore reveal that KDR/FLK-1+ human secretory cells display a conserved capacity to generate AT2 cells via Notch inhibition. Our results demonstrate the functional role of a IL-1β-Notch-Fosl2 axis for the fate decision of secretory cells during injury repair, proposing a potential therapeutic target for human lung alveolar regeneration.

Cover page

COVER ILLUSTRATION: The cover image is based on the Original Article Alveolar ridge regeneration in two-wall-damaged extraction sockets of an in vivo experimental model by Jung-Seok Lee et al., https://doi.org/10.1111/clr.13791.

The cell cycle inhibitor P21 promotes the development of pulmonary fibrosis by suppressing lung alveolar regeneration

The cell cycle inhibitor P21 has been implicated in cell senescence and plays an important role in the injury–repair process following lung injury. Pulmonary fibrosis (PF) is a fibrotic lung disorder characterized by cell senescence in lung alveolar epithelial cells. In this study, we report that P21 expression was increased in alveolar epithelial type 2 cells (AEC2s) in a time-dependent manner following multiple bleomycin-induced PF. Repeated injury of AEC2s resulted in telomere shortening and triggered P21-dependent cell senescence. AEC2s with elevated expression of P21 lost their self-renewal and differentiation abilities. In particular, elevated P21 not only induced cell cycle arrest in AEC2s but also bound to P300 and β-catenin and inhibited AEC2 differentiation by disturbing the P300–β-catenin interaction. Meanwhile, senescent AEC2s triggered myofibroblast activation by releasing profibrotic cytokines. Knockdown of P21 restored AEC2-mediated lung alveolar regeneration in mice with chronic PF. The results of our study reveal a mechanism of P21-mediated lung regeneration failure during PF development, which suggests a potential strategy for the treatment of fibrotic lung diseases.

Tempo-spatial regulation of the Wnt pathway by FAM13A modulates the stemness of alveolar epithelial progenitors.

BackgroundFamily with Sequence Similarity 13, Member A (FAM13A) gene has been consistently associated with COPD by Genome-wide association studies (GWAS). Our previous study demonstrated that FAM13A was mainly expressed in the lung epithelial progenitors including Club cells and alveolar type II epithelial (ATII) cells. Fam13a−/− mice were resistant to cigarette smoke (CS)–induced emphysema through promoting β-catenin/Wnt activation. Given the important roles of β-catenin/Wnt activation in alveolar regeneration during injury, it is unclear when and where FAM13A regulates the Wnt pathway, the requisite pathway for alveolar epithelial repair, in vivo during CS exposure in lung epithelial progenitors.MethodsFam13a+/+ or Fam13a−/− mice were crossed with TCF/Lef:H2B-GFP Wnt-signaling reporter mouse line to indicate β-catenin/Wnt-activated cells labeled with GFP followed by acute (1 month) or chronic (7 months) CS exposure. Fluorescence-activated flow cytometry analysis, immunofluorescence and organoid culture system were performed to identify the β-catenin/Wnt-activated cells in Fam13a+/+ or Fam13a−/− mice exposed to CS. Fam13a;SftpcCreERT2;Rosa26RmTmG mouse line, where GFP labels ATII cells, was generated for alveolar organoid culture followed by analyses of organoid number, immunofluorescence and gene expression. Single cell RNA-seq data from COPD ever smokers and nonsmoker control lungs were further analyzed.FindingsWe found that FAM13A-deficiency significantly increased Wnt activation mainly in lung epithelial cells. Consistently, after long-term CS exposure in vivo, FAM13A deficiency bestows alveolar epithelial progenitor cells with enhanced proliferation and differentiation in the ex vivo organoid model. Importantly, expression of FAM13A is significantly increased in human COPD-derived ATII cells compared to healthy ATII cells as suggested by single cell RNA-sequencing data.InterpretationOur findings suggest that FAM13A-deficiency promotes the Wnt pathway-mediated ATII cell repair/regeneration, and thereby possibly mitigating CS-induced alveolar destruction.FundThis project is funded by the National Institutes of Health of United States of America (NIH) grants R01HL127200, R01HL137927, R01HL148667 and R01HL147148 (XZ).

Alveolar ridge regeneration in two-wall-damaged extraction sockets of an in vivo experimental model.

AimTo determine the healing outcome following grafting with deproteinized porcine bone mineral (DPBM) with or without collagen membrane coverage in two‐wall (both buccal and lingual)‐damaged extraction sockets.Materials and methodsDistal roots of three mandibular premolars in six beagle dogs were extracted, and the whole buccal and lingual bony walls were surgically removed. Three treatment protocols were then applied according to the following group allocation: no graft (None), grafting DPBM (BG), and grafting DPBM with coverage by a collagen membrane (BG + M). Two observational periods (2 and 8 weeks) were used with the split‐mouth design, and quantitative and qualitative analyses were performed by microcomputed tomography and histology.ResultsThe dimensions of the alveolar ridge at both grafted sites (BG and BG + M) remained similar to those of the pristine ridge in the histologic and radiographic analyses, whereas the ungrafted sites (None) collapsed both vertically and horizontally. Both grafting protocols produced substantial bony regeneration, but the addition of a covering membrane enhanced the proportion of mineralized tissue within the augmented area, and the BG + M group also showed a significantly larger area of regenerated ridge than the None group (p < .05).ConclusionsBone grafting with collagen membrane can maintain the alveolar ridge dimensions with substantial bone regeneration in a two‐wall‐damaged extraction socket.

SOX9 inactivation affects the proliferation and differentiation of human lung organoids

BackgroundThe regulation of the transcription factor sex-determining region Y-box transcription factor 9 (SOX9) in lung development has been described in mouse, but the same principles apply to human lung development is unknown due to a lack of appropriate experimental approaches and models.MethodsHere, we used gene editing technology to inactivate SOX9 in human embryonic stem cells that were then induced to differentiate into lung organoids to investigate the role of SOX9 in human lung epithelium development.ResultsComplete knockout of the transactivation domain of SOX9 by gene editing resulted in indels in both alleles of SOX9. SOX9−/− hESCs could be induced to differentiate into lung progenitor organoids. In vitro long-term expansion showed that SOX9 inactivation did not affect the differentiation of pulmonary epithelial cells, but promoted apoptosis and reduced proliferative capacity in the organoids. When lung progenitor organoids were transplanted under the kidney capsule of immunodeficient mice, expression of the club cell marker secretoglobin family 1A member 1 (SCGB1A1) was detected in SOX9−/− transplants but was absent in wild-type (WT) transplants. The maturation of goblet cells was also affected by SOX9 inactivation, as evidenced by the presence of mucin 5 AC (MUC5AC) in the cytoplasm of SOX9−/− grafts as compared to WT grafts in which most MUC5AC was secreted into the lumen. In vivo lung orthotopic transplantations showed that SOX9 inactivation had a limited effect on the differentiation of alveolar cells and lung regeneration in injured mice.ConclusionsSOX9 modulates the proliferative capacity of lung epithelium but is not an indispensable transcription factor in the regulation of human lung epithelium development.

VEGF receptor 2 (KDR) protects airways from mucus metaplasia through a Sox9-dependent pathway.

Mucus-secreting goblet cells are the dominant cell type in pulmonary diseases, e.g., asthma and cystic fibrosis (CF), leading to pathologic mucus metaplasia and airway obstruction. Cytokines including IL-13 are the major players in the transdifferentiation of club cells into goblet cells. Unexpectedly, we have uncovered a previously undescribed pathway promoting mucous metaplasia that involves VEGFa and its receptor KDR. Single-cell RNA sequencing analysis coupled with genetic mouse modeling demonstrates that loss of epithelial VEGFa, KDR, or MEK/ERK kinase promotes excessive club-to-goblet transdifferentiation during development and regeneration. Sox9 is required for goblet cell differentiation following Kdr inhibition in both mouse and human club cells. Significantly, airway mucous metaplasia in asthmatic and CF patients is also associated with reduced KDR signaling and increased SOX9 expression. Together, these findings reveal an unexpected role for VEGFa/KDR signaling in the defense against mucous metaplasia, offering a potential therapeutic target for this common airway pathology.

Age-dependent alveolar epithelial plasticity orchestrates lung homeostasis and regeneration.

Regeneration of the architecturally complex alveolar niche of the lung requires precise temporal and spatial control of epithelial cell behavior. Injury can lead to a permanent reduction in gas exchange surface area and respiratory function. Using mouse models, we show that alveolar type 1 (AT1) cell plasticity is a major and unappreciated mechanism that drives regeneration, beginning in the early postnatal period during alveolar maturation. Upon acute neonatal lung injury, AT1 cells reprogram into alveolar type 2 (AT2) cells, promoting alveolar regeneration. In contrast, the ability of AT2 cells to regenerate AT1 cells is restricted to the mature lung. Unbiased genomic assessment reveals that this previously unappreciated level of plasticity is governed by the preferential activity of Hippo signaling in the AT1 cell lineage. Thus, cellular plasticity is a temporally acquired trait of the alveolar epithelium and presents an alternative mode of tissue regeneration in the postnatal lung.

Human-Based Advanced in vitro Approaches to Investigate Lung Fibrosis and Pulmonary Effects of COVID-19.

The coronavirus disease 2019 (COVID-19) pandemic has caused considerable socio-economic burden, which fueled the development of treatment strategies and vaccines at an unprecedented speed. However, our knowledge on disease recovery is sparse and concerns about long-term pulmonary impairments are increasing. Causing a broad spectrum of symptoms, COVID-19 can manifest as acute respiratory distress syndrome (ARDS) in the most severely affected patients. Notably, pulmonary infection with Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2), the causing agent of COVID-19, induces diffuse alveolar damage (DAD) followed by fibrotic remodeling and persistent reduced oxygenation in some patients. It is currently not known whether tissue scaring fully resolves or progresses to interstitial pulmonary fibrosis. The most aggressive form of pulmonary fibrosis is idiopathic pulmonary fibrosis (IPF). IPF is a fatal disease that progressively destroys alveolar architecture by uncontrolled fibroblast proliferation and the deposition of collagen and extracellular matrix (ECM) proteins. It is assumed that micro-injuries to the alveolar epithelium may be induced by inhalation of micro-particles, pathophysiological mechanical stress or viral infections, which can result in abnormal wound healing response. However, the exact underlying causes and molecular mechanisms of lung fibrosis are poorly understood due to the limited availability of clinically relevant models. Recently, the emergence of SARS-CoV-2 with the urgent need to investigate its pathogenesis and address drug options, has led to the broad application of in vivo and in vitro models to study lung diseases. In particular, advanced in vitro models including precision-cut lung slices (PCLS), lung organoids, 3D in vitro tissues and lung-on-chip (LOC) models have been successfully employed for drug screens. In order to gain a deeper understanding of SARS-CoV-2 infection and ultimately alveolar tissue regeneration, it will be crucial to optimize the available models for SARS-CoV-2 infection in multicellular systems that recapitulate tissue regeneration and fibrotic remodeling. Current evidence for SARS-CoV-2 mediated pulmonary fibrosis and a selection of classical and novel lung models will be discussed in this review.

Effect of Slit/Robo signaling on regeneration in lung emphysema

Emphysema, a pathological component of chronic obstructive pulmonary disease, causes irreversible damage to the lung. Previous studies have shown that Slit plays essential roles in cell proliferation, angiogenesis, and organ development. In this study, we evaluated the effect of Slit2 on the proliferation and migration of mouse lung epithelial cells and its role in regeneration in an emphysema lung mouse model. Here, we have shown that Slit2/Robo signaling contributes to the regeneration of lungs damaged by emphysema. Mouse epithelial lung cells treated with Slit2 exhibited increased proliferation and migration in vitro. Our results also showed that Slit2 administration improved alveolar regeneration in the emphysema mouse model in vivo. Furthermore, Slit2/Robo signaling increased the phosphorylation of ERK and Akt, which was mediated by Ras activity. These Slit2-mediated cellular signaling processes may be involved in the proliferation and migration of mouse lung epithelial cells and are also associated with the potential mechanism of lung regeneration. Our findings suggest that Slit2 administration may be beneficial for alveolar regeneration in lungs damaged by emphysema.

Alveolar epithelial cell fate is maintained in a spatially restricted manner to promote lung regeneration after acute injury.

SUMMARYAlveolar epithelial type 2 (AT2) cells integrate signals from multiple molecular pathways to proliferate and differentiate to drive regeneration of the lung alveolus. Utilizing in vivo genetic and ex vivo organoid models, we investigated the role of Fgfr2 signaling in AT2 cells across the lifespan and during adult regeneration after influenza infection. We show that, although dispensable for adult homeostasis, Fgfr2 restricts AT2 cell fate during postnatal lung development. Using an unbiased computational imaging approach, we demonstrate that Fgfr2 promotes AT2 cell proliferation and restrains differentiation in actively regenerating areas after injury. Organoid assays reveal that Fgfr2-deficient AT2 cells remain competent to respond to multiple parallel proliferative inputs. Moreover, genetic blockade of AT2 cell cytokinesis demonstrates that cell division and differentiation are uncoupled during alveolar regeneration. These data reveal that Fgfr2 maintains AT2 cell fate, balancing proliferation and differentiation during lung alveolar regeneration.

ROBO/SLIT in Obesity‐dependent Changes in Angiogenesis and Lung Regeneration

More than 35 million people in the US suffer from chronic lung diseases, and a number of these patients have pre-existing conditions including obesity. Although lung transplantation is one of the options for end-stage lung diseases, due to the shortage of donors and low survival rate, this approach is suboptimal. In particular, there is an association between obesity and a poor outcome in lung transplantation. Thus, stimulating the regenerative ability of the lungs could be one of the therapeutic strategies for end-stage lung diseases in obese patients. Angiogenesis –the formation of new blood vessels- plays an important role in lung regeneration. Obesity is accompanied by endothelial cell dysfunction. However, the effects of obesity on lung vascular and alveolar regeneration remain unclear. Here we found that compensatory lung growth after unliteral pneumonectomy (PNX) was inhibited in leptin-deficient ob/ob mice treated with a high-fat diet compared to that in control lean mice. RNA sequencing analysis demonstrates that the levels of endothelial guidance molecules such as Roundabout proteins (ROBO/SLIT), which contribute to angiogenesis, are significantly higher in post-PNX obese mouse lungs compared to those from lean mouse lungs. The mRNA levels of Robo1, 2, and 4 and Slit2 are higher in the mouse lung tissues collected from post-PNX obese mice compared to those from lean mouse lungs when analyzed using quantitative RT-PCR. Network analysis reveals that the genes listed in the top 20 BP GO terms of cellular responses including cell migration, cell proliferation and extracellular matrix assembly interact with ROBO/SLIT during post-PNX lung growth. These results suggestthat ROBO/SLIT signaling may mediate the inhibition of post-PNX regenerative lung growth in obese mice. ROBO/SLIT may be the key signaling mechanism to restore lung regeneration in obese population.

Pretreatment of aged mice with retinoic acid supports alveolar regeneration via upregulation of reciprocal PDGFA signalling

ObjectivesIdiopathic pulmonary fibrosis (IPF) primarily affects the aged population and is characterised by failure of alveolar regeneration, leading to loss of alveolar type 1 (AT1) cells. Aged mouse models of...

ALVEOLAR RECONSTRUCTION WITH THE USE OF ESTROMAL FRACTION OF BONE MARROW AUTOLOGOUS: A CASE REPORT

<h3>Background</h3> There is a great challenge in restoring the function and aesthetics of the patient using dental implants, especially when there are alveolar bone defects due to dental losses, often requiring grafting techniques for repair and reconstruction of the affected area. The objective of this study was to analyze the progress of alveolar regeneration and osteogenesis of a clinical case with bone grafting combined with the use of the stromal fraction of bone marrow, identifying the effectiveness of bone repair and exploring possibilities of use in the dental and medical area. <h3>Methods</h3> The 54-year-old patient with alveolar bone loss and atretic ridge was submitted to a graft with synthetic bone and platelet-rich fibrin (PRF), optimized through autologous cell grafting, for osteoinduction and faster alveolar reconstruction for later treatment with dental implants and prosthetic rehabilitation. <h3>Results</h3> The results analyzed through clinical and tomographic examinations, performed periodically, showed the effectiveness of the bone repair process of the patient, with consistent and quick alveolar reconstruction. <h3>Conclusion</h3> The use of stromal fraction of bone marrow combined with PRF proved to be safe and viable for the optimization of bone repair of alveolar defects.

The Glycoprotein/Cytokine Erythropoietin Promotes Rapid Alveolar Ridge Regeneration In Vivo by Promoting New Bone Extracellular Matrix Deposition in Conjunction with Coupled Angiogenesis/Osteogenesis.

The loss of bone following tooth extraction poses a significant clinical problem for maxillofacial esthetics, function, and future implant placement. In the present study, the efficacy of an erythropoietin-impregnated collagen scaffold as an alveolar ridge augmentation material versus a conventional collagen scaffold and a BioOss inorganic bovine bone xenograft was examined. The collagen/Erythropoietin (EPO) scaffold exhibited significantly more rapid and complete osseous regeneration of the alveolar defect when compared to bone xenograft and the collagen membrane alone. The new EPO induced extracellular matrix was rich in Collagen I, Collagen III, Fibronectin (Fn) and E-cadherin, and featured significantly increased levels of the osteogenic transcription factors Runt-related transcription factor 2 (Runx2) and Osterix (Osx). Histomorphometric evaluation revealed a significant two-fold increase in the number of capillaries between the EPO and the BioOss group. Moreover, there was a highly significant 3.5-fold higher level of vascular endothelial growth factor (VEGF) in the collagen/EPO-treated group compared to controls. The significant effect of EPO on VEGF, FN, and RUNX2 upregulation was confirmed in vitro, and VEGF pathway analysis using VEGF inhibitors confirmed that EPO modulated extracellular matrix protein expression through VEGF even in the absence of blood vessels. Together, these data demonstrate the effectiveness of an EPO-impregnated collagen scaffold for bone regeneration as it induces rapid matrix production and osseoinduction adjacent to new capillaries via VEGF.

The History and Mystery of Alveolar Epithelial Type II Cells: Focus on Their Physiologic and Pathologic Role in Lung.

Alveolar type II (ATII) cells are a key structure of the distal lung epithelium, where they exert their innate immune response and serve as progenitors of alveolar type I (ATI) cells, contributing to alveolar epithelial repair and regeneration. In the healthy lung, ATII cells coordinate the host defense mechanisms, not only generating a restrictive alveolar epithelial barrier, but also orchestrating host defense mechanisms and secreting surfactant proteins, which are important in lung protection against pathogen exposure. Moreover, surfactant proteins help to maintain homeostasis in the distal lung and reduce surface tension at the pulmonary air–liquid interface, thereby preventing atelectasis and reducing the work of breathing. ATII cells may also contribute to the fibroproliferative reaction by secreting growth factors and proinflammatory molecules after damage. Indeed, various acute and chronic diseases are associated with intensive inflammation. These include oedema, acute respiratory distress syndrome, fibrosis and numerous interstitial lung diseases, and are characterized by hyperplastic ATII cells which are considered an essential part of the epithelialization process and, consequently, wound healing. The aim of this review is that of revising the physiologic and pathologic role ATII cells play in pulmonary diseases, as, despite what has been learnt in the last few decades of research, the origin, phenotypic regulation and crosstalk of these cells still remain, in part, a mystery.