Monoclonal gammopathy cases of unknown significance occur mainly in aging people. Furthermore, it is now well-established that all cases of multiple myeloma, an aggressive lethal cancer of the bone marrow leading to bone destruction, emerge from such cases of gammopathy. In order to improve the management of myeloma, it is therefore critical to discover all of the specific factors responsible for their emergence. Here, we suggest that bone factors, i.e., bone senescence and bone inflammaging, could facilitate the emergence of these pathologies in aging people. The generalized bone loss observed in gammopathy and myeloma corresponds at the histological level to a disruption of the endosteal niche, which is both the site of hematopoiesis and the bone remodeling compartment and consists of a coupling between bone-forming osteoblasts and bone-resorbing osteoclasts. This niche undergoes a shift from an osteoblastic to an osteoclastic profile as soon as the physiological bone experiences senescence. Such a disrupted endosteal niche could represent the permissive microenvironment necessary for gammopathy and myeloma. Inside this environment, osteoclasts, through their capacity to suppress T cells but also present antigens in the bone marrow, could locally create immune changes favorable to the maintenance of gammopathy and its malignant transformation by favoring cellular instability.Multiple Myeloma (MM) and its preexisting stage, termed Monoclonal Gammopathy of Undetermined Significance (MGUS), have long been considered mainly as genomic diseases. However, the bone changes observed in both conditions have led to a reassessment of the role of the bone microenvironment, mainly the endosteal niche in their genesis. Here, we consider the disruption of the endosteal niche in the bone marrow, that is, the shift of the endosteal niche from an osteoblastic to an osteoclastic profile produced by bone senescence and inflammaging, as the key element. Thus, this disrupted endosteal niche is proposed to represent the permissive microenvironment necessary not only for the emergence of MM from MGUS but also for the emergence and maintenance of MGUS. Moreover, the excess of osteoclasts would favor the presentation of antigens (Ag) into the endosteal niche because osteoclasts are Ag-presenting cells. As such, they could significantly stimulate the presentation of some specific Ag and the clonal expansion of the stimulated cells as well as favor the expansion of such selected clones because osteoclasts are immunosuppressive. We also discuss this scenario in the Gaucher disease, in which the high incidence of MGUS and MM makes it a good model both at the bone level and the immunological level. Finally, we envisage that this endosteal niche disruption would increase the stochasticity (epigenetic and genetic instability) in the selected clones, according to our Tissue Disruption-induced cell Stochasticity (TiDiS) theory. All Multiple Myeloma (MM) cases emerge from a pre-existing stage termed Monoclonal Gammopathy of Undetermined Significance (MGUS), confirming the ‘2-hits’ hypothesis presented by SE Salmon and M Seligman 50 years ago . ActuallTogether with MGUS, MM-induced bone changes (lytic bone lesions that are characteristic of MM) are the second major component of MM oncogenesis . In addiIndeed, END, as a direct consequence of bone senescence and as abnormally augmented/accentuated by sterile bone inflammaging, could represent the permissive micro-environment necessary for the emergence of MGUS cases, their maintenance, and their subsequent malignant transformation into MM. Thus, we suggest that the END plays a critical role in the emergence of MGUS preceding their malignant transformation, and that this explains some immunological observations in both MGUS and MM. This scenario could also explain the high incidence of both MGUS and MM observed in Gaucher Disease (GD) cases , where bBone cells are composed of osteoblasts (OB), their descendant osteocytes and osteoclasts (OC). Bone homeostasis is maintained through persistent remodeling mediated by bone-forming OB and bone-resorbing OC. Under normal conditions, the resorption of old bone is compensated by an equal amount of new bone formation, with a coupling between the two processes that should ensure the maintenance of stable bone mass. Any event inducing an imbalance between these processes by increasing bone resorption or decreasing bone formation, or both, can result in generalized bone loss, as observed in aging and osteoporosis. Indeed, bone architecture is altered with aging: trabecular thickness and numbers decrease, porosity increases, cortical bone mass is lost, and adiposity increases in the bone marrow. In humans, bone mass loss begins between 20 and 30 years in both men and women , with anThe molecular and cellular bases underlying the pathogenesis of age-related bone changes are now well-known . An incrThe declines in differentiated cell function and stem cell reserves linked to senescence are important factors in age-related pathologies, and an increase in cellular senescence is particularly involved in bone aging ,16. IndeThe bone area of the aging endosteal niche is characterized by a reduction in the number of OB and the release of osteopontin, which negatively regulates stem cell pool size . MoreoveMGUS is also characterized by associated bone changes that include (i) generalized bone loss, leading to bone fragility with increased fracture risk (reviewed in ), and  that accumulate in the bone marrow. PCs correspond to the terminal stage of B-cell lineage differentiation. They secrete large amounts of antibodies and are the effectors of humoral immunity. In lymph nodes , secondary antigen presentation with T cells helps generate long-lived and highly mutated memory B cells and PCs, both of which are able to circulate and persist inside the bone marrow for many years. Memory B cells can be reactivated to enrich the pool of PCs. These highly mutated and long-lived PCs are the normal counterparts of MM cells .Bone marrow niches in mice are known to facilitate the survival, maintenance, and differentiation of hematopoietic multipotent progenitors and PCs. Specific stromal cells and various factors in the niches are involved, and similarities between the microenvironments necessary for the establishment and the maintenance of these two immune cell subsets exist, including the CXCL12/CXCR4 signaling axis . AlthougMM-induced bone changes are the hallmark of MM. The mechanisms of MM-induced bone changes are specific to MM, suggesting that these bone changes could be more than simply induced and viewed as a fatal attraction regarding the interactions of MM cells for/with bones, as it could also be primarily involved in the oncogenesis of MM from MGUS. Although such lytic bone lesions result from a disruption of the bone remodeling compartment, which is characterized by an uncoupling between an excess of bone resorption and a suppression of bone formation, we were the first to show that the major factor responsible for such lytic bone lesions was the suppression of bone formation, which is a disruption of the mesenchymal stromal to OB transition . Of noteWe previously proposed that age-related disruption of the BME and endosteal niches would increase the pool of PCs that cannot niche into the BME during polyclonal B cell activation and transient MGUS, when the number of PCs needing to fully differentiate is enhanced ,40. As aIn summary, an early (pre-MGUS) END, in relation to increased bone senescence , prefigures the disruption of the bone remodeling compartment observed first in MGUS/MGSS and then in MM. We propose  that theUntil now, infection, inflammation, and immune deficiency have been the most documented contexts for the emergence of MGUS. In these contexts, the role of Ag appears to be of major importance. Many studies have now demonstrated the presence of immunoglobulin (Ig)-gene mutations in both MGUS and MM, showing that both MGUS and MM clones are clones engaged in an immune response . MoreoveThen, the central question is: can the bone marrow present Ag by itself ? It is iFrom our model, we propose that END can favor the presentation of these specific Ag by the OC . Indeed,Recent elegant studies in patients with GD, in whom an excess of both MGUS and MM was observed, are highly relevant to this concept. GD is characterized by an increased incidence of hyper-gammaglobulinemia (100%), MGUS (30%), and MM (4%) with this characteristic sequence: hyper-gammaglobulinemia, then MGUS, then MM . Thus, GStrikingly, this antigenic specificity for the sphingolipids of GD-associated monoclonal immunoglobulins is also observed in 15–20% of sporadic MGUS , suggestInside the bone marrow, now considered as an antigen-presenting site (at least for certain antigens as auto-antigens), our comprehensive bone scenario for both MGUS and MM initiates with bone senescence and bone inflammaging (END) that has three consequences: (i) abnormal Ag presentation through OC as Ag-presenting cells in excess, (ii) local immune defects through OC, and (iii) increased stochasticity (epigenetic and genetic instability) in selected clones. END, through the activation of OC as Ag-presenting cells, could significantly stimulate the presentation of some specific Ag and the clonal expansion of such stimulated cells. The presence of such Ag inside the bone marrow could also facilitate the maintenance of such selected clones. This scenario that integrates bone senescence, immune stimulation, the BME, and the genesis of MGUS and MM provides a new perspective at the tissue level that appears to better reconcile the different sides of the disease than canonical, genome-centered perspectives.


This case report presents a rare occurrence of basal cell carcinoma (BCC) in the periungual region of the thumb. BCC is the most common type of skin cancer, typically found in sun-exposed areas. The discussion explores the underlying pathogenesis mechanisms, including the role of ultraviolet exposure, the absence of pilosebaceous units, and the involvement of the sonic hedgehog (SHH) pathway. Understanding the complexities of BCC in atypical locations is essential for effective prevention and treatment strategies. Basal cell carcinoma (BCC) is the most prevalent form of cancer globally, and its incidence is on the rise . While BA 54-year-old man with a history of well-controlled hypertension visited with a complaint about a finger lesion that had been present for two years Figure . Upon exDermoscopy revealed characteristic features of a non-melanocytic lesion, including redness, a dotted appearance, globules, arborescent vessels, and areas of disrupted white coloration Figure .Dermatopathologic analysis confirmed the presence of basaloid cell nests arranged in a palisading pattern within the top layer of the skin Figure . Based oBCC accounts for approximately 80% of all skin cancers, making it the most common type of cancer . AlthougTo explore the existing literature, we conducted a search on PubMed using the terms "basal cell carcinoma" and "nail fold" or “periungual” spanning from 1931 to 2022. Our search yielded 32 cases, with the thumb being the most affected finger. Interestingly, most cases had a disease duration of four to five years, and a significant number had received different initial diagnoses, highlighting the low index of suspicion for BCC in these locations. It is worth noting that most of these misdiagnoses occurred prior to the use of dermoscopy. We believe that the widespread adoption of dermoscopy has facilitated the detection of more cases, even in situations where there is no suspicion of BCC. Also, most of the cases affected fingernails, according to Mortada et al., who conducted a recent systematic review of BCC in the hands and identified 34 cases out of 2,051 patients with involvement of the fingernails, which also underscored the high variability of morphological features [Regarding BCCs occurring in atypical locations, Carter-Grine et al. proposed interesting hypotheses about the underlying oncogenic mechanisms. They suggested that the rarity of BCC in these locations may be associated with the absence of pilosebaceous units, as the oncogenesis of BCC is typically related to hair follicles . HoweverThe development of BCC is influenced by multiple factors, including UV exposure, the presence of pilosebaceous units, and intricate mechanisms within and beyond the SHH pathway. Understanding these complex interactions is crucial for comprehending the occurrence of BCC in atypical locations and advancing our strategies for prevention and treatment.It is conceivable that somatic mutations play a role in influencing these mechanisms in certain individuals, potentially rendering specific anatomical regions more prone to developing BCC at these less common sites. This case encourages us to think beyond the conventional and consider the possibility of BCC at uncommon sites.


Aims: Medication-related osteonecrosis of the jaw (MRONJ) is a drug-related adverse reaction characterized by bone destruction and necrosis in the jaw. This case series aims to evaluate the treatment approaches and outcomes in MRONJ patients. Materials and methods: The retrospective study was conducted at the Dental Unit of the University of Bari, Italy. Patients with MRONJ were treated and followed up for 60 months. The treatment approach involved piezosurgery and concentrated growth factor (CGF). Six clinical cases from this group are described in detail. Results: None of the patients showed recurrence of necrotic MRONJ lesions during the follow-up period. The surgical interventions, including bone resections and the application of CGF, resulted in successful mucosal healing and the prevention of disease progression. Conclusions: This study highlights the complexity of managing MRONJ and the importance of a multidisciplinary approach. Conservative treatment options and minimally invasive surgery have shown efficacy in controlling symptoms and improving patients’ quality of life. However, the optimal treatment approach remains a challenge, and further studies are needed to evaluate alternative therapies and resective surgery. A comprehensive preoperative evaluation and collaboration among dental, endocrinology, and oncology specialists are crucial for personalized and multidisciplinary management. Ongoing research efforts are necessary to explore new therapeutic modalities and improve our understanding of MRONJ management, providing better support to patients dealing with this complex condition. Identified in the dental literature since 2003 , medicatMRONJ pathogenesis is based on the activity of medications that target osteoblasts, which are involved in bone remodeling, turnover, and vascularization. Osteoblasts deposit in the mineralized matrix and convert into osteocytes, which live for about 180 days ,5. They When the osteocyte decays, OPG is no longer produced and, as a result of RANK-receptor binding on the osteoclast, necrotic or dysfunctional bone tissue is resorbed. This mechanism is the basis of bone homeostasis, which guarantees the elasticity and load-bearing characteristics of the skeletal structure ,9.Bone turnover in the maxilla is 10 times faster than in the long bones, especially in the alveolar process of the post-extraction alveoli, the postero-lingual area, the maxillary sinus, and the torus [The mandibular vascularization is terminal;The mucosa and underlying periosteum are intrinsically exposed to trauma during masticatory phases;There is a high concentration of bacteria in the salivary biofilm;The periodontal ligament is also present ,8,11.MRONJ is a pathology that primarily affects the jaw bones because they have the following distinctive features:The risk level is determined by the kind of medication administered, the dose, the frequency, and the length of administration, and the timing of the last dose. Bisphosphonates (BPs): BPs are pyrophosphate analogues that form strong bonds with hydroxyapatite, which is the mineral component of bone. They are commonly used for the treatment of osteoporosis, bone metastases, and other bone-related conditions. However, long-term use of BPs has been linked to an increased risk of developing MRONJ .Tyrosine kinase inhibitors (such as sunitinib): these drugs are used in the treatment of various cancers, including kidney cancer and gastrointestinal stromal tumors. They work by inhibiting the activity of specific enzymes involved in cell signaling pathways. While they have shown efficacy in cancer treatment, they have also been associated with the development of MRONJ .Monoclonal antibodies (such as denosumab): monoclonal antibodies are designed to target specific proteins or receptors involved in disease processes. Denosumab, for example, is a monoclonal antibody used for the treatment of osteoporosis and bone metastases. However, like the other drugs on this list, denosumab has been linked to an increased risk of MRONJ .Angiogenesis inhibitors (such as bevacizumab): these drugs inhibit the formation of new blood vessels, which can be beneficial in the treatment of cancer and other conditions. However, they can also interfere with the normal healing process of the jawbone, leading to the development of MRONJ .Fusion proteins (such as aflibercept): fusion proteins are created by combining different protein components to target specific molecules involved in disease processes. Aflibercept, for instance, is a fusion protein used in the treatment of certain cancers. However, its use has been associated with an increased risk of MRONJ .mTOR inhibitors (such as Everolimus): mTOR inhibitors are a class of drugs that inhibit the mammalian target of rapamycin (mTOR), a protein involved in cell growth and division. These drugs have shown efficacy in cancer treatment, but they have also been associated with an increased risk of MRONJ .Radiopharmaceuticals (such as radium-223): radiopharmaceuticals are drugs that contain radioactive substances used for diagnostic or therapeutic purposes. Radium-223, for example, is used in the treatment of metastatic prostate cancer. However, its use has been linked to an increased risk of MRONJ .Estrogen inhibitors : estrogen inhibitors are drugs used in the treatment of hormone-receptor-positive breast cancer and osteoporosis. Raloxifene, for instance, is an estrogen inhibitor that has been associated with an increased risk of MRONJ .Immunomodulators (such as methotrexate and corticosteroids): immunomodulators are drugs that modify the immune response. Methotrexate, a commonly used immunosuppressive drug, and corticosteroids, which have potent anti-inflammatory properties, have both been linked to an increased risk of MRONJ .Several drugs have been identified in recent research as being associated with MRONJ . These dMRONJ mostly affects patients taking one or more of these drugs because they present oncological diseases, osteoporotic diseases, or osteometabolic diseases, subsequent to exposure to a local triggering factor .The most common local factor is tooth extraction (about 61%); however, MRONJ can also be initiated by chronic inflammation caused by untreated periodontitis, bone biopsies, the clinical elongation of crowns, surgery on the bone, and the insertion of a dental implant, all of which lead to trauma and the need for bone turnover, which these drugs inhibit .Many categories have been developed over the years to help surgeons to determine the best treatment method based on the severity of the clinical picture, the cost–benefit ratio, and the patients’ different comorbidities .In 2014, Favia et al. presented the dimensional staging of MRONJ to appropriately assess treatment plans.Stage 0: this stage is characterized by the presence of clinical symptoms and nonspecific radiological signs, but there is no exposure of the underlying bone.Stage 1: in this stage, there is exposed bone measuring less than 2 cm, with or without associated pain.Stage 2: here, the exposed bone measures between 2 and 4 cm, and there is pain that can be managed with non-steroidal anti-inflammatory drugs (NSAIDs).Stage 3: this is the most advanced stage, where the exposed bone measures more than 4 cm. The pain experienced in this stage is not responsive to NSAIDs. Additionally, complications such as fistulae  or the involvement of the maxillary sinus or the inferior alveolar nerve may be present.It is important to note that these stages provide a framework for assessing the severity of MRONJ and guiding treatment decisions. Each stage represents a progression in the disease, with Stage 3 being the most severe and complex . The effOriginally, surgical therapy was prioritized; however, medical professionals are now encouraged to adopt more medical, non-surgical treatments at an early stage . AccordiThe study describes patients with a diagnosis of MRONJ who were treated at the Dental Unit of the University of Bari.Our treatment protocol for MRONJ followed a multidisciplinary approach involving medical and surgical interventions. The initial treatment approach was non-surgical or conservative, focusing on the management of infection and pain. This included the use of antimicrobial mouth rinses, antibiotic therapy, and non-steroidal anti-inflammatory drugs (NSAIDs). The specific medications and the treatment duration were based on individual patient needs.For patients who did not respond to conservative treatment or had advanced stages of MRONJ, surgical intervention was recommended. The surgical approach involved the use of piezosurgery, which is a minimally invasive technique for bone resection. The necrotic bone tissue was removed using piezosurgery, and debridement and sequestrectomy were performed as needed. Flap design and suturing techniques were employed to promote proper healing.Autologous preparations derived from the patient’s blood, such as concentrated growth factor (CGF), were used to enhance bone healing and regeneration. CGF was applied during the surgical procedures to promote stable repair over time.The clinical assessment of the MRONJ lesions included radiographic examinations, cone-beam computed tomography (CBCT), and orthopantomography (OPT). The severity of the lesions was classified according to Favia’s staging system, which categorizes MRONJ into four stages based on the extent of bone involvement and associated symptoms.Patients were followed up for a period of 60 months to monitor the healing of the MRONJ lesions and assess for any recurrence. Follow-up evaluations included clinical examinations, radiographic imaging, and assessments of pain symptoms.The results of the study are presented as clinical case descriptions for six patients with MRONJ who underwent treatment at the Dental Unit of the University of Bari. The treatment approach, including medical therapy, surgical intervention, and the use of CGF, is described for each case. The clinical and radiographic outcomes at different time points are also presented to demonstrate the effectiveness of the treatment approach.The current retrospective clinical cases were treated in complete conformity with ethical standards, including the World Medical Association Declaration of Helsinki and the extra requirements of Italian legislation, at the Dental Unit of the University of Bari . Additionally, the University of Bari in Italy designated the project as being free from the need for ethical assessment since it includes only minor risks and the use of available information that consists only of non-identifiable data about humans. A formal informed consent form was signed by the patients. Our research group collaborated with the Phan Chau Trinh University of Medicine to carry out this restrospective study.Our research group evaluated the 60-month follow-up of patients treated for MRONJ lesions with piezosurgery and CGF. None of the patients showed recurrences of necrotic MRONJ lesions. In the following, we illustrate the treatment of six clinical cases belonging to this group.This case featured a 78-year-old female patient affected by MRONJ following the intravenous administration of BPs. Radiographically, there was the presence in the fourth quadrant of a voluminous stage III lesion, according to Favia’s classification, which resulted in the fracture of the mandibular corpus . The patA 74-year-old female patient who was affected by osteoporosis received intravenous BPs that caused MRONJ in areas 33–35. The margins and size of the lesion were assessed by radiographic examination and CBCT ; clinicaA 77-year-old male patient who received BPs therapy developed a degree I lesion in the second quadrant, according to Favia’s classification . The thrFollowing the use of oral BPs, a 75-year-old female patient developed MRONJ in the area of teeth 35–36. The lesion was discovered radiologically using orthopantomography. (OPT) .® tip, and the bone plug was removed once more  excision of a skeletal segment with the interruption of its anatomical continuity is referred to as segmental resective surgery ,44.The most frequent form of mandibular reconstructive surgery is a mandibulectomy. It always leads to a loss of symmetry and occlusion of the lower half of the face. After surgery, titanium reconstruction plates, anatomical mandibular replicas, or vascularized bone flaps can be used to restore missing bone . When tiAn upper jaw maxillectomy, a segmental resective procedure, is typically identified by the vertical and horizontal expansions of the defect caused by the removal of aberrant tissue . The remAccording to the research, individuals with MRONJ have significant rates of recurrence/dehiscence following surgical resection, which results in more hospital stays and subsequent operations . As a reThe management of MRONJ presents several challenges. Non-surgical or conservative approaches are recommended in the early stages of the disease to stabilize the clinical picture and limit disease progression. Conservative treatment focuses on infection and pain management by using antimicrobial mouth rinses, antibiotic therapy, and non-steroidal anti-inflammatory drugs (NSAIDs). Surgical intervention is typically recommended for patients who do not respond to conservative treatment or have advanced stages of MRONJ. Surgical approaches involve debridement, sequestrectomy, and bone resection using techniques such as piezosurgery. The choice of treatment approach depends on the severity of the disease and individual patient needs.The efficacy and success rates of non-surgical and surgical treatments for MRONJ have been demonstrated in clinical cases. Non-surgical approaches aim to stabilize or downstage the lesions, while surgical interventions aim for the complete removal of necrotic bone tissue. The use of autologous preparations derived from the patient’s blood, such as concentrated growth factor (CGF), has shown promising results in promoting bone healing and regeneration.Complications and side effects associated with MRONJ treatment include the risk of recurrence or dehiscence following surgical resection, postoperative infections, and the need for subsequent operations. It is crucial to carefully evaluate resection margins and ensure the complete removal of pathological bone tissue while preserving healthy tissue.MRONJ can significantly impact the quality of life of affected patients, causing pain, functional limitations, and aesthetic concerns. The effective management of MRONJ requires a multidisciplinary approach, the close monitoring of symptoms, and tailored treatment strategies based on the specific stage and individual patients’ needs.The study on medication-related osteonecrosis of the jaw (MRONJ) presented in this article has several limitations that need to be considered. Firstly, the study design is a case series, which has inherent limitations in terms of generalizability and the establishment of causal relationships. Additionally, the study was conducted in a single dental unit, which may limit the representativeness of the patient population and the applicability of the treatment protocol to other settings.Furthermore, the retrospective nature of the study introduces potential biases and limitations in terms of data collection and analysis. The absence of a control group makes it difficult to assess the comparative effectiveness of different treatment approaches or to draw definitive conclusions about the efficacy of the interventions used.Moreover, the follow-up period of 60 months may not be sufficient to fully evaluate the long-term outcomes and recurrence rates of MRONJ. Longer-term follow-up studies are needed to assess the durability of the treatment effects and to identify any potential late complications or relapses.Additionally, the article does not discuss potential confounding factors, such as comorbidities or concomitant medications, which may influence the outcomes of MRONJ treatments. These factors can introduce variability and limit our ability to attribute the observed outcomes solely to the interventions used in the study.Further research, including well-designed randomized controlled trials with larger sample sizes and longer follow-up periods, is needed to establish more robust evidence and guidelines for the management of MRONJ.In conclusion, this clinical case series of medication-related osteonecrosis of the jaw (MRONJ) caused by bisphosphonates and monoclonal drugs highlights the complexity and importance of effectively managing this debilitating condition. The available conservative treatment options, including topical therapies, antibiotics, and minimally invasive surgery, have demonstrated their efficacy in controlling symptoms, slowing down disease progression, and enhancing the quality of life of affected patients.However, selecting the optimal treatment approach still poses a significant challenge due to the need for further studies to thoroughly evaluate the benefits and risks of certain alternative therapies, such as teriparatide therapy or resective surgery.The significance of a comprehensive preoperative evaluation, which encompasses the analysis of necrotic lesions and resection margins, cannot be overstated, as it ensures complete and stable healing over time.While our understandings of and treatments for MRONJ continue to evolve, it is crucial for clinicians to maintain close collaboration with specialists from diverse disciplines such as dentistry, endocrinology, and oncology. This interdisciplinary collaboration facilitates the development of personalized and multidisciplinary approaches that are tailored to the specific needs of each patient.Lastly, ongoing research efforts are imperative to explore new therapeutic modalities and enhance our understanding of MRONJ management. By making such efforts, we can provide the best possible support to patients dealing with this complex and continuously evolving condition.


COVID-19 has highlighted challenges in the measurement quality and comparability of serological binding and neutralization assays. Due to many different assay formats and reagents, these measurements are known to be highly variable with large uncertainties. The development of the WHO international standard (WHO IS) and other pool standards have facilitated assay comparability through normalization to a common material but does not provide assay harmonization nor uncertainty quantification. In this paper, we present the results from an interlaboratory study that led to the development of (1) a novel hierarchy of data analyses based on the thermodynamics of antibody binding and (2) a modeling framework that quantifies the probability of neutralization potential for a given binding measurement. Importantly, we introduced a precise, mathematical definition of harmonization that separates the sources of quantitative uncertainties, some of which can be corrected to enable, for the first time, assay comparability. Both the theory and experimental data confirmed that mAbs and WHO IS performed identically as a primary standard for establishing traceability and bridging across different assay platforms. The metrological anchoring of complex serological binding and neuralization assays and fast turn-around production of an mAb reference control can enable the unprecedented comparability and traceability of serological binding assay results for new variants of SARS-CoV-2 and immune responses to other viruses. Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in unprecedented disruptions to society, but also led to impressive innovations in the fields of diagnostics, vaccines, and therapeutics. Serology assays have been and continue to be vital for managing COVID-19 ,4. In thSerological binding assays are known to have large variations across different laboratories due to the multi-component nature of the assays ,12. AmonDue to biological complexities, the standardization of serology assays has been extremely challenging . It is gMonoclonal antibodies (mAb), including anti-SARS-CoV-2 spike-neutralizing antibodies (nAb), can be generated rapidly with the current technologies ,23. mAbss,r determined using Equation (5) are shown as red squares along with harmonized antibody concentrations s,r. The randomness associated with assay- and sample-dependent uncertainties as Δgr,n a. The core clear c.s,r due to the reference-dependent bias When the same analysis procedure was implemented using each of the three mAb as the standard, we observed the following: identical trends for tainties , all conp < 0.000, between pvNT and the surrogate assay  curves to evaluate the performance of the assays d 26]. T. T26]. TConsidering pvNT as an optimal predictor of the presence of the neutralizing antibodies in this study, we constructed the probability model connecting consensus-binding concentrations to their neutralization counterparts. The left plot of From a practical standpoint, however, the relationship expressed by Equation (8) and illustrated in The results shown in s,r, assay- and reference-dependent (systematic) bias s,r cannot be determined in a diagnostic setting where only one assay is used. Thus, the harmonized measurement s,r, and the variance Our analysis examines the choice of reference through the lens of uncertainty quantification (UQ). Specifically, our new analysis method (Equation (5)) shows that the average log antibody concentration s,r, One unexpected finding is that, through the new analysis methods, we were able to generate quantitative metrics, namely, cn curves , not jusOur interlaboratory study includes assays spanning live-virus MN assay to surrogate the neutralization assay, enabling the assessment of the strengths and weakness of different neutralization assay formats. Live-virus MN assays are thought to provide the most biologically relevant data but must be conducted in a biosafety level 3 (BSL3) containment facility. They are also time-consuming, labor-intensive, and known to have relatively large uncertainties . Hence, The surrogate assay clearly showed the highest precision . ConsistA fundamental goal of serology testing is to understand the extent to which an individual is protected from further infection. Because neutralization assays are essential for determining the infection rate, predicted humoral protection, as well as vaccine efficacy during clinical trials and after large-scale vaccination, the different platforms of neutralization assays, including MN, pvNT, and surrogate assay, have been developed . An incrChief among these is the recognition that assessing immunity, whether expressed in NT50 levels or otherwise, amounts to decision making under uncertainty. This is most clearly expressed by This approach to UQ-based decision making is also well adapted to incorporating information from our harmonization analysis. As discussed in previous sections, the uncertainty lso ref.  for moreThe study materials consisted of WHO IS, 3 anti-SARS-CoV-2 spike mAbs, and 62 human serum/plasma samples. Two of the three mAbs were provided by Regeneron via a material transfer agreement. Another mAb and 47 convalescent serum samples consisting of 24 negative samples (collected pre-pandemic) and 23 positive samples  were donated by Abbott Laboratories. Fifteen convalescent plasma/serum samples provided by CDC were from vaccinated individuals, and five samples from each vaccine modality: Johnson and Johnson, Moderna, and Pfizer. All convalescent sample curations were IRB-approved by the respective organizations. The study was also approved by the IRB office of the National Institute of Standards and Technology (NIST) through an individual material transfer agreement with each participating organization.A detailed study design is provided in the Six serological binding assays and three neutralization assays were incorporated in the centralized data analysis of this interlaboratory study ) and B is an arbitrarily assigned binding antibody unit (BAU) concentration, e.g., associated with the WHO IS or another standard. Here and throughout the paper, the subscripts s, n, and r are reserved for a sample, assay, and reference, respectively. This observation motivated us to define normalization as the task of estimating the antibody quantity relative to a chosen reference for a fixed assay. The normalized antibody concentration To better utilize the entire set of data from the dilution measurements that reduces random variations, we developed a novel analysis method that generalizes the slope-correction method  and direurve see . In otheIn contrast, harmonization is intuitively defined as the process of altering the numerical values of normalized concentrations so that the  measurements of each assay become interchangeable for a fixed sample. This leads to the concept of a consensus antibody concentration ts, etc. . ImportaEquation (5) provides the bases for determining the harmonized antibody concentration, Effectively, AbN is the total number of labeled Ab immobilized on surface and maxN and minN are the maximum and minimum numbers of the labeled AbN, respectively. NT50 equals to the dilution at which half of the immobilized antigen molecules are filled and n is the Hill coefficient. Equation (7) is an excellent representation of Gibbs free energy function characterizing the thermodynamic interaction system between immobilized antigen molecules and antiviral antibodies in the sample. A detailed procedure for applying Equation (7) for the three neutralization assays platformed is provided in the For serology measurements, diluting samples containing antiviral antibodies changes the number of antibodies bound to the assay surface, as given by the Hill equation, shown in Equation (7) .(7)NAb=To construct the probability models that connect binding and neutralizing assays, we used the consensus values This motivated us to postulate a minimal model in which To specify an experimental procedure as completely as possible, certain commercial materials, instruments, and equipment were identified in this manuscript. In no case does the identification of the manufacturer of particular equipment or materials imply a recommendation or endorsement by the National Institute of Standards and Technology or the Centers for Disease Control and Prevention, nor does it imply that the materials, instruments, and equipment identified are necessarily the best available for the purpose. The findings and conclusions in this article are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention or the Agency for Toxic Substances and Disease Registry.The results from this interlaboratory study and accompanying new analysis methods provide a means to achieve unprecedented serological-binding assay harmonization. Importantly, our analysis methods afford a more rigorous quantitation of antibody concentrations as well as sources of uncertainties, thus allowing a better understanding of assay performance. Generally, harmonization is sought through experimental design that requires all laboratories use the same reagents to perform the identical assay. Because of logistics and supply chain issues associated with this method of assay harmonization and the existence of diverse serological assay platforms using different assay reagents during the COVID-19 pandemic, the harmonization of serological-binding assays has not been accomplished at present. In this paper, we conclusively demonstrated that a single anti-spike mAb can be used alone for assay harmonization when utilizing the new analysis method. Considering the presence of different viral variants respective to the original Wuhan-Hu-1 strain investigated in this study, a cocktail of mAbs with different binding specificities to different viral strains would be better to harmonize serological-binding assays across different viral strains. This means that mAb-based reference materials can be rapidly developed to support emerging infections. Together, and for the first time, we can establish methods to enable traceability and comparability across different assay platforms and reference materials.This interlaboratory study also directly compared surrogate, pvNT, and MN neutralization assays and showed that the pvNT assay had the highest sensitivity and specificity compared to the other methods. Using NT50 values from the pvNT assay, we showed good correlations with binding assays used in the study, consistent with literature reports of the two markers, anti-spike antibody titer and neutralizing antibody titer, serving as CoPs for the efficacy evaluation of vaccines and COVID-19 disease management.This approach was developed and demonstrated for COVID-19, although we expect the thermodynamics underpinning of binding assays to be generally applicable to broader serological assays, including efforts to develop new vaccines for a variety of diseases, such as RSV and pan-influenza, for seroprevalence monitoring and in the assessment of pre-existing immunity prior to the administration of therapeutics, such as emerging gene therapies.


The degeneration of osteochondral tissue represents one of the major causes of disability in modern society and it is expected to fuel the demand for new solutions to repair and regenerate the damaged articular joints. In particular, osteoarthritis (OA) is the most common complication in articular diseases and a leading cause of chronic disability affecting a steady increasing number of people. The regeneration of osteochondral (OC) defects is one of the most challenging tasks in orthopedics since this anatomical region is composed of different tissues, characterized by antithetic features and functionalities, in tight connection to work together as a joint. The altered structural and mechanical joint environment impairs the natural tissue metabolism, thus making OC regeneration even more challenging. In this scenario, marine-derived ingredients elicit ever-increased interest for biomedical applications as a result of their outstanding mechanical and multiple biologic properties. The review highlights the possibility to exploit such unique features using a combination of bio-inspired synthesis process and 3D manufacturing technologies, relevant to generate compositionally and structurally graded hybrid constructs reproducing the smart architecture and biomechanical functions of natural OC regions. The aging population, increasing obesity, and a surge in the number of sports-related injuries are the main drivers for the worldwide rising prevalence of bone and osteochondral (OC) diseases. Osteoarthritis (OA) refers to a rheumatic musculoskeletal disorder characterized by the progressive deterioration of the articular cartilage and sclerosis of the subchondral bone, together with changes in the synovial tissues, as a result of mechanical and biological processes that modify the cartilage homeostasis ,2. InjurFor several decades, both scientists and clinicians have been actively searching for an ideal cartilage implant that can reproduce the mechanical and biological properties of healthy, native cartilage. The inhibitory potential of some sulphated polysaccharides against various catabolic enzymes involved in joint degradation, such as hyaluronidases, elastases and matrix metalloproteases, was reported  and propThe altered joint environment and regenerative capacities make osteochondral regeneration even more challenging since this complex biologic process has to be sustained by a combination of mechanical and biological stimuli. Osteochondral defects affecting most of the condyle surface cannot yet be regenerated nor treated except using highly invasive approaches. Indeed, the implantation of metallic prosthesis is highly invasive and implies the ablation of a large amount of bone to assure the fixation. On the other hand, the existing regenerative devices have inadequate mechanical performance and insufficient bioactivity, hampering the re-growth of articular cartilage with a sliding surface and shock-absorption ability.To overcome these bottlenecks, new approaches should be pursued that might be powered using novel smart materials and innovative engineering methods. In this respect, marine-derived materials are eliciting great interest as bioactive molecules or to develop scaffolds for biomedical applications such as wound healing and as drug delivery systems. Such an interest is raised by the outstanding performance of marine-derived tissues in terms of mechanical responsiveness and of multiple biologic abilities including anti-inflammatory, anticancer and antioxidant properties ,6,7,8.Boosted by such an interest, marine biomaterials experienced various stages of development, starting from coralline bone grafts and then moving to polysaccharide and protein-based biomaterials . RelevanAmong proteins, collagen extracted from marine sources elicits great interest since it has an amino acid composition that is similar to human type I collagen ; in addiSuch considerations suggest that marine-derived materials are very promising compounds to be investigated for the development of multi-layered scaffolds for osteochondral regeneration. In this respect, the present review describes in more details the unique mechanical performance of the natural osteochondral unit and the relevance in reproducing such a performance to obtain effective osteochondral regeneration using scaffolds exhibiting high compositional and structural biomimicry. To this end, the development of multiphasic scaffolds is highlighted, as these are uniquely able to reproduce compositional and structural gradients typical of the different bony and cartilaginous compartments. On the other hand, the review focuses on the combination of bio-inspired mineralization processes and 3D manufacturing as promising technologies which might be synergistically combined to generate biomimetic multiphasic nanofibrillar structures, hierarchically organized from the nano to the macroscopic scale, as exhibited by the natural osteochondral unit.2 is recommended) or for patients over 35 [Therapeutic interventions on damaged osteochondral regions should not only tackle cartilaginous damage, but the overall joint environment: indeed, the subchondral bone is always damaged in advanced OA and it may be further compromised by osteoporotic or inflammatory conditions, greatly influencing the course of the disease and treatment success. Until now, most attempts to regenerate osteochondral defects, particularly if affected by OA, have not resulted in a stable and reproducible restoration of joint function ,27,28. V over 35 ,50,51,52 over 35 .To date, current cartilage scaffolds do not replicate the biomechanical performance of the natural osteochondral unit but mainly the ability to withstand compressive forces and to ensure sliding effects at the articular surface. These aspects are key in OC regeneration; however, most of the efforts in OC scaffold development have been focused only on the biological performance, leading to articular mechanical instability and failure.lamina splendens) accounts for extremely low friction and allows the transfer of multi-axial loads to the underlying subchondral bone, with the thin tidemark layer minimizing the stiffness gradient between the rigid bone and the more pliable cartilage [lamina splendens represents a crucial element in the maintenance of virtually null friction conditions due to its unique ultrastructure and the constant production of molecules exerting a lubrication effect. Changes in the cartilage surface lead to the increase in the friction and result in the transformation of orthogonal into tangential forces with progressive deformation of the tissue architecture. In this respect, a major limitation in the existing cartilage repair devices is their inability to reproduce such a surface structure giving low friction conditions and correct load-dependent behavior, thus resulting in the progressive degeneration of the surrounding cartilage with time. The ensemble of the self-lubricating ability of the cartilage surface and the dynamically adaptive mechanisms, such as the fluid pressurization effect, are believed to be at the base of the unique shock-absorption properties of the articular cartilage and the key for the health of the entire osteochondral unit.The articular cartilage and its supporting subchondral bone represent a functional unit which is of paramount relevance for the joint mechanical performance : the dynartilage . Going fartilage ,58. SuchWith this concept in mind, a general design that may drive the development of effective osteochondral scaffolds should recapitulate the composition and mechanically functional architecture of bony and cartilaginous tissues. This implies the realization of bioactive multiphase scaffolds, i.e., including both inorganic and organic compounds, engineered in such a way to reproduce the hybrid porous structure of the subchondral bone and the active mechanical responsiveness to multi-axial mechanical forces typical of the surmounting articular cartilage. In this respect, the outstanding physico-chemical, mechanical and biological properties of marine-derived compounds are very promising.While there is no single material or fabrication method that has proven to be superior in developing an OC scaffold, a multiphase scaffold seems to be the best approach to creating tailored environments to regrow each of the tissue elements within the complex OC unit.To date, one of the most promising strategies to develop osteochondral substitutes comprises the generation of heterogeneous scaffolds that are obtained through the combination of distinct but integrated layers corresponding to the cartilage and bony regions. Such a design should be based on the recognition of the different requirements to regenerate the cartilaginous and bony parts of an osteochondral defect, and simultaneously prevent the risk of delamination of different components if these are adjacent but physically separated. To overcome this problem, a biomineralization process was previously developed, focusing on obtaining hybrid constructs through supramolecular assembling and mineralization of collagenous proteins, inspired by the natural phenomena occurring in the formation of several biologic structures such as shells, exoskeletons of marine animals and including bone and osteochondral regions . The pro2+ and CO32− to induce the heterogeneous nucleation of the apatitic phase on the collagen fibers during their assembling [In detail, such a scaffold was built by stacking a bone-like mineralized layer produced according to the newly developed technique, an intermediate layer with a reduced amount of mineral phase to mimic the tidemark and an upper layer formed by collagen and hyaluronic acid, thus reproducing the cartilaginous environmental cues . The biosembling . The ressembling .In parallel, additive fabrication technologies have emerged in recent years, providing new tools for engineering living tissues, including bone and cartilage ,63. HoweVarious 3D printing technologies are currently in development, and their full capabilities are an area for potential improvement in the design of biomimetic OC scaffolds see . Howeverlamina splendens with its sliding surface is relevant to achieve resistance to shear forces, a key aspect to improving the biomechanical performance of the scaffold under loading [In the native tissue, the articular cartilage is continuous, with an average pore size of only 6 nm; calcified cartilage has pores within the unmineralized areas which accounted for ≈22% of the tissue; and subchondral bone has an overall porosity of 64–94% ,80,81. A loading ,94.From the material perspective in OC scaffold development, natural polymers are commonly used in hydrogel form where their polymer networks are capable of holding a large amount of water, thereby creating a fully hydrated 3D environment comparable to that of the natural ECM ,96. ThisOn the other hand, natural polymers typically possess weak mechanical properties, which can lead to deformation in load-bearing areas . NoticeaThe surface wettability of the polymeric network is strongly increased by the presence of biomimetic HA powder inside and on the surface of PCL pillars, thus favoring cell adhesion and colonization. The final scaffold was obtained by mimicking the three osteochondral layers as above described and infiltrating them inside the macro-porous PCL structure. The final scaffolds were implanted in critical knee osteochondral defects created in a group of 6 sheep. In spite of the presence of the Mg-HA/Col hybrid filling all the space inside the polymeric network, very promising to enhancing the bioactivity of the scaffold , abundanAdditionally relevant is polyethylene glycol (PEG), which is primarily used in the cartilage region of the scaffold ,117,118.In recent decades, the extrusion technology has been widely implemented to develop solutions for tissue engineering ,126.Considering the advantages and drawbacks related to different elective approaches for OC scaffold development, the association of bio-inspired assembling/mineralization processes and 3D fabrication techniques is a promising approach for a leap forward in the development of multi-layered scaffolds suitable for OC tissue regeneration.2+, substituting Ca2+ in dynamic equilibrium with the physiological environment, is known to stimulate the regenerative process, while the presence of Sr2+ affects the osteoblast–osteoclast cross-talk favoring the osteoblast activity, chiefly relevant to contrast osteoporosis [It is well-known that porosity, particularly when organized in hierarchical architectures, greatly influences the mechanical properties with reference to viscoelastic deformation in hybrid scaffolds. However, to achieve the cell chemotaxis and thus activate appropriate cell behavior yielding regeneration of healthy OC region with its outstanding mechanical performance, the chemical composition and scaffold biomimicry are also a key aspect. In fact, the appropriate choice of materials for OC scaffold permits the tuning of the bioactivity by exploiting biomolecular recognition by stem cells and chondrocytes; at the same time, however, the functional structural arrangement of constituting elements is a major aspect determining the mechanical performance of the regenerated OC region. To implement an appropriate synergy between chemistry and 3D architecture in OC scaffolds, marine-derived materials can be a relevant source of inspiration, particularly echinoderms endowed with smart connective tissues made of mutable collagenous proteins (MCP) able to dynamically modify the stiffness of the extracellular matrix. The biomimetic composition and structure could provide the OC scaffold with a high regenerative ability and mechanical competence as a result of the activation of chemical mechanisms modulating the interfibrillar cohesion and the elastic–plastic stress transfer dependent on the strength of the applied load ,128. As oporosis ,130. Besoporosis , due to For the cartilaginous layer, the use of MCP, prevalently formed of marine-derived mutable collagen, can be effective to achieve an enhanced mechanical performance. Indeed, contrary to mammalian-derived collagen, MCP does not require harsh processes for the extraction and purification of fibrils . In factIn association with MCP, marine-derived polysaccharides such as fucoidan (FUC) elicit great interest as a result of their excellent bioactivity and unique structure. Given the presence of negatively charged functional groups, FUC can be chemically linked to MCP by activating reactions between the sulfate group and free amines; this is suitable to achieve stronger inter-fibrillary interactions and marked increase in the compressive strength. This can be relevant to dynamically regulate interfibrillar shear stress and the fibril–fibril sliding under loading, in turn affecting the elastic–plastic stress transfer . As FUC In consideration of the above-reported, an interesting future perspective in the development of OC scaffolds with enhanced regenerative ability is given by the convergence of novel 3D fabrication technologies with materials endowed with unique bioactive and structural properties, such as marine-derived proteins and polysaccharides. Such an approach is promising as regards important advances in terms of still unmet mechanical and biologic performance of scaffolds for regenerative medicine, particularly addressing the regeneration of large, load-bearing OC defects .The present review focused on osteochondral regeneration as a still unmet clinical need from a material science perspective. In this respect, we investigated the existing literature, highlighting the most important aspects relevant for the achievement of clinically effective scaffold-based solutions. We thus highlighted the relevance of using marine-derived ingredients given their outstanding mechanic and biologic properties as a new concept guiding material scientists in the development of osteochondral scaffolds.In particular, we presented a rationale sustaining the combination of bio-inspired synthesis methods with advanced 3D manufacturing technologies, as a synergistic approach allowing the translation of the unique biomechanical and regenerative performances of marine-derived ingredients into new bioactive multiphase scaffolds for osteochondral regeneration. Firstly, mutable collagen proteins (MCP), characterized by adaptive interfibrillar cohesion, can be explored for their mechanical competence and high regenerative ability. In particular, MCP can be assembled and mineralized using ion-doped apatitic phases acting as bioactive chemical signals for stem cells, as well as promoting the rising of bio-stimulating piezo/tribo-electric currents, as occurs in the natural osteochondral region. On the other hand, the non-mineralized cartilage-like layer can be obtained by functionalizing MCP matrix with fucoidans, which are able to entrap water molecules and cross-link the MCP matrix, responding to applied mechanical loads. This ability is promising and represents the missing piece to reproduce the fluid pressurization effect, which is a key mechanism in ensuring the mechanical integrity and the health of the whole osteochondral unit.lamina splendens, and are able to prevent cartilage disruption caused by shear forces. Such a challenging achievement is considered a cornerstone for the development of osteochondral scaffolds effective for the regeneration of large, load-bearing defects.Finally, we highlighted the synergistic use of 3D manufacturing technologies to generate multi-scale, mechanically competent arch-like architectures able to cope with multi-axial compressive shocks. In this respect, electrospinning and electrowriting techniques are techniques of particular interest as they can be implemented to generate wear-free surfaces mimicking the Importantly, the inherent anti-inflammatory and antioxidant properties of marine-derived ingredients are of great interest to contrast osteoarthritis and osteoporosis-related degenerative diseases.


This paper reports the proceedings of a meeting convened by the Research Group on Thoracic Ultrasound in Older People of the Italian Society of Gerontology and Geriatrics, to discuss the current state-of-the-art of clinical research in the field of geriatric thoracic ultrasound and identify unmet research needs and potential areas of development. In the last decade, point-of-care thoracic ultrasound has entered clinical practice for diagnosis and management of several respiratory illnesses, such as bacterial and viral pneumonia, pleural effusion, acute heart failure, and pneumothorax, especially in the emergency–urgency setting. Very few studies, however, have been specifically focused on older patients with frailty and multi-morbidity, who frequently exhibit complex clinical pictures needing multidimensional evaluation. At the present state of knowledge, there is still uncertainty on the best requirements of ultrasound equipment, methodology of examination, and reporting needed to optimize the advantages of thoracic ultrasound implementation in the care of geriatric patients. Other issues regard differential diagnosis between bacterial and aspiration pneumonia, objective grading of interstitial syndrome severity, quantification and monitoring of pleural effusions and solid pleural lesions, significance of ultrasonographic assessment of post-COVID-19 sequelae, and prognostic value of assessment of diaphragmatic thickness and motility. Finally, application of remote ultrasound diagnostics in the community and nursing home setting is still poorly investigated by the current literature. Overall, the presence of several open questions on geriatric applications of thoracic ultrasound represents a strong call to implement clinical research in this field. In the last decade, bedside thoracic ultrasound has become extremely popular in the hospital care of acute patients with respiratory symptoms . In factThe diagnostic advantages of thoracic ultrasound are particularly emphasized in older patients . FrailtyThe emergence of the coronavirus disease 2019 (COVID-19) pandemic, which has particularly hit older patients with multi-morbidity and frailty, has allowed us to explore the use of thoracic ultrasound in other settings of geriatric care, especially in nursing homes –10. In fDespite these interesting applications, the use of thoracic ultrasound remains relatively limited in geriatric settings, as suggested by a survey conducted among Italian geriatricians before the pandemic emergency . ResearcTherefore, the aim of this narrative review is to identify knowledge gaps in geriatric thoracic ultrasound research and propose novel areas of development for optimizing the use of chest ultrasound in older patients and, ultimately, improving clinical care.The Research Group on Thoracic Ultrasound in Older Patients  of the Italian Society of Gerontology and Geriatrics was founded in 2018 with the aim of promoting knowledge and diffusion of thoracic ultrasonography in geriatric settings through educational and research initiatives. It is composed of clinicians and academicians with recognized expertise in thoracic ultrasonography, mainly of geriatric background but also with competence in emergency medicine, intensive care, and respiratory medicine. It also includes some residents in geriatric and emergency medicine with experience in thoracic ultrasonography.This paper represents the summary of the proceedings of a meeting of the GRETA Group held in May 2023 in Florence, Italy. Each member of the group was given the task of summarizing the literature state-of-the-art on specific topics related to the use of thoracic ultrasound in various fields of geriatric medicine Table , and ideDespite thoracic ultrasound has emerged as a highly sensitive and specific tool for diagnosing lung and pleural diseases and for guiding invasive procedures –20, a unThe research groups by Schmickl and Duggan have separately performed in vitro and in vivo studies to identify the optimal settings of ultrasound equipment in order to find a compromise between image quality and visualization of artifacts of diagnostic value, namely B lines, in thoracic ultrasound , 26. B lAnother open issue concerns the best methodology of ultrasound scan. The recent TUONO consensus has suggested that, in an intensive care setting, the 12-field study protocol  represents the best compromise between examination speed and diagnostic accuracy . This prWhen reporting examinations conducted with this protocol, the description of physiological findings, including pleural sliding synchronous with respiration and horizontal artifacts called A lines, could be omitted . ConversThe applicability of these ultrasound settings, protocols of assessment and methodology of reporting to older patients, especially those with frailty, cognitive impairment, and mobility disability, is still uncertain. More specifically, the use of standardized methods of ultrasound assessment and reporting could facilitate follow-up of acute and chronic respiratory illnesses, monitor aging of the respiratory system and, ultimately, assist clinicians in outcome predictions.Studies conducted in acute geriatric patients suggest that bedside thoracic ultrasound is superior to traditional chest radiographs, especially in terms of sensitivity, in detecting pulmonary consolidations caused by pneumonia , 35. PoiAspiration due to oropharyngeal dysphagia is also an important etiologic factor leading to pneumonia in older individuals , 38. AspAccording to different systematic reviews and meta-analyses, lung ultrasound (LUS) has a very good diagnostic accuracy in identifying lung parenchymal consolidations related to pneumonia in the emergency/urgency setting , 43. In However, it is unclear whether there is a typical ultrasonographic pattern associated with aspiration pneumonia in older individuals, allowing us to discern aspiration from other forms of pneumonia, including CAP, healthcare-associated pneumonia (HAP), or ventilator-associated pneumonia (VAP).To date, very few studies are available in the scientific literature with the specific aim of describing the ultrasonographic appearance of aspiration pneumonia. In one of these studies, conducted in a group of 17 dogs, a pattern similar to those present in humans with CAP  was found, with a typical localization at the level of the right anterior regions, suggesting that the presence of consolidations on LUS in these regions may be a sign of aspiration pneumonia . The onlDespite the interest in these studies, there is still uncertainty on how LUS can help clinicians to distinguish the etiology of pneumonia in older patients. It is also unclear whether specific LUS signs can help predict the clinical course of pneumonia, either caused by aspiration or infection, because studies on this issue are limited to anecdotical reports , 48. TheThoracic ultrasound is more sensitive and specific than X-rays in detecting pleural effusions, and is able to distinguish qualitative characteristics more effectively than CT , 50. DesVarious methods of quantitative estimation of pleural effusions are described in the literature , 52. MulIn a recent observational prospective study conducted in 188 patients with a diagnosis of heart failure, Lindner and colleagues developed a semi-quantitative ultrasound score ranging from 0 to 4 for each hemithorax  . The scoAnother major challenge in ultrasonographic evaluation of pleural effusions is represented by the differential diagnosis between exudative and transudative effusions . Older pRecent studies have examined echogenicity of pleural effusions using quantitative methods, such as the Hypoechogenicity Index (HI) and pixel density, which appear to correlate with the chemical and the physical characteristics of pleural effusions . Soni etNovel technological applications of ultrasound assessment of pleural effusions also include the use of contrast-enhanced ultrasonography (CEUS). In a recent prospective study, the utility of CEUS in guiding pleural drainage was evaluated . In compThe European Society for Medical Oncology 2022 Clinical Practice Guidelines for diagnosis, treatment, and follow-up of malignant mesothelioma do not include thoracic ultrasound among the clinical procedures recommended for the diagnostic approach to pleural malignancies and for monitoring patients with a history of asbestos exposure . Chest CTherefore, thoracic ultrasound may not be suitable for definitive diagnosis, but could play a role in the population screening of subjects at risk of mesothelioma. In a recent study, pleural abnormalities were detected on ultrasound or CT in 57% of 117 subjects with long history of asbestos exposure, and related to older age and duration of the exposure, but not to job or smoking habits . PositivThe use of color-flow Doppler (CFD) could be also implemented as an adjunct to the traditional B-mode ultrasonography to assist the diagnostic workflow of pleural lesions. Specifically, it could be used to identify vasculature within a lesion, because pleural malignancies are often supplied by the bronchial arterial system. According to one study, CFD has a specificity of 98% and a positive likelihood ratio of 27 in differentiating malign from benign pleural lesions . CFD couThoracic ultrasound can also serve as a guidance instrument when sampling suspected pleural lesions. According to a study by Sconfienza and colleagues , ultrasoThoracic ultrasound has already entered clinical practice in the evaluation of diseases causing interstitial syndrome, such as acute heart failure, pulmonary edema, primary or secondary pulmonary fibrosis, and COVID-19 pneumonia , 31. In In the last years, artificial intelligence (AI), especially machine learning (ML) and deep learning (DL) techniques, has emerged as an effective method in mining and integrating large-scale and heterogeneous medical data for clinical practice . As it iIn clinical practice, the common approach used to detect different lung ultrasound patterns is based on the counting and visual interpretation of artifacts. This approach was made official with the release of the Lung Ultrasound Score (LUS) in 2020, consisting of grading interstitial syndrome from 0 to 4 for each thoracic field, originally conceived for assisting the diagnosis of COVID-19 pneumonia, but applicable to all patients with interstitial syndrome . AlthougAutomatic quantification of B lines by AI has the potential to decrease inter-operator variability in ultrasound assessment of interstitial syndrome. Machine-assisted quantification of lung ultrasound artifacts, according to studies conducted in small groups of patients, generally performs well . HoweverInter-operator variability in thoracic ultrasonography represents an issue particularly for novice operators, although the learning curve is steep and a limited number of examinations is generally considered sufficient to guarantee an appropriate evaluation of common lung patterns . To dateMachine-learning algorithms could improve the accuracy of thoracic ultrasound assessments made by novice operators, especially regarding the quantification of interstitial syndrome severity. In a recent prospective multicenter study conducted on patients with decompensated heart failure, the quantification of B lines by an AI software on ultrasound images acquired by novice operators was compared with the quantification made by expert reviewers on the same images . The corFinally, AI could assist physicians in discriminating between similar ultrasonographic patterns of interstitial syndrome. In interstitial lung disease, B-line pattern is not homogenous and generally associated with irregular and thickened pleural line, whereas in heart failure, B lines generally appear more homogeneous, bright, and bilateral, with laser beam pattern and no alterations of the pleural line . Thus, tArntfield and colleagues evaluated the capacity of a convolutional neural network to discriminate between 612 lung ultrasound videos with similar appearance but different etiology  from 243 patients, in comparison with an expert evaluation by 61 physicians with at least 3 years of thoracic ultrasound experience . InteresThe diaphragm is the main respiratory muscle, and its dysfunction has been associated with prolonged mechanical ventilation and weaning failure , 91. RecFurthermore, recent studies have encouraged the use of US in the evaluation of diaphragmatic dysfunction (DD) as a reliable technique during an exacerbation of chronic obstructive pulmonary disease (COPD) and in predicting the response to non-invasive ventilation (NIMV) therapy , 96. In Despite muscle atrophy occurring in mechanically ventilated patients, other aspects could affect DD during an acute exacerbation of COPD (AECOPD). Some authors hypothesized that the progressive development of dynamic hyperinflation may cause a change in the geometry of the chest wall and diaphragm, with tidal breathing closer to the individual’s total lung capacity (TLC) . As suchOn the contrary, Similowsky et al.  showed tAt the current state of knowledge, no studies have investigated DD as a prognostic parameter during an AECOPD in undernourished patients with “respiratory sarcopenia”, the muscle wasting typical of advanced lung disease . This syThis aspect underlines the multiplicity of causes related to DD during COPD and the need of identification of early predictors of NIMV failure, in order to assist clinicians in stratifying the risk of negative outcomes and identifying the candidates to endotracheal intubation. Thus, early evaluation of patients under NIMV with worse survival prospects related to sarcopenia throughout a practical and low-cost tool as the diaphragmatic ultrasound could be extremely useful in clinical practice. Early identification of NIMV outcome predictors in patients with COPD who are experiencing acute hypercapnic respiratory failure consequent to exacerbation or pneumonia still remains a critical issue deserving to be further investigated even with the support of diaphragmatic ultrasound.Much of the knowledge on the respiratory involvement in COVID-19 derives from studies focused on the acute phase of the disease, and there are still few data available on the long-term respiratory sequelae of the infection. Subjective dyspnea and shortness of breath are among the main symptoms frequently reported in the long-COVID syndrome, but their correlation with pulmonary imaging and respiratory functional tests still remains to be evaluated , 106.Starting from the assumption that high-resolution chest CT is considered the gold standard for the evaluation of persistent interstitial lung disease , there aA study by Giovannetti et al., that included 38 patients previously hospitalized in a specialist respiratory Intensive Care Unit (ICU) for COVID-19, found a high degree of overlap of chest ultrasound and HRCT abnormalities of lung parenchyma on examinations performed three months after discharge . More reEven more recently, Barbieri et al. demonstrated a clear improvement in the ultrasound and CT findings, with excellent correlation between the two methods, in 232 post-COVID patients compared to baseline . InteresTherefore, as suggested by these studies and other expert opinions, lung ultrasound can be proposed as a reliable first-level method for the long-term follow-up of patients with previous COVID-19, reserving CT as a second-level method for specific cases –114. HowThe technique of providing health care services remotely through the use of information and communications technology (ICT) is known as telemedicine, a term first used in the 1970s. However, elementary forms of telemedicine have been in place since the invention of telephone, which was used by physicians to provide remote counseling for non-urgent cases , 117. ThThe massive development of ICT in the medical field and the parallel diffusion of ultrasonography in clinical practice have recently led to the development of tele-ultrasound, i.e., the use of ultrasound in conjunction with a telemedicine connection to another instructor or other means of communication . ThoraciDespite the promising results, the implementation of thoracic tele-ultrasound in the geriatric population encounters important barriers . First, Thoracic tele-ultrasound may be better applied in the nursing home setting, where healthcare professionals, not necessarily of medical background, could perform the examination, delivering images remotely to expert physicians able to give a diagnostic interpretation. The use of telemedicine in geriatrics has been studied, in fact, mainly in the nursing home setting, with promising results in terms of care organization, patient outcomes, and caregiver satisfaction , 129.Despite the established use in clinical practice for evaluation of respiratory failure and other acute respiratory illnesses, there are several open issues regarding the application of thoracic ultrasound, especially in the field of geriatric medicine. The main ones, identified by panel discussion among the members of the GRETA Group and discussed in the previous sections of this paper, are summarized in Table Thoracic ultrasound is a modern, accurate, quick, and inexpensive technique that has a great potential of improving the care of older patients with a wide array of acute and chronic conditions. The possible frontier applications of thoracic ultrasound, listed in Table


Optical coherence tomography (OCT) is an emerging imaging technique for diagnosing ophthalmic diseases and the visual analysis of retinal structure changes, such as exudates, cysts, and fluid. In recent years, researchers have increasingly focused on applying machine learning algorithms, including classical machine learning and deep learning methods, to automate retinal cysts/fluid segmentation. These automated techniques can provide ophthalmologists with valuable tools for improved interpretation and quantification of retinal features, leading to more accurate diagnosis and informed treatment decisions for retinal diseases. This review summarized the state-of-the-art algorithms for the three essential steps of cyst/fluid segmentation: image denoising, layer segmentation, and cyst/fluid segmentation, while emphasizing the significance of machine learning techniques. Additionally, we provided a summary of the publicly available OCT datasets for cyst/fluid segmentation. Furthermore, the challenges, opportunities, and future directions of artificial intelligence (AI) in OCT cyst segmentation are discussed. This review is intended to summarize the key parameters for the development of a cyst/fluid segmentation system and the design of novel segmentation algorithms and has the potential to serve as a valuable resource for imaging researchers in the development of assessment systems related to ocular diseases exhibiting cyst/fluid in OCT imaging. Optical coherence tomography (OCT) is a non-invasive high-resolution imaging technique that enables the visualizing of the characteristics of biological samples . It was However, manual segmentation of cyst/fluid in OCT images, which requires retinal expert identification, is prone to deviations in subjective interpretation and poor repeatability. Additionally, the large amount of imaging data per patient makes manual assessment extremely time-consuming. These challenges have motivated lots of researchers to propose automated algorithms for cyst segmentation/measurement to speed up the process and reduce human labor. Automated segmentation techniques to the cysts/fluid can provide ophthalmologists with valuable tools for improved interpretation and evaluation of retinal features, which is helpful for more accurate diagnoses and guided intervention. An illustration of cyst/fluid segmentation is shown in Recently, artificial intelligence (AI) has experienced significant growth with the advent of deep learning, offering a better representation of data . These aThe process of cyst segmentation can be broken down into three distinct steps, namely image denoising, retinal layer segmentation, and cyst/fluid segmentation. The initial image denoising step serves to eliminate speckle noise and enhance edge information in OCT images. Retinal layer segmentation identifies potential regions where cysts or fluids may exist, reducing the possibility of false positive errors in subsequent steps. Finally, the cyst/fluid segmentation step identifies the precise location of each cyst or fluid region within the OCT images. In a cyst segmentation system, all three of these steps are used to-gather to accurately and efficiently identify and segment cystic structures in OCT images.In this paper, we review existing works for retinal cyst/fluid segmentation in OCT images, with an emphasis on the importance of machine learning techniques. Given the critical role of image denoising and retinal layer segmentation in many cyst/fluid segmentation algorithms, we also review the state-of-the-art methods in these two areas and discuss their advantages and disadvantages. This review is distinct from other related works in the field, which focus only on the core cyst/fluid segmentation algorithm. We provide a holistic overview of each component of the cyst segmentation system, including image denoising, retinal layer segmentation, and cyst/fluid segmentation, as depicted in The main contributions of our study can be summarized as follows: Firstly, we present a comprehensive overview of the current state of retinal cyst segmentation in optical OCT and highlight the key findings of machine learning techniques. Secondly, we provide a summary of the publicly available OCT datasets for cyst/fluid segmentation. Thirdly, we discuss the challenges, opportunities, and future directions for the application of machine learning in retinal cyst segmentation. Overall, this review provides valuable insight into the development of new segmentation algorithms and the improvement of existing ones.The process of the cyst segmentation system can be divided into three steps, which includes image denoising, retinal layer segmentation, and cyst/fluid segmentation. The image denoising step removes speckle noise and enhances the edge information in the OCT images. The retinal layer segmentation step identifies the potential regions where fluid or cysts may exist and reduces the likelihood of false positive errors in the subsequent step. The cyst/fluid segmentation step finally locates the position of each cyst or fluid region in the OCT images.The limited spatial bandwidth of the interference signals in OCT imaging lead to the occurrence of speckle noise, which significantly hinders subsequent analysis and segmentation in OCT. The dual role of speckle noise as both a source of noise and a carrier of information regarding tissue structure presents challenges in accurately identifying the boundaries of retinal layers and other retinal features in OCT, such as cysts/fluids. To address these difficulties, a number of techniques for speckle denoising have been proposed in the literature, including spatially adaptive wavelet filter , BayesiaThe spatially adaptive wavelet filter (SAWF)  is a wavWong et al.  presenteThe use of anisotropic diffusion-based methods ,20,21 haTaking advantage of recent developments in deep learning, various learning-based approaches have been put forth to tackle the problem of speckle noise in OCT images. Saba et al.  introducRetinal layer segmentation in OCT is a critical pre-processing step in retinal cyst/fluid segmentation, as illustrated in Based on statistical models, traditional methods for segmenting retinal layers in OCT images involve partitioning the image into homogeneous segments according to characteristics, such as intensity or texture. For instance, Hee et al. segmented the entire retinal layer and the nerve fiber layer by the variations of pixel gray value of pixels . TraditiRetinal layer segmentation in OCT images has also been investigated in deformable-model-based methods. Lu et al. formulated the OCT datasets as probability density fields and applied a level set model to outline the retinal nerve fiber layer . More spGraph-cut-based algorithms have been explored for layer segmentation in OCT. Garvin et al. introduced a graph-theoretic segmentation approach that optimized the simultaneous segmentation of multiple 3D surfaces through the application of the minimum cut algorithm in a graph . This meIn the realm of classical machine learning, Karri et al. presented a structured random forest to detect eight retinal layer boundaries employing traditional graph theory for AMD . The metThe segmentation of cysts/fluid in OCT images is still challenging, even after the application of denoising and layer segmentation techniques. This was due to several factors that complicate the efficacy of conventional segmentation algorithms. The intensity patterns observed in these images are a result of light absorption and scattering in the retinal tissue, leading to a decrease in the intensity of homogeneous areas with increasing depth. This made it difficult for segmentation algorithms that rely solely on intensity variations to accurately segment the cysts/fluid. The presence of low optical contrast in certain regions of the OCT images caused by optical shadowing of retinal blood vessels further exacerbates the problem. Additionally, motion artefacts and sub-optimal imaging conditions negatively impacted the accuracy of segmentation techniques.To address these challenges, various algorithms for cyst/fluid segmentation in OCT images have been proposed and can be grouped into two main categories: classical machine learning methods and deep-neural-network-based learning methods.Classical machine learning is a branch of machine learning that encompasses traditional algorithms that were developed prior to the advent of deep learning. These algorithms are based on mathematical and statistical models and are widely used for various tasks, including regression and classification. The most common classical machine learning algorithms include K-nearest neighbor (k-NN), support vector machine (SVM), random forest, kernel-based regression, graph search-graph cut method, and AdaBoost. K-NN is a simple and intuitive algorithm that classifies a new data point based on the classes of its k nearest neighbors . SVM is The majority of classical machine-learning-based works in the field of cyst segmentation adopt a simple classification approach that leverages hand-crafted or texture-based features. The typical framework of these methods consists of four stages, including pre-processing, retinal layer segmentation, cyst segmentation, and post-processing  as illu. The objInitially, due to the limitations of data and computational power, simple methods were employed by researchers in the field of classical machine learning for cyst segmentation. In 2010, Quellec et al.  proposedRecently, the increased involvement of researchers in the field of OCT segmentation has led to the publication of several annotated OCT datasets. In 2015, the MICCAI conference hosted the OPTIMA Cyst Segmentation Challenge, releasing an OPTIMA dataset comprising 30 volumes from four different OCT devices used in ophthalmology . On the basis of this dataset, Gopinath et al.  proposedThe application of graph theory in image segmentation has gained significant attention in the field due to its ability to effectively represent the relationships between pixels or voxels in an image. Chen et al.  investigThe utilization of deep learning for automated segmentation in the field of medical imaging has garnered substantial interest. Compared with classical methods, these methods need a more extensive dataset, do not require human-engineered features, and simplify the system design by training an end-to-end classifier. Deep neural networks simplify the entire system design, as a single neural network is capable of detecting retinal-layer-bound cysts and segmenting cysts. Since 2017, various CNN and FCN architectures have been proposed for the purpose of cyst/fluid region segmentation in OCT imaging. FCNs, a subcategory of CNNs, are specifically designed for semantic segmentation tasks, where the objective is to assign a label to each pixel in an image. The structure of FCNs is similar to that of CNNs, yet it employs a unique architecture that maintains the spatial information of the image throughout the network. The utilization of CNNs in image recognition tasks offers several advantages, such as the capacity to automatically learn and extract features from data, the ability to handle high-dimensional data such as images, and the capability to generalize effectively to new data. FCN benefits from the strengths of CNNs and further allows for handling dense prediction tasks, such as semantic segmentation.Chen et al.  used a fIn recent years, more complex network structures were applied for the retinal fluid/cyst segmentation. A CNN-based model based on the DeepLab framework was presented by Wang et al.  for the In addition to the network structures, revisions of loss functions have also been considered to improve the segmentation performance. Liu et al. used an uncertainty-aware semi-supervised framework to tackle the issue of small fluid area detection and to prevent close regions from being identified as a unified entity. The framework consists of a teacher network and a student network that share a common architecture and employ three decoders to predict the probability map, contour map, and distance map of the image, thus enabling segmentation . In lighAnother area of research in deep learning for segmentation is based on the U-Net architecture. This architecture, a well-established type of FCN, is frequently utilized for the segmentation of medical images. The U-Net design features a contracting path and an expanding path, which facilitates the localization of features with precision and the training of the network in an efficient manner. This unique design has proven to be effective in a variety of biomarker segmentation research applications.In the study conducted by Roy et al. , a novelRecently, new modules were implemented in the U-Net construction. Chen et al.  introducThe process of training a cyst/fluid segmentation model requires the use of annotated OCT datasets, where the location of fluid in each image is labeled by experts. This labeling process is labor-intensive, which is why there are numerous public OCT datasets available for use. 3 and is centered on the macula. The dataset has been further divided into 15 training and 15 testing volumes, which have been chosen to reflect the broad range of scans encountered in clinical settings, as well as the diverse appearances and distributions of IRF. The IRF in the training data was annotated by two expert graders at the Medical University of Vienna.The OPTIMA dataset  was madeThe RETOUCH dataset  was deriThe DME dataset , made avThe UMN dataset  was gathOCT is a non-invasive imaging technique that utilizes the principle of interferometry to produce cross-sectional views of the retina. The technology has a resolution capability of at least 10–15 microns, which has led to its widespread adoption as an ophthalmic diagnostic tool. OCT was first introduced in 1991 and has since been used to image various ocular conditions, including DR and AMD. The versatility of OCT has extended beyond ophthalmology, as it has been utilized to image non-transparent tissues in other fields. In ophthalmology, it has become a crucial tool in the diagnosis, prognosis, and management of various ocular conditions. The non-invasive imaging capability of OCT to produce detailed images of the retina non-invasively has revolutionized the way ocular conditions are diagnosed and treated and has facilitated early detection and effective management of conditions. OCT’s precise and reliable images have made it a critical tool in advancing ocular medicine. Its ability to non-invasively image the retina has enabled a greater understanding of retinal structure and function, leading to improved patient outcomes.The implementation of an automatic cyst/fluid segmentation system has the potential to greatly aid ophthalmologists in their research endeavors. For example, in a study conducted by Pennesi. et al. , an autoFor the methods we reviewed, the classical machine learning or statistic-based methods require human-designed features. Still, they have the benefits of explainability for results and flexibility for a small dataset. The deep-learning-based method is more powerful but data-hungry. The cyst segmentation framework has three components: image denoising for removing the speckle in OCT, layer segmentation for finding the research region of the cyst, and cyst segmentation for finding the cyst region. These three parts are designed independently. The perspective of our review provides a research direction to test each possible combination of these components and find the most suitable method in each step. On another hand, with the benefits of deep learning, numerous works are designed as an end-to-end system, which only includes one neural network to handle tasks from all three components. They need fewer computational resources and avoid the error propagation from the layer segmentation to the cyst segmentation. In the current situation, no specific evaluation exists to determine which types of methods are superior and which should be investigated in the future.We presented a comparison of the performance of classical machine learning methods and deep learning algorithms. Classical machine learning approaches, such as González  and GopiIn the domain of deep learning, researchers commonly favor the utilization of public datasets, with the RETOUCH dataset being the prevailing choice. Among the deep learning algorithms evaluated in this study, faster R-CNN by Chen et al. , modifieThe advances in AI/machine learning and its applications in ophthalmology have brought significant benefits, but also present several limitations. One of the limitations is the lack of integration of domain knowledge in current OCT cyst segmentation algorithms. Typically, deep neural networks are trained using large amounts of supervised data. However, manual annotation of these datasets is a tedious and time-consuming process, particularly in the medical field where specialized experts are required. For cyst segmentation, doctors have concrete domain knowledge and rules to distinguish the cyst area in the OCT. However, most current works ignore this part, and the whole learning structure is designed purely from raw data. So, encoding this knowledge is a more scalable solution which potentially allows for finding better models. The techniques can be involved in the design of the algorithms: Bayesian-style learning method and knowledge-based learning.Another limitation is the volume and diversity of data available for training. Existing machine learning algorithms are highly data-dependent, and while publicly annotated cyst datasets are available, they are not enough to train powerful neural networks. As a result, techniques, such as transfer learning and few-shot learning, should be employed to enhance the size and quality of training datasets. Furthermore, current datasets are often lacking in representation of diverse ethnicities and may not be inclusive of all relevant features. To maximize the generalizability and applicability of deep learning algorithms, it is important to train these algorithms on data obtained from various hardware devices and not limit them to a specific one. As the application of AI in healthcare continues to grow, the exchange of medical information between patients and physicians through it is becoming increasingly prevalent. This highlights the importance of balancing data privacy and data utilization efficacy. One solution to this challenge is the use of federated learning, which enables AI institutions/hospitals to collaborate in a cooperative manner, utilizing data from multiple sources while avoiding privacy issues. Federated learning is a novel approach to training machine learning algorithms, which diverges from the centralized transactional approach. In federated learning, algorithms are trained on multiple decentralized servers/hospitals, each containing local data samples, without the need for data exchange between servers/hospitals. This novel approach to machine learning provides a solution to the data privacy and data utilization efficacy challenges.Lastly, current OCT cyst segmentation algorithms lack explanation and trustworthiness, which is a critical issue in fields affecting people’s health. It is imperative to understand the reasoning behind these decisions, as this could provide insight into the rules used by the models and prevent potential unintended consequences. Thus, the development of human-understandable explanations for these models is a crucial aspect of the practical implementation of artificial intelligence in healthcare. In the case of the cyst and fluid segmentation, providing explanations of the segmentation results can increase the trustworthiness of the recognition results. Most of them are viewed as black boxes with unknown mechanisms. It is essential to understand why decisions are made, which helps us understand the rules the models use to make decisions and prevent potentially unexpected situations from happening. Generating human-understandable explanations for the models is an essential topic for using AI in the real world. In the cyst/fluid segmentation, the explanation of the segmentation made the recognition results more trustworthy for doctors.OCT images are increasingly used in the diagnostic and prognostic of retinal disease. It is essential to evaluate retinal cysts/fluid for the purpose of determining initial diagnosis and retreatment. However, the manual evaluation of large OCT datasets is time-consuming and error prone. In order to effectively process the increasing amount of images generated by modern OCT devices and provide accurate cyst or fluid detection, automated learning-based methods are required. Ideally, the method should not be restricted to one disease or device and could be applied to a wide range of diseases.In this study, we reviewed the typical existing works in OCT cyst/fluid segmentation, while emphasizing the significance of machine learning applications. We delve into the three fundamental steps of the problem, including image denoising, layer segmentation, and cyst segmentation, highlighting the most prominent state-of-the-art algorithms for each step and comparing their efficacy. Moreover, the challenges, opportunities, and potential avenues for future growth in the application of AI in OCT cyst segmentation were also explored. The insights gained from this review can serve as a valuable resource for those seeking to apply the current segmentation technique for disease diagnosis or advance the field through novel algorithmic designs.