The treatment landscape for pediatric cancers over the last 11 years has undergone a dramatic change, especially with relapsed and refractory B-cell acute lymphoblastic leukemia (ALL), due to the introduction of chimeric antigen receptor-T (CAR-T) cell therapy. Because of the success of CAR-T cell therapy in patients with relapsed and refractory B-cell ALL, this promising therapy is undergoing trials in multiple other pediatric malignancies. This article will focus on the introduction of CAR-T cell therapy in pediatric B-cell ALL and discuss past and current trials. We will also discuss trials for CAR-T cell therapy in other pediatric malignancies. This information was gathered through a comprehensive literature review along with using first hand institutional experience. Due to the potential severe toxicities related to CAR-T cell therapy, safe practices and monitoring are key. These authors demonstrate that nurses have a profound responsibility in preparing and caring for patients and families, monitoring and managing side effects in these patients, ensuring that study guidelines are followed, and providing continuity for patients, families, and referring providers. Education of nurses is crucial for improved patient outcomes. In April 2012, the treatment landscape for pediatric relapsed B-cell acute lymphoblastic leukemia (ALL) was transformed. Using targeted immunotherapy, the first pediatric patient with relapsed B-cell ALL was infused with anti-CD19 chimeric antigen receptor-T (CAR-T) cells.In patients with relapsed and refractory malignancies, cancer becomes more difficult to treat due to dose-limiting toxicities and chemotherapy resistance, indicating the need for novel therapies such as CAR-T cell therapy. Ongoing research in genetics, disease biology, and immunology has impacted the discovery of effective and less toxic targeted therapy in pediatric cancer.CAR-T cell therapy is a form of targeted immunotherapy that uses tumor-specific antigen recognition. Chimeric antigen receptors are customized receptors that target specific antigens on a malignant cell.CAR-T cells have shown remarkable clinical activity in B cell cancers, and the current Food and Drug Administration (FDA)-approved CAR-T cell product in pediatrics is directed at relapsed and refractory B cell ALL. With CAR-T cell therapy becoming more widely used and with the introduction of CAR-T cell trials in other pediatric cancers, nurses will be at the forefront of caring for these patients both inpatient and outpatient. They will be responsible for educating patients and their families and providing care to these patients during the CAR-T cell experience.Pediatric ALL is the most common cause of cancer in children.Due to the risk of dose-limiting toxicities and chemotherapy resistance, research in targeted immunotherapy led to phase 1 and phase 2 CAR-T cell trials. A phase 1-2a single-center anti-CD19 CAR-T cell trial for pediatric and young adult patients with relapsed or refractory B-cell ALL showed promising results with a complete response (CR) rate of 93%.These pivotal phase 2 global, multisite trial response rates and safety results were integral in leading to FDA approval of tisagenlecleucel, the first commercialized anti-CD19 CAR-T cell product, in August 2017 for the treatment of pediatric relapsed or refractory B cell ALL. The first report of tisagenlecleucel in the real world shows similar response rates and safety compared with the phase 1-2 a single center trial and the phase 2 global, multisite trial. With a median follow-up of 13.4 months, the initial CR rate was 85.5% and the 12-month duration of response was 60.9%. Cytokine release syndrome (CRS), a systemic inflammatory response involving elevated cytokines in response to immune system activation, occurred in 55% of the patients. Immune effector cell-associated neurotoxicity syndrome (ICANS) was seen in 27% of patients.Unfortunately, patients can relapse after anti-CD19-directed CAR-T cell therapy in one of two ways. A patient can relapse with CD19-positive B cell ALL, meaning that the ALL still expresses the CD19 antigen. This often happens when CAR-T cells have short-term persistence; therefore, disease surveillance is gone. Short term persistence can be related to immune-mediated rejection or CAR-T cell exhaustion. CD19-negative relapses can happen due to the loss of expression of CD19 on the tumor cell surface. Patient outcomes for B-cell ALL relapse post-anti-CD19 CAR-T cell therapy are poor.B cell aplasia, an on-target/off-tumour toxicity following CAR-T cell infusion, is used as a marker for CAR-T cell persistence. Loss of B-cell aplasia indicates loss of CAR-T cell persistence.Most CAR-T cells contain an extracellular antigen-binding domain that is derived from a murine monoclonal antibody, possibly leading to an antimurine immune response causing CAR-T cell rejection. Studies have been done and are ongoing looking at replacing the murine component with a humanized component to avoid this immune rejection and improve long-term persistence.For some patients, successful collection and manufacturing of T cells is extremely difficult due to multiple factors including heavy chemotherapy pretreatment, a high level of circulating blasts, and age at diagnosis. A study showed that naïve and early memory T cells in the collection product correlate with successful CAR-T cell performance and that deficits in naïve T cells can be seen in patients who have been heavily pretreated, contributing to harvesting and manufacturing failure. There is also a concern about the quality of T cells collected in patients with infant ALL.Universal CAR-T cells are products using T cells collected from healthy allogeneic donors. These cells are also referred to as “off-the-shelf” CAR-T cells. This is a promising therapy for patients who have had unsuccessful T cell collections or who have a highly aggressive disease and may have disease progression before an autologous CAR-T cell product is ready.A universal CAR-T cell trial that enrolled patients aged 9 months to 62 years from 2016 to 2018 showed that universal CAR-T cell therapy is feasible. The median time from trial consent to beginning lymphodepletion was 11 days. This trial included 7 pediatric patients and 14 adult patients with relapsed or refractory B cell ALL. CRS was experienced by 91% of the patients, ICANS by 38%, and GVHD by 10%. Viral infections, including cytomegalovirus, adenovirus, human metapneumovirus, and BK virus, were seen in some patients. Sixty-seven percent achieved CR 28-days post CAR-T cell infusion, and 71% of the responding patients proceeded to stem cell transplant (SCT).As previously mentioned, patients who relapse post anti-CD19 CAR-T cell therapy can relapse with CD19-negative disease. Evidence shows that around 50% of patients receiving anti-CD19 CAR-T cell therapies may relapse within the 1st year.The results of a phase 1 anti-CD22 CAR-T cell trial were recently released. Fifty-eight patients were infused, 87.9% after receiving anti-CD19 CAR-T cells. CRS occurred in 86.2% of patients, and mild neurotoxicity occurred in 32.8% of patients. The complete remission rate was 70%. Seventy-five percent of these patients eventually experienced relapse. It was noted that patients who received prior CD22 targeted therapy had shorter remission durability and decreased MRD negative complete remission rates.Anti-CD22 CAR T cell trials for relapsed and refractory B cell ALL showing anti-leukemia responses have opened the door to using dual-targeting CAR-T cells. A central belief in the initial treatment of ALL is to use multi-agent chemotherapy to avoid relapse due to drug resistance.Dual CAR trials targeting both CD19 and CD22 have been done and show that there is no increased toxicity. Results from a dual CAR-T cell trial targeting CD19 and CD22 were recently released. Two hundred and twenty-five patients younger than 20 years were enrolled in a phase 2 trial, and 194 patients were infused. A complete response rate of 99% was seen. Event-free survival (EFS) at 12 months was 73.5%, and overall survival at 12 months was 87.7%. Improved EFS was seen with patients proceeding to SCT and in patients with B cell aplasia, indicating CAR-T cell persistence lasting longer than 6 months. CRS occurred in 88% of patients and ICANS in 20.9%. Three deaths occurred during CRS and/or ICANS.,Most pediatric patients with mature B-cell NHL have excellent prognoses with chemotherapy. Relapses are rare, but relapsed disease has decreased chemosensitivity and a dismal prognosis.Results from a phase 2 global, multicenter study using tisagenlecleucel in pediatric patients with relapsed or refractory B-cell NHL showed safety and efficacy. ORR was encouraging, although better in patients with large B-cell lymphoma (46%) vs. Burkitt lymphoma (20%). Neurologic events occurred in 27% of patients, and CRS occurred in 70% of patients.Developing successful CAR-T cell therapies for non-B-cell hematologic cancers has been more challenging. Some challenges have included finding tumor-specific antigens to target and avoiding severe, unacceptable on-target/off-tumor toxicities that attack normal, healthy cells. T cell aplasia as a result of anti-T cell ALL CAR-T cells and myeloid aplasia as a result of anti-AML CAR-T cells are examples of unacceptable long-term off-tumor toxicities.Despite the success seen with CAR-T cell therapy for B-cell ALL, the development of successful strategies for T-cell ALL has been more difficult. T-cell ALL accounts for 10%–15% of cases of ALL.,There are unique challenges with CAR-T cells for T cell ALL including CAR-T cell product contamination with T lymphoblasts, CAR-T cell fratricide (the self-killing of CAR-T cells), and the on-target/off-tumor toxicity of T cell aplasia, leading to life threatening immunodeficiency.Early-stage trials are being done using autologous and universal CAR-T cells. A recent phase 1, human universal anti-CD7 CAR-T cell trial has shown some promising results. The use of a universal CAR-T cell from a healthy donor alleviates the problem of product contamination with leukemic cells. Twenty patients were infused. There were no dose-limiting toxicities. The CAR-T cells expanded, 90% of patients achieved a complete response (CR), and this was proven to be a safe therapy.,Relapsed and refractory AML remains challenging to treat due to chemotherapy resistance. The only potential curative therapy for this patient population is SCT; therefore, alternative therapies are needed for these patients.,Studies are ongoing to create CAR-T cells that minimize the severity and duration of myelosuppression in relapsed and refractory AML.,There are several barriers to solid tumor CAR-T cells that are distinct from B-cell ALL. These barriers include identifying tumor-specific antigens, heterogenous expression, inefficient trafficking, and the immunosuppressive tumor microenvironment (TME). The most ideal tumor-specific antigen to target is one that is highly expressed on tumor cells but not expressed on normal cells.,Tumor-specific antigens in solid tumors can be very individualized. This individualization is often due to tumor-specific mutations that help evade the immune system.The TME can also impede the effectiveness of CAR-T cells. The micro-environment surrounding solid tumors consists of a physical barrier of stromal cells, immune cells, and inflammatory cytokines, as well as metabolic barriers that include hypoxia and nutrient starvation.•GD2 is a disialoganglioside that is highly expressed in neuroblastoma cells. It is also expressed on neurons, peripheral nerves, and skin melanocytes.•,GPC2 is a protein that plays an important role in the growth and differentiation of axons. It is highly expressed in neuroblastoma cells but has low expression in normal cells. Murine preclinical trials have shown encouraging results for disease regression without toxicity.•,B7–H3 is a checkpoint inhibitor highly expressed in neuroblastoma cells but restricted in normal cells. It is involved in tumor immune evasion and tumor promoting signals.•,LICAM (CD171) is overexpressed in neuroblastoma cells but has limited expression in normal cells. Phase 1 trials are ongoing. They have been shown to be safe with toxicities including skin rash and transient hyponatremia, but as with other CARs for neuroblastoma, they are showing limited response.Neuroblastoma is the most prevalent extracranial solid tumor in children. About 50% of patients present with metastatic disease. Though overall survival for high-risk patients has improved, it is still around 50% with chemotherapy, autologous peripheral stem cell transplant, radiation, and immunotherapy. Therefore, just as in relapsed and refractory B-cell ALL, novel therapies are needed. Neuroblastoma cells tend to be quite heterogenous; they have low expression of HLA class 1 antigens, allowing the tumor to evade the immune system, and the TME is quite hypoxic, making it highly immune suppressive.,Brain tumors are the most common non-hematologic cancers in children and are the leading cause of pediatric cancer deaths.,There are specific challenges that are unique to brain tumors including the blood brain barrier, the potential for on-target/off-tumor toxicity, and tumor inflammation associated neurotoxicity (TIAN).Brain tumors that are located in the brainstem, spinal cord, and thalamus are sensitive to edema from inflammation.•GD2 is highly expressed on diffuse intrinsic pontine gliomas (DIPGs) and other diffuse midline gliomas (DMGs). It is also expressed on neurons, peripheral nerves, and skin melanocytes.•B7–H3 (CD276) is a transmembrane protein highly expressed in brain tumors such as ATRT, DIPG, DMG, medulloblastomas, and high-grade gliomas.•(HER2) is found in glioblastomas. It is also expressed in epithelial tissues, skin, and muscle.Human epidermal growth factor receptor 2 Another difference in brain tumor CAR-T cells is the possible delivery mechanisms. Administration can be intravenous, intrathecal/ventricular, or local/regional (intratumor). Systemic administration may have limitations with CAR-T cell delivery to the tumor bed due to the blood-brain barrier. To overcome this barrier, higher doses may need to be used, which could lead to a higher risk of CRS.52CAR-T cell therapy is an evolving treatment used for hematologic malignancies and, more recently, solid tumors. The patients who receive this therapy have refractory or relapsed disease. These patients and their families experience a significant amount of stress coping with the diagnosis, in addition to traveling far for this therapy and often leaving support systems at home. Therefore, a multidisciplinary team approach is needed to provide seamless, safe care and to guide them through this process. At our institution, this team includes intake coordinators, nurse navigators, insurance specialists, inpatient and outpatient nurses, physicians, nurse practitioners, social workers, psychologists, physical and occupational therapists, nutritionists, and child life specialists. The complex care of these patients also involves successful collaboration with other departments in the hospital including interventional radiology, anesthesia, apheresis, the emergency department, the intensive care unit (ICU), neurology, and the cell and gene therapy lab. A strong research team is necessary to enable these trials to continue, and this team includes clinical research coordinators, data coordinators, research nurses, and a program manager. Clear and open communication is critical for a successfully functioning team.Since this is an evolving therapy, it requires specialized nursing care and knowledge. Oncology nurses play a crucial role in the care of patients receiving CAR-T cell therapy from the very beginning with patient referral through the monitoring of long-term effects. At our institution, these nursing roles include nurse navigators, apheresis nurses, research nurses, inpatient bedside nurses, outpatient nurses, and nurse practitioners, all working collaboratively to meet the unique needs of this complex population of patients.Nurses are integral to the CAR-T cell process and the success of these programs. As nurses, one of our biggest roles is that of educators. Patients and families are often very knowledgeable and have thoughtful questions regarding this therapy; therefore, nurses need to be educated and aware of the latest updates. We are responsible for the anticipatory guidance of patients and families, the education of our own staff, and those from referring institutions. Nurses need to be educated regarding who is eligible, what it is, how it works, and the short- and long-terms side effects to be able to provide education for patients and families. At our institution, we encourage our nursing staff to attend conferences at both the local and national levels. Attending conferences gives them the opportunity to expand their knowledge on CAR-T cell therapy and share what they have learned when they return. Locally, we share our knowledge through bi-annual nursing education days, where information is formally presented to nursing staff via content experts. These education days offer nurses protected time away from the unit to learn. The information is organized, and there are ample opportunities to ask questions and have educational discussions. Lectures and in-services regarding CAR-T cell therapy are also given throughout the year, especially when new trials are opening. Lectures are recorded so that new staff can watch them. Lectures are given by nurse practitioners to master's degree nursing students at the adjacent nursing school so that they are prepared when they graduate. Nurses and nurse practitioners educate and share information with colleagues at other institutions through presentations at national and international meetings, by presenting posters, and by publishing articles. As nursing pioneers in this field, it is important for us to share what we know and what we have learned.With anticipatory guidance, our hope is to decrease anxiety and stress for patients and their families. Education is family-centered, age-appropriate, and requires an individualized approach. The establishment of a relationship with consistent nurses and nurse practitioners allows for the development of trust and comfort. At our institution, we use each interaction with patients and families as a timepoint for education. In most cases, this has been a long and complicated journey, and patients and their families have spent many years at their home institution being treated for their cancer. Families and children are often stressed by the new routines at the CAR-T cell institution, such as central line care and clinic processes. To help alleviate this stress, families are given documents that include a map for navigating the hospital, oncology-specific information like inpatient routines if admitted, phone numbers to call the clinic and to call overnight and weekends, reasons to call, written material describing CAR-T cell therapy and how it works, and suggested websites to read if they are interested.Patients travel from all over for this therapy, and they interact with nurses throughout their entire journey. Nurse navigators are often the first contact. They are integral in collecting information to determine eligibility and maintaining communication with referrers and families. They need to discuss with the patient and family what to expect with the first visit including CAR-T cell consent conversation and T cell collection via apheresis, which often involves an apheresis catheter placement. Apheresis nurses provide education regarding T cell collection and are experts in the collection process.Outpatient clinic nurses are the first nurses to meet patients and families in person and play a role in lymphodepleting chemotherapy education and management of side effects. Outpatient and inpatient oncology nurses are responsible for reviewing and discussing the potential side effects and monitoring for them as well. Depending on the institution, the inpatient or outpatient nurse monitors the patient on the infusion day for any infusion related complications such as anaphylaxis. These nurses are also integral in assessing for the first signs of CRS and neurotoxicity.Bedside oncology nurses play a critical role in monitoring and assessing patients for early signs and symptoms of CAR-T cell toxicity and are at the forefront of providing supportive care.Cytokine release syndrome is the most common toxicity associated with CAR-T cell therapy. CRS is a systemic inflammatory response involving elevated cytokines that occurs with immune system activation. Fever is the hallmark sign of CRS, and symptoms can appear similar to those of an infection. In many patients, CRS is mild, and patients present with flu-like symptoms, including fever, chills, tachycardia, myalgias, fatigue, and headache. In contrast, other patients develop more fulminant CRS with multisystem organ failure.In addition to CRS symptoms, nurses need to be educated on how to carefully assess patients for neurologic toxicities, including headaches, confusion, tremors, ataxia, dysphasia, and seizures. Immune effector cell-associated neurotoxicity of any severity occurs in 20–60 percent of patients treated with CAR-T cell therapy.The nurse's role in the care of patients at risk for neurotoxicity after CAR-T cell therapy includes frequent assessments, symptom management, coordination of care, and timely escalation of care for patients experiencing sequelae of therapy. If signs and symptoms are recognized, neurologic assessment should occur more frequently to identify further neurologic deterioration. Supportive care includes reorientation, respiratory support, and the treatment of coagulopathies. In most cases, symptoms are transient and don't result in long-term deficits. With prompt recognition and appropriate supportive interventions, the acute symptoms of ICANS are reversible in nearly all patients.At our institution, we work very closely with our colleagues in the ICU and emergency department. We share important information about the care of our CAR-T cell patients through our unit-based nursing experts. When patients are admitted to the hospital for CAR-T cell therapy or for post-infusion complications, our unit-based clinical nurse experts reach out to the experts on the unit caring for the patient. We help to coordinate care, review side effects of chemotherapy, provide anticipatory guidance surrounding common post-infusion complications, and offer our expertise for educating and supporting patients and their families.Early collaboration with the ICU is essential for patients developing severe CRS and ICANS. Nurse practitioners and nurses play a central role in identifying early signs of severe CRS and consulting the critical assessment team to facilitate the safe transfer of patients to the ICU if needed. Another means of collaborating with our colleagues in the ICUs is by making our high-risk patients “medical watchers”. By identifying these patients as watchers, our critical care outreach team, which consists of an ICU physician, ICU nurse, and respiratory therapist, is made aware of the patients who are at potential risk for clinical deterioration. They round on these patients twice per day and will follow their clinical status from afar. This offers both teams an opportunity for ongoing collaboration, early detection, and a smoother transfer of care should the patient become acutely and/or critically ill.Because anti-CD19 CAR-T cells are targeted to destroy cells that express CD19, healthy B cells are also impacted. Persistence of CAR-T cells leads to B-cell aplasia, resulting in hypogammaglobulinemia, which increases the risk of infection and viral reactivation. Almost all patients who respond to anti-CD19 CAR-T cell therapy develop B cell aplasia after infusion. Immunoglobulins should be monitored and intravenous immunoglobulin replacement given until B cell function has been restored.In addition to outpatient and bedside nurses, research nurses are extremely important to the functioning of the CAR-T cell team, especially in this current era of multiple phase 1 and phase 2 trials. Some aspects of the role include ensuring the study guidelines are followed, communicating with the primary investigator, monitoring for adverse events, reporting adverse events and serious adverse events, and assuming regulatory responsibilities.Nurse practitioners play a crucial role in the care of CAR-T cell patients and their families including providing anticipatory guidance and monitoring for and managing adverse events in the inpatient and outpatient settings. In our nurse practitioner-run clinic, nurse practitioners build rapport and relationships with the patient and family, provide support in answering questions, and help coordinate care along with diagnosing and managing adverse events. The nurse practitioner provides continuity for the patient and family and with the referring provider following CAR-T cell infusion. Education is a major responsibility, and this includes educating patients and families, educating staff at the treating institution, and being a resource for other disciplines within the institution and for outside referring providers. Inpatient and outpatient nurse practitioners provide continuity of care between the inpatient and outpatient worlds. It is imperative to share pertinent patient information such as complications and required follow-up, to ensure a seamless transition between the inpatient and outpatient settings.As CAR-T cell therapy continues to advance, this opens many opportunities for future research. Nurses are in a unique position as we are involved throughout the entire CAR-T cell process. Since nurses are so closely involved in providing anticipatory guidance and in assessing and managing side effects related to this therapy, they are at the forefront of generating research related to CAR-T cell therapy. One area of possible research is using evidenced-based practice within nursing to guide the assessment and early identification of complications such as CRS and ICANS. Patient-reported outcomes are another area that can benefit from additional research and investigation. Since this is a newer therapy, information gained in this area can be incredibly helpful in providing anticipatory guidance and symptom management to these patients during their therapy. Since this therapy has only been available for the last 11 years and the number of patients who have received it is still small, there are limited data regarding survivorship and long-term effects. This is an area where nursing research will be extremely beneficial that will need further investigation. As this therapy advances, there will continue to be numerous research opportunities for nurses to contribute to this field. Therefore, it is crucial for nurses to be educated regarding this therapy so that advances can be made, nursing practice improved, and more optimal outcomes can be achieved for patients .Fig. 4NuCAR-T cell therapy has made great strides in pediatric oncology over the past 11 years. This transformative therapy began with treating patients with relapsed and refractory B-cell ALL. Although this is a promising and effective therapy for many patients, some still experience relapse. These relapsed patients require novel therapies, which led to more trials being developed for B cell ALL relapses after CAR-T cell therapy. This success has also led to trials for other relapsed and refractory pediatric hematologic malignancies and solid tumors such as neuroblastoma and brain tumors. Nurses, in various clinical and research roles, will always be at the forefront of these innovative therapies and are responsible for providing family-centered education, safe and expert patient care, anticipatory guidance, care coordination, maintaining research standards, and ongoing education of one another.During the preparation of this work the authors did not use AI in writing this manuscript.


Vascular endothelial (VE)-cadherin, an endothelium-specific adhesion protein, is found in the junctions between endothelial cells (ECs). It’s crucial to maintain the homogeneity of ECs. Keeping and controlling the contact between ECs is essential. In addition to its adhesive function, VE-cadherin plays important roles in vascular development, permeability, and tumour angiogenesis. Signal transfer, cytoskeletal reconstruction, and contractile integrating, which are crucial for constructing and maintaining monolayer integrity as well as for repair and regeneration, are the foundation of endothelial cell (EC) junctional dynamics. The molecular basis of adhesion junctions (AJs), which are closely related and work with actin filaments, is provided by the VE-cadherin-catenin complex. They can activate intracellular signals that drive ECs to react or communicate structural changes to junctions. An increasing number of molecules, including the vascular endothelial growth factor receptor 2 (VEGFR2) and vascular endothelial protein tyrosine phosphatase (VE-PTP), have been connected to VE-cadherin in addition to the conventional VE-cadherin-catenin complex. This review demonstrates significant progress in our understanding of the molecular mechanisms that affect VE-cadherin’s function in the regulation of EC behaviour during angiogenesis. The knowledge of the molecular processes that control VE-cadherin’s role in the regulation of EC behaviour during angiogenesis has recently advanced, as shown in this review.  Angiogenesis is a complicated process whereby already-existing blood vessels are exploited to create new ones . Angiogeα-catenin to the cytoskeleton of F-actin, finally forming a VE-cadherin-catenin complex. The mutual reactions between VE-cadherin and its binding partners are adjusted depending on the condition of the endothelium, resulting in a suitable cellular response. However, it is yet unclear what molecular processes underlie the spatiotemporal cycling of AJs between ECs. In this review, we go over the molecular processes that influence the behaviour of ECs induced by the VE-cadherin-catenin complex during angiogenesis.Endothelial cell-cell adhesion, which is a vital part of preserving vascular integrity, is made up of VE-cadherin, commonly referred to as CDH5 . ECs expDifferences in VE-cadherin activity reflect the diverse behaviours of ECs during vascular quiescence and angiogenesis . In prolAngiogenesis is a complex process that involves several steps. It is currently unclear how the local microenvironment selectively acts on ECs to cause capillaries to grow in an orderly manner through sprouting and intussusception. Vascular sprouting, however, also needs AJs disruption. Junctional strength is regulated by VE-cadherin endocytosis, and vascular stability is decreased by a reduction in surface VE-cadherin levels. When sprouting occurs, functional EC rearrangement is driven by VE-cadherin junction dynamics . RemodelThe differentiation of ECs into tip cells and stalk cells with unique morphologies and functional characteristics is necessary for sprout initiation. ECs stimulated with VEGF adopt a promising phenotype. Tip cells are the ECs that are at the head of the developing capillaries . Tip celVEGF is one of the initiators of angiogenic factors, which have a very strong stimulatory effect on the proliferation, migration, and chemotaxis of the vascular endothelium in various organs and even the whole body . The VEGin vitro and in vivo when VE-cadherin expression is somewhat decreased  Vav2 via Src . When VEecreased . Togetheecreased . The VE-Integrins primarily mediate cell-to-cell and cell-to-extracellular matrix adhesion. It acts as a receptor for matrix molecules, receives signals from extracellular molecules and transduces them through signaling pathways to influence gene expression, thereby regulating processes such as cell proliferation, cell cycle and cell migration . In angiVEGFR2 signaling is regulated by several receptors, including integrins. Signaling crosstalk between integrins and VEGFR2 is necessary for a fully realized angiogenic program . IntegriDuring sprouting angiogenesis, ECs actively compete for the location of tip cells, and (Neurogenic locus notch homolog protein) Notch signaling is associated with the dynamic rearrangement of ECs . Specialin vivo and in vitro showed that Notch is key for maintaining junctional heterogeneity  interacts to VE-cadherin via its proximal structural region that resembles fibronectin . TyrosinYes-associated protein 1 (YAP) and transcription regulator 1 (TAZ), effectors of the Hippo signaling pathway, control proliferation and apoptosis and are crucial for organ development . SeveralYAP is also a mechanotransduction protein that constantly shuttles between the nucleus and cytoplasm, with nuclear localization being a key determinant of its activation to regulate downstream target genes.YAP/TAZ delivers mechanotransduction cues from the extracellular matrix to control cell survival mediated through Rho and the cytoskeleton . ARHGAP1in vitro in a 3D vascular experiment (A member of the (CKLF-like MARVEL transmembrane domain) CMTM gene family is CMTM1-8. This gene family’s products have a significant impact on the immune system, the male reproductive system, and tumor formation . At baseperiment . Similarα-catenin and β-catenin proteins. Actomyosin’s cytoskeleton or external forces of mechanics are in charge of controlling this binding  JAIL was the name given to these structures . E-cadhein vitro and ex vivo environments, leading to the rupture of barrier integrity  ECM modification, all of which are essential for angiogenesis, according to studies . In vascntegrity . Angiogein vitro and in vivo sprouting angiogenesis, according to Gomez-Escudero et al.  has been identified to trigger VE-cadherin mobility and govern the dynamic dissociation and remodelling process of the VE-cadherin complex . To be mVasculogenic mimicry (VM) is the ability of cancer cells to arrange themselves into vascular-like structures for the goal of acquiring nourishment and oxygen without the aid of conventional blood vessels or angiogenesis . Over thThe precise regulation of gene expression is fundamental to normal cell development and function. The mechanisms of angiogenesis remain largely unknown. Vascular endothelial cells have been extensively studied for triggering a series of signaling pathways involved in endothelial differentiation and vascular stabilisation, but little is known about VE-cadherin and its associated epigenetic regulatory mechanisms. Gene regulation occurs at multiple levels, including DNA-binding transcription factors, chromatin and histone post-translational modifications . PolycomThe expression and aggregation of vascular endothelial cells upregulate the endothelial specific genes of claudin-5, VE-PTP and VWF, which play a key role in vascular stability, by inhibiting the activity of polycomb on the promoters of specific genes . The VE-VE-cadherin contributes to important and distinctive functional characteristics of ECs during angiogenesis. In particular, VE-cadherin interacts with β-catenin/α-catenin proteins to connect to the actin cytoskeleton . The cytAtherosclerosis, pulmonary hypertension, and cardiac fibrosis are vascular disorders that are impacted by the recombination of vascular structures. The precise regulatory mechanism of vascular remodelling during the formation of neointima and the generation of neointimal cells is yet unknown. Understanding the neointimal cell genesis and the molecular processes that control their development is essential for our comprehension of vascular disorders. The preservation of endothelial cell connections during  EndMT is facilitated by EC-CD45 expression in ECs, as demonstrated by research using an EC-specific lineage tracking system (VE-cadherin-BAC-Cre ERT2 mice) . AdditioTo allow maternal blood to access the placenta during the development of the mammalian placenta, trophoblasts penetrate the meconium of the mother and modify the spiral arteries . Endovasin vitro induces a significant increase in Ang2 mRNA and protein, which in turn stimulates the phosphorylation and degradation of VE-cadherin (Some retinal diseases, particularly diabetic retinopathy and age-related macular degeneration, are characterized by a breakdown in the structure and function of blood vessels, leading to abnormal vascular hyperplasia. Angiopoietin dysregulation is associated with diabetic retinopathy. Exposure of human retinal endothelial cells to hyperglycemia cadherin , therebySepsis is a dysregulated immune response to infection that results in multiple organ dysfunction and death . BreakdoTherefore, VE-cadherin-centered AJs are important for angiogenesis. A crucial component of the AJs connecting vascular ECs is VE-cadherin, whose structural and functional defects can cause AJs to break down. The regulatory mechanisms are unclear, although a range of pathogenic stimuli acting on ECs can result in aberrant activation and damage, instability of VE-cadherin and endocytosis , a reduc


Tissue engineering and regenerative medicine (TERM) holds great promise for addressing the growing need for innovative therapies to treat disease conditions. To achieve this, TERM relies on various strategies and techniques. The most prominent strategy is the development of a scaffold. Polyvinyl alcohol-chitosan (PVA-CS) scaffold emerged as a promising material in this field due to its biocompatibility, versatility, and ability to support cell growth and tissue regeneration. Preclinical studies showed that the PVA-CS scaffold can be fabricated and tailored to fit the specific needs of different tissues and organs. Additionally, PVA-CS can be combined with other materials and technologies to enhance its regenerative capabilities. Furthermore, PVA-CS represents a promising therapeutic solution for developing new and innovative TERM therapies. Therefore, in this review, we summarized the potential role and functions of PVA-CS in TERM applications. Tissue engineering (TE) is an interdisciplinary field that uses the principles of engineering and biological sciences to create biological substitutes that can reinstate, sustain, or improve tissues or organs functionality in the body . It provTissue engineering and regenerative medicine (TERM) were explored for over 30 years. Due to their similar goals, these two fields converged in recent years and became a potential strategy to satisfy the future requirements of patients . TissuesHydrogels are three-dimensional (3D) systems composed of hydrophilic polymeric materials, which exhibit a structure conducive to water absorption. This feature enables one to retain significant amounts of water. Their high-water content, softness, and porosity make them resemble living tissue and exhibit excellent biocompatibility ,16. TheyPolyvinyl alcohol (PVA) is a synthetic, biocompatible polymer extensively studied for developing scaffolds owing to its high strength, resistance to deformation under load, capacity for water retention, and porous architecture . At the 2) in its chemical structure, which can form hydrogen bonds with the hydroxyl groups of PVA  are emerging as a better option due to their unique properties and potential to promote tissue regeneration and repair . MSCs arIn 2015, Dashtdar et al. investigated whether MSCs seeded in PVA-CS hydrogel could result in comparable or even better cartilage healing than that of previously established alginate-transplanted model. This study confirmed that the PVA-CS-MSCs construct leads to comparable treatment outcomes in the rabbit cartilage defect model, thus suggested for for clinical applications in cartilage regeneration . In exteNour-Eldeen et al. established a scaffold that allowed adipose-derived mesenchymal stem cells (ADSCs) to proliferate and differentiate into chondrocyte-like cells using PVA-CS nanofiber scaffolds . CharactMoreover, the osteoconductive and tissue regeneration performance of the fabricated scaffold was demonstrated with and without AT-MSCs in vivo rat model . AbazariRecently, Wee et al. investigated the impact of transforming growth factor-beta 1 (TGF-β1) and -β3 on the chondrogenic differentiation of rBM-MSCs, grown on the PVA-CS-PEG (polyethylene glycol) scaffold in comparison to pellet cultures . The stuMohammadi et al. developed a novel 3D nanofiber hybrid scaffold of poly(ε-caprolactone), PVA, and CS for bone tissue engineering using MSCs via a multi-jet electrospinning method . The scaInterestingly, CS was shown to stimulate osteoblast differentiation and mineralization. Mathews et al. demonstrated the osteogenic potential of CS in a 2D culture system. This study presented novel findings indicating that CS enhanced mineralization by upregulating the genes involved in mineralization as well as calcium-binding proteins such as OPN, Integrin binding sialoprotein (IBSP), Collagen type I alpha 1 chain (COL1A1), ALP, and OCN . Chen etFor cardiovascular TE, PVA/CS or PVA-CS was used as a coating material for cardiovascular stents to improve their biocompatibility and reduce the risk of restenosis . In addiHydrogel delivery systems are used clinically and can provide therapeutically beneficial effects. The use of hydrogels allows for the precise control of the time and location of therapeutic agent delivery, including small-molecule drugs, macromolecular drugs, and cells .PVA-CS was used as a drug delivery system for various therapeutic agents in TE. This combination was shown to enhance drug solubility and stability, increase drug uptake by cells, and improve drug release kinetics. The hydrophilic nature of PVA and the cationic characteristic of CS provides an ideal environment for drug loading and delivery.In drug delivery, PVA-CS can be used in multiple forms, such as nanoparticles, microparticles, and hydrogels. The different forms have different advantages and can be tailored to meet specific drug delivery requirements. Mahato et al. developed PVA-CS lactate hydrogel and investigated it as a matrix for the continuous and gradual release of hydrophilic drugs . The devIn recent years, PVA-CS nanoparticles were used to encapsulate various drugs, including anticancer drugs, antibiotics, and anti-inflammatory agents. They were shown to increase drug bioavailability and potentially target specific cells or regions. In 2011, Parida et al. included Cloisite 30B in the formulation of PVA-CS as a matrix material component, and curcumin was prepared at various concentrations and loaded with PVA-CS/C 30B nanocomposites to investigate the in vitro drug delivery system for anticancer drugs . They stShagholani et al. has improved the interaction between PVA and CS hydrogel by magnetite nanoparticles, making them a favorable option for drug delivery and clinical applications . They syMicroparticles, microcapsules, and microspheres are common constituents of multiparticulate drug delivery systems. Microparticles are spherical particles ranging in size from 1 to 1000 µm and are used as multiparticulate drug delivery systems to improve efficacy, tolerability, and patient compliance . Micropa3. Insulin was incorporated into the formulated delivery system, resulting in a final solution with a concentration of 1 IU of insulin per 200 µL. The in vitro insulin release assay showed that glucose levels were maintained for 6 h, while in the in vivo experiment, the greatest reduction was observed 4 h after administration [In 2010, the hydrogel PVA-CS was developed to deliver insulin through the nasal cavity . The PVAstration . This sustration . The autstration . Thus, uPVA-CS was extensively studied for its potential use in wound healing, as its water retention capacity and antibacterial activity indicate that it is a perfect material for wound treatment ,45. CS wEscherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus subtilis) in wound sites [Escherichia coli and Staphylococcus aureus was evaluated by the inhibition method, and the wound healing properties were tested in mice skin. The PVA-CS -ZnO showed excellent antibacterial and wound-healing activity, suggesting its potential use for wound-healing applications [PVA-CS hydrogels and films are most used as wound dressings because they can retain water, which is critical for healing. This hydrogel absorbs exudate and creates a protective barrier that shields the wound from external contaminants. Several studies reported the antibacterial activity and healing properties of PVA-CS loaded with other active ingredients ,63,87,88nd sites . Furtherications .High cell proliferation capacity is critical for wound healing. Lin et al. reported that the combination of PVA, CS, and dextran exhibited high cell proliferation ability, making them ideal for wound dressing . RecentlDue to their high surface area and ability to mimic natural tissue structure, PVA-CS nanofibers can be used as wound dressings. Nanofibers can increase cell adhesion and migration, critical for wound healing. PVA-CS nanofibers can also be loaded with active ingredients to improve wound healing. Electrospun nanofibers are well suited as wound dressing materials because they have a high surface area ratio, variable pore size distribution, and oxygen permeability . MoreoveFatahian et al. developed a hybrid fiber mat through a co-electrospun hybrid of PVA, CS, and silk fiber mats. The hybrid fiber mat characteristics, including porosity, degradability, pore size, tensile strength, and hydrophilic properties for wound healing, were investigated in vitro and in vivo by localizing BMMSC keratinocytes on the mat . ComparePVA-CS composite is also commonly used in other applications, including periodontal treatment , ophthalSeveral studies showed that PVA-CS has properties that can be useful in periodontal treatment. Its biocompatibility, drug delivery, antibacterial, and wound healing properties showed promising treatment options for periodontal diseases. Dong et al. synthesized CS-decorated metronidazole (MTZ) microcapsules (CS@MTZ) and used them as cross-linkers for injectable PVA hydrogel preparation for periodontal drug delivery . The stuIn periodontitis treatment, local administration of drugs or antimicrobial agents is an appropriate strategy when an infection is intensely localized in the pockets and does not respond well to mechanical debridement and systemic antibiotic treatment. Constantin et al. synthesized a PVA-CS film containing silver nanoparticles and ibuprofen to treat periodontal disease . The filIn recent years, TE scaffolds emerged as a potential treatment strategy for the repair and regeneration of tissue defects in periodontal disease ,102. A rPVA-CS blend was evaluated for its potential for ophthalmic applications as an ocular drug delivery system. It is used to improve drug bioavailability and residence time in the eye ,103,104.Gene therapy is a procedure that involves delivering genetic material to cells to treat or prevent diseases. Viral and non-viral vectors are commonly utilized to transport therapeutic genes into specific cells for effective treatment . Non-virCS-based polymeric nanoparticles can be used as a non-viral vector for gene delivery. It has numerous advantages over viral vectors, such as safety and low immunogenicity . In cancPVA-CS was widely used in the cosmetics industry. A combination of PVA-CS can offer several benefits to cosmetic products, especially skin and hair care products ,117. SinMelaleuca alternifolia) essential oil and tested them on Wistar rats through subdermal implantations [Castor et al. synthesized four distinct composite films composed of PVA-CS and tea tree (ntations . This stntations .PVA-CS showed potential material for TERM , but theFabrication of the PVA-CS scaffold itself may be challenging due to difficulties in optimizing the pore size and porosity of the scaffold. Poor pore size and porosity of a scaffold can impair cell adhesion, proliferation, and differentiation. The shape of the scaffold can influence its porosity and pore size in several ways. For example, a flat, planar scaffold may have lower porosity than a 3D scaffold with more complex geometry, such as a cylindrical or spherical shape. This is because a planar scaffold has a lower surface area-to-volume ratio, which limits the number of pores created in the material.On the other hand, a more complex-shaped scaffold may provide a greater surface area-to-volume ratio, allowing a larger number of pores to form ,120. In PVA-CS nanoparticles are pH-dependent , which mThe limited availability of CS, which is derived from crustacean shells, can increase the cost of production. Additionally, PVA-CS must undergo rigorous testing and regulatory approval when combined with other biologically active components and polymer materials before it can be widely used in TERM applications.PVA-CS showed promising results in the laboratory, but producing large quantities of PVA-CS products for commercial use can be challenging. The process of synthesizing PVA-CS products can be complex and time-consuming, and scaling up the production process while maintaining desired quality and consistency can be difficult.The PVA-CS blend is a promising material for TERM applications due to its biocompatibility, antimicrobial properties, and ability to support cell growth and tissue regeneration. The best is that it can be fabricated and customized to fit the specific requirements of different cells, tissues, and organs, making it a versatile tool in RM. PVA-CS nanoparticles were investigated for their potential use as drug delivery vehicles, as they can protect the encapsulated drug from degradation and improve its bioavailability. Films prepared from PVA and CS were shown to have good mechanical strength, biocompatibility, and controlled drug release. On the other hand, hydrogels based on PVA-CS can be used for TE and drug delivery due to their large water absorption capacity and 3D network for cell growth. PVA-CS can also be used in the form of scaffolds for TE and wound healing applications as they provide mechanical support and promote cell growth.


A crisis is defined as a disruption in equilibrium at the failure of own personal resources which results in important distress and functional impairment. Particularly after Covid-19 pandemic we have been attending an increased number of this kind of consultations in the mental health services.To expose a new mental health program called PAIC (Ambulatory Intensive Community Program) started in March 2019 and addressed to people who are suffering a crises period, and to evaluate its effectiveness. The aim of this program is to reach prior stability, trying to avoid chronicity, clinical status perpetuation and sociofunctional impairment, as well as assessing suicidal risk and perform early prevention interventions.LTE and Clinical Global Impression (CGI-I). Changes in CORE-OM Scale were analysed using t-student test and a descriptive analysis was used for CGI-results.PAIC is proposed as an early, intensive, focused and psychotherapeutic intervention led by psychiatrist. Consist in an 8-week program of 30 to 60 minute weekly individual sessions. A total of a 205 patients were attended in PAIC during 2021 and 84,6% of the patients completed the program. Mixed-methods, clinical interviews and measurements using validated self-administrated questionnaires were used: CORE-OM pre/post treatment , List of Threatening Experiences, CORE-OM  showed improvement in all of four dimensions: subjective well-being (TW), problems/symptoms (PT), general function (TF) and risk (TR).Perception of clinical improving measured by CGI was 81,6%. There were no cases of clinical worsening.We conclude that intensive and early programs are effective reducing the intensity of symptoms and the level of disability in people who are experiencing a psychological crisis. Also, it supports primary health care as well as helps to avoid saturation of specialized system.None Declared


Advances in next-generation sequencing technology have identified many genes responsible for inborn errors of immunity (IEI). However, there is still room for improvement in the efficiency of genetic diagnosis. Recently, RNA sequencing and proteomics using peripheral blood mononuclear cells (PBMCs) have gained attention, but only some studies have integrated these analyses in IEI. Moreover, previous proteomic studies for PBMCs have achieved limited coverage (approximately 3000 proteins). More comprehensive data are needed to gain valuable insights into the molecular mechanisms underlying IEI. Here, we propose a state-of-the-art method for diagnosing IEI using PBMCs proteomics integrated with targeted RNA sequencing (T-RNA-seq), providing unique insights into the pathogenesis of IEI. This study analyzed 70 IEI patients whose genetic etiology had not been identified by genetic analysis. In-depth proteomics identified 6498 proteins, which covered 63% of 527 genes identified in T-RNA-seq, allowing us to examine the molecular cause of IEI and immune cell defects. This integrated analysis identified the disease-causing genes in four cases undiagnosed in previous genetic studies. Three of them could be diagnosed by T-RNA-seq, while the other could only be diagnosed by proteomics. Moreover, this integrated analysis showed high protein–mRNA correlations in B- and T-cell-specific genes, and their expression profiles identified patients with immune cell dysfunction. These results indicate that integrated analysis improves the efficiency of genetic diagnosis and provides a deep understanding of the immune cell dysfunction underlying the etiology of IEI. Our novel approach demonstrates the complementary role of proteogenomic analysis in the genetic diagnosis and characterization of IEI. Genetic diagnosis plays a central role in the clinical management of patients with inborn errors of immunity (IEI). However, the diagnostic yield for IEI based on the sequencing of germline DNA is still low and is estimated to be approximately 30%. This study shows the utility of integrated analysis with proteomics and targeted RNA sequencing (T-RNA-seq) of peripheral blood mononuclear cells. We identified the molecular cause and immune cell defects in patients with IEI, increasing the diagnostic yield by 6%. Notably, even in cases missed by T-RNA-seq, proteomics could identify the genetic etiology of the disease, suggesting the pivotal role of proteomic analysis in diagnosing IEI. Our novel approach improves the efficiency of the genetic diagnosis and elucidates the pathogenesis of IEI.Patients with inborn errors of immunity (IEI), previously known as primary immunodeficiency disorders, demonstrate increased susceptibility to infectious diseases, autoimmunity, autoinflammatory diseases, allergies, and malignancies . These cRNA sequencing (RNA-seq) has been well employed as one of the most valuable tools to study Mendelian disorders , 13, becMore recently, researchers have shown an increasing interest in proteomics due to technological advances in mass spectrometry (MS)-based protein identification , 28. To Here, we propose a state-of-the-art method for diagnosing IEI, providing notable insights into the pathogenesis of IEI. Our single-shot DIA-MS approach, which was high-throughput and cost-effective, enabled proteomic analysis of PBMCs at greater depth. Furthermore, this improved analytical depth achieved protein coverage nearly equivalent to the depth of transcriptome analysis by RNA-seq and allowed integrated analysis with T-RNA-seq. This study aims to highlight the complementary role of integrated analysis of proteomics and T-RNA-seq to canonical genomic analysis in determining the molecular pathogenesis of IEI.SI Appendix, Table SI Appendix, SI Appendix, Fig. The current study encompassed a cohort of 70 patients diagnosed with IEI but without a known genetic etiology. Of these, 48 patients underwent WES, and the remaining 22 underwent T-NGS of a 400 IEI gene panel. Prior genetic analysis was conducted based on criteria established by the American College of Medical Genetics and Genomics (ACMG), along with the patients’ phenotypes and the disease’s inheritance mode. However, no pathogenic variants were identified that satisfied these criteria.  based on the PCA for raw data  and mRNA   but a considerable reduction in protein expression   Fig. A and B, ly) Fig. .SI Appendix, Fig. XIAP region identified a large deletion containing a noncoding exon with promoter activity. These results were previously reported by Sbihi et al., and “patient 2” corresponded to this case  in a two-group comparison (B-cell-deficient clusters vs. others). The results showed that many genes involved in B-cell function were strongly downregulated in the B-cell-deficient group, even in the total protein profile, suggesting that the clustering results were valid . This finding is consistent with previous findings by Rivas et al., who demonstrated that variants generating premature stop codons and predicted to trigger NMD were prone to demonstrate ASE . NeverthAnother important finding was that target enrichment of RNA-seq allowed us to identify the genomic basis of an ADA2-deficient case. The expression levels of aberrant transcript was very low due to mRNA instability; Leafcutter results show that the number of aberrant splicing reads is only 0.008% of the cluster. However, target enrichment increased the read depth and revealed the aberrant splicing with intronic variant. These results reflect those of Gildea et al. who also found that target RNA-seq method increased the efficiency of identification of rare splice isoforms, which was difficult with standard RNA-seq . Given tAICDA. This rather contradictory result may be due to inadequate target enrichment of AICDA; in fact, some cases showed missing values. In contrast to B-cell analysis, T-cell analysis showed that the proteins involved in ribosome biogenesis and ribosomal RNA processing were downregulated to the same extent as those involved in T-cell function. However, paradoxically, these results coincide with those of well-regarded studies indicating that T-cell activation via T-cell receptor signaling enhances ribosome biosynthesis  were excluded due to the large number of samples containing MVs for that protein. Moreover, these proteins included 85 genes covered in the T-RNA-seq , which may have caused some bias in the results of correlation analysis. Additionally, analyzing only at a one-time point may underestimate the correlation as proteins and mRNAs have different temporal contexts , 48. In Seventy IEI patients were recruited from five institutions in three cohorts, with 34, 28, and 8 patients, respectively. In addition, six HCs participated in another period. Throughout this paper, we refer to the cohorts as Batch1 (B1), Batch2 (B2), Batch3 (B3), or Ctrl (C), and patients are identified by group and a unique ID, for example, B1_P1, B2_P35, or B3_P63. Clinical information, such as classification from IUIS, presumptive diagnosis, and candidate genes, was obtained from clinicians. The primary inclusion criterion for IEI patients was the lack of genetic diagnosis via a canonical diagnostic approach such as WES or T-NGS; that is, patients without pathogenic variants in genes consistent with their clinical features and mode of inheritance, and the interpretation of “pathogenic” was according to the ACMG criteria . TherefoThe local ethics boards approved this study of Hiroshima University, Tokyo Medical and Dental University, National Defense Medical College, Gifu University, and Kyoto University.Methods for sample preparation are described in “SI methods.”Methods for Mass spectrometry-based proteomics and T-RNA-seq are described in “SI methods.”To understand the etiology and pathogenesis of IEI, we carried out three different approaches using R v4.1 and Bioconductor v3.14 packages.z-scores. The absolute value of the z-score greater than two was defined as significant change. The absolute value of the z-score greater than or equal to 2 was defined as significant change. We also analyzed the quantitative differences between each case and the HCs to obtain further information about the biological significance. We calculated the log fold-change (LFC) and mean expression values using limma  on the comprehensive proteomic data to further validate the clustering results. DEA was compared in the cell-deficient cluster vs. others and was performed using DEP  enrichment analysis of significantly suppressed proteins using ClusterProfiler  proteins with higher specificity and (ii) proteins without MVs or with fewer MVs. In T-RNA-seq, on the other hand, the analysis was based on gene profiles selected based on the criteria described in (i), since T-RNA-seq data are already target-enriched and contain no MVs. We then normalized the data with the nefilter , and perHeatmaps . The k vsing DEP , which bsing DEP . In the Profiler . GO termpgad104_Supplementary_DataClick here for additional data file.


Stent thrombosis was the only secondary clinical outcome that showed a significant difference, favoring the IVUS-guided approach (p = 0.01). Procedural outcomes revealed that IVUS-guided procedures had longer stents, larger diameters, and longer procedure and fluoroscopy times . Stent number and contrast volume did not significantly differ between the approaches . In summary, routine IVUS use did not significantly improve clinical outcomes, except for reducing stent thrombosis. Decisions in CTO-PCI should be individualized based on patient characteristics and supported by a multi-parametric approach.Chronic total occlusions (CTO) in coronary angiographies present a significant challenge nowadays. Intravascular ultrasound (IVUS) is a valuable tool during CTO-PCI, aiding in planning and achieving procedural success. However, the impact of IVUS on clinical and procedural outcomes in CTO-PCI remains uncertain. This meta-analysis aimed to compare IVUS-guided and angiography-guided approaches in CTO-PCI. The study included five studies and 2320 patients with stable coronary artery disease (CAD) and CTO. The primary outcome of major adverse cardiac events (MACE) did not significantly differ between the groups ( Coronary artery disease (CAD) is the third major global cause of mortality ,2. DespiThe Cochrane Collaboration and PRISMA criteria were followed for conducting this meta-analysis ,17.https://pubmed.ncbi.nlm.nih.gov/) and Cochrane Library (https://www.cochranelibrary.com/) accessed until 22 March 2023). We used the following keywords: CTO or chronic total occlusions, (pci or PTCA), IVUS.Scientific literature was systematically searched for studies reporting clinical and procedural outcomes for different strategies of revascularization (IVUS-guided versus angiography-guided) in patients undergoing CTO-PCI. Articles were searched for on the following public databases: PubMed  independently reviewed search records. Divergences were settled by discussion and agreement. Studies were considered eligible if they met all of the following inclusion criteria: (a) any clinical study in which an IVUS-guided CTO-PCI was compared to an angiography-guided CTO-PCI approach; (b) the clinical scenario was CAD with evidence of a least one CTO; and (c) clinical and procedural outcomes were reported. Exclusion criteria were as follows: clinical scenarios different from stable CAD with evidence of CTOs; studies in which intravascular imaging performed was not IVUS; editorials or review articles; case reports or case series; meta-analyses; clinical and procedural outcomes not reported. Data extraction was carried out by the same co-authors .The primary clinical outcome was the incidence of major adverse cardiac events (MACE), defined as a composite of cardiovascular (CV) death, non-fatal myocardial infarction (MI), and target vessel revascularization (TVR) at the longest follow-up available. Secondary clinical outcomes were cardiovascular death, all-cause mortality, MI, TVR, target lesion revascularization (TLR) and stent thrombosis. Total stent length, stent diameter, total number of stents, procedure time, fluoroscopy time, and contrast volume were also analyzed as procedural outcomes.Two co-authors evaluated study quality . Divergences were handled by discussing and coming to a consensus. According to the ROBINS-II tool, the risk of numerous forms of bias was assessed: randomization process, deviation from the intended intervention, missing outcome data, outcomes’ measurement, and selection of the reported results .2). PROSPERO, international prospective register of systematic reviews, has the analytic procedure listed (PROSPERO record ID: CRD42023410342).Discrete data were reported as percentages, whereas continuous values were summarized as mean and standard deviation. The Mantel–Haenszel random effects model was used to compute summary effect sizes, and the findings of clinical outcomes were shown as risk ratios (RR) with 95% confidence intervals (95% CI), while differences regarding continuous procedural outcomes were expressed as standardized mean difference (SMD) with 95% CI. To determine the appropriate sample size to demonstrate or reject a 20% relative risk reduction with IVUS-guided CTO PCI with an alpha error of 5% and a statistical power of 80%, trial sequential analysis (TSA) was conducted. Meta-analysis was performed using OpenMetaAnalyst 10  and Comprehensive Meta-analysis Software . The TSA computer application  was used to carry out TSA. Study bias was evaluated by examining funnel plots graphically and using Egger’s and Begg’s tests. Heterogeneity of studies was assessed with Cochran’s Q test, and it was measured using the Inconsistency index (IFive studies (2320 patients with stable complex CAD and evidence of CTO) were included in this analysis out of the 871 studies that were screened  19,20,2,22,23. Tp = 0.40; From the 2320 patients included in the primary analysis, 143 patients (13.7%) presented with the primary outcome of MACE in the IVUS-guided group, versus 200 patients (15.6%) in the angiography-guided group . MI occurred in 56 patients (5.3%) in the IVUS-guided group, as compared to 99 patients (7.7%) in the angiography-guided group . All-cause mortality occurred in 21 patients (3.0%) in the IVUS-guided group, versus 31 patients (4.4%) in the angiography-guided arm . TVR was found in 46 patients (4.4%) of the IVUS-guided group, and in 60 patients (4.6%) of the angiography-guided group . TLR occurred in 46 patients (5.3%) in the IVUS-guided group, versus 56 patients (5.1%) in the angiography-guided group . Finally, stent thrombosis incidence was significantly lower in the IVUS-guided group as compared to the angiography-guided group . All the results of the secondary endpoints are showed in A CV death was registered in 12 patients (1.1%) in the IVUS-guided group and in 25 patients (1.9%) in the angiography-guided group  and stent diameter  were significantly higher in the IVUS-guided group, while no difference was apparent in the mean stent number . Since heterogeneity for this outcome was very high (I2 = 93%), we performed sensitivity analysis showing a large proportion of heterogeneity attributable to the study by Kalogeropulos et al. In fact, removal of this study yielded a change to moderate heterogeneity (I2 = 63%). Conditional analysis without this study showed a higher number of stents implanted in the IVUS-guided group  (p = 0.03) and fluoroscopy time . Lastly, no difference in mean contrast volume was found between the two approaches . All the results of the procedural outcomes analysis are depicted in Total stent length  . In addiA low to moderate risk of bias was found with the included studies . HeterogThe use of IVUS during CTO-PCI can solve a number of practical issues in a complex and lengthy procedure. Yet, the general clinical impact of IVUS use in this scenario is difficult to measure. In fact, in a consensus paper published by the European Association of Percutaneous Cardiovascular Interventions, it was emphasized that IVUS guidance compared to angiography could significantly improve clinical outcomes . HoweverThe ongoing and rapid advancement of therapeutic methods, interventional techniques, and materials frequently causes research results to become out of date once they are published, as is typical with interventional studies of percutaneous interventions, especially in CTO. Some of the studies included are from 2014 and 2015. Moreover, in the AIR-CTO study, only less then 50% of the patients were treated with a 2nd generation drug eluting stent (DES). Therefore, not all materials and interventional techniques used would reflect the actual state of the art, which may restrict the application of our findings to a typical nowadays scenario. Also, observational studies were included in this meta-analysis, leading to a possible risk of bias. Nevertheless, sensitivity analysis showed that the general outcomes’ outlook was not affected by the exclusion of non RCTs. Not all the studies included reported all data regarding both clinical and procedural outcomes, as well as some baseline characteristics. More in detail, the PROGRESS-CTO trial did not report data regarding all-cause mortality, stent thrombosis, and stent diameter, while the study from Kalogeropulos et al. did not report data regarding stent thrombosis and TLR. Finally, in the K-CTO trial, no data of procedure time, fluoroscopy time, and contrast amount were provided. The overall number of patients included is another drawback of this meta-analysis. In fact, there are limited studies comparing IVUS guidance with angiographic guidance in patients with CTO. Therefore, other randomized controlled trials ongoing (NCT04917432) will provide information about this important topic. The recent RENOVATE-COMPLEX-PCI trial shed new lights on the significant role of intravascular imaging in complex PCI to reduce MACE. In this trial, there were 319 patients whom complex CAD was characterized by CTOs. In the CTO subgroup analysis performed in this trial, the use of intravascular imaging significantly reduced the primary outcome in contrast to angiography guide . As this trial included both IVUS and optical coherence tomography as intravascular imaging device, we could not include those patients in our analysis. However, the results are in line with this meta-analysis and support our findings.A routinary use of IVUS to guide CTO-PCI was associated with a significant reduction in stent thrombosis rate. However, no impact on other clinical outcome measure was observed. IVUS guidance was associated with a significant increase in stent length and diameter, as well as procedure and fluoroscopy time. Results of ongoing randomized trials will hopefully provide significant and much awaited insight to this topic. Meanwhile, available data and expert consensus documents suggest that IVUS imaging is crucial in CTO-PCI to maximize obtain procedural success, as it provides relevant information to inform the selection of the most appropriate technique upfront and to guide the procedure in itinere. Therefore, despite that IVUS imaging is crucial in CTO-PCI to obtain procedural success, decisions should be tailored to the patient’s profile and supported by a multi-parametric approach.


Sports-related rotator cuff muscle injury is one of the most prevalent pathologies affecting overhead sports athletes. Since the COVID-19 pandemic and its subsequent stay-at-home protocols, physical therapy has transited into a new realm of telehealth. Current evidence regarding examining and managing RTC strain in telehealth physical therapy is minimal.A self-referred 14-year-old female Chinese semi-professional tennis player presented with an acute right RTC strain. The mechanism of injury involved forehand strokes with left trunk rotation. No ligamentous or labral damage was observed on Magnetic Resonance Imagining. The individualized care plan included virtual partner-assisted assessment, online instructions on therapeutic exercises, and education with psychosocial considerations.After a 6-week intervention, the patient demonstrated complete shoulder range of motion, full muscle strength, complete return-to-practice, 0% Quick DASH disability index, and 6/68 on the Tampa Scale for kinesiophobia.This case report demonstrated that telehealth is an accessible and cost-effective option for youth tennis athletes with RTC strain. This unique case showed a detailed roadmap from examination to discharge of this plan of care. There are also barriers including test and measure validity, and communication difficulties to be considered. Despite the challenges, this case was a good example of telehealth being an effective, repeatable, and cost-efficient option for patients with poor healthcare access. The American Physical Therapy Association defined TH as “the use of electronic information and telecommunication technologies to remotely provide healthcare information and services” –5. With Overall, the development of TH allows PT to break geographical restrictions. Despite its current progression, an effective and reliable method to measure the strength and special tests from TH requires further exploration. The purpose of this case report is to describe the successful management of an acute RTC injury in a youth semi-professional tennis player through telehealth. CARE guideline (for CAse REports) was followed when preparing this manuscript.1.2.The patient was a 14-year-old female Chinese semi-professional tennis athlete. She first developed mild right shoulder pain after a forehand tennis stroke with left trunk rotation in practice which disappeared after two days of resting. The second injury occurred five days later with the same mechanism of injury and resulted in 8/10 pain with immediate loss of upper extremity function. An x-ray and Magnetic Resonance Images (MRI) of the patient's shoulder were ordered by her primary care physician. The imagines ruled out epiphyseal plate, ligamentous, or labral damage, but showed a supraspinatus muscle strain. The patient self-referred to our team for TH due to the limited access to local sports-medicine resources. Oral consent was obtained from her guardian. During TH history taking, the patient complained of weakness with shoulder flexion, abduction, and external rotation (ER), as well as kinesiophobia. These combined physical and psychological limitations negatively impacted the patient's activities of daily living (ADL) and tennis. , and special tests. For range of motion (ROM) measurements, the clinicians screenshot the patient's shoulder active ROM (AROM), then measured the angles through a smartphone 2-Dimensional application (Angle Meter 360) after the assessment  Figure . For str1.3.Based on patient demographic, history, inspection, and ROM testing, the clinicians highly suspected a strain/minor tear of one or more RTC muscles, with less likely diagnoses of subacromial impingement, deltoid strain, and acromioclavicular joint sprain. Since the quality of tests for specific RTC muscles is lacking, the test cluster of Painful Arc, Drop-arm, and Infraspinatus test was used, which has a 91% probability of RTC tear if all three are positive . With tw1.4.Telehealth re-evaluation and treatments were delivered once per week, over 6 weeks. Each re-evaluation session included 10 min of subjective report related to overall exercises intensity, pain level, 10–15 min of re-assessment, and 15 min for patient education including home-exercise-program technique instructions, intensity tapering using rate of perceived exertion scale (RPE), activity modification and precautions, and psycho-social management. The patient's guardian assisted with the re-evaluation of ROM, and MMT in the same manner as the initial evaluation to ensure reliability. Between visits, the patient recorded herself performing the home exercises program (HEP) and sent videos to therapists weekly. The clinicians provided asynchronous education and feedback in the form of video and writing, which focused on form correction, RPE monitoring and tapering, and patient encouragement, within 12 h of receiving the recordings. The asynchronous nature was practically unavoidable due to time-zone differences, where the patient was located in China and the clinicians in different parts of the US.The plan of care was divided into three phases. The initial phase (0–2 weeks) aimed to reduce pain, improve shoulder ROM and increase strength of the scapular muscles. Therapeutic exercises included active-assistive-ROM  with low1.5.At 2 weeks follow-up, the patient demonstrated full pain-free shoulder ROM and self-reported 70% muscle strength on the modified partner-assisted MMT compared to the healthy side of flexion/abduction and 90% of ER. At week 4, the patient self-reported 100% strength in all shoulder muscles, with fully functional ADL including carrying, lifting, and pulling. Special test cluster showed all negatives when re-tested. After 5 weeks of rehabilitation, the patient started tennis-specific movements such as no-ball contact strokes and serves practice, then gradually returned to play with light-intensity ball contact. Before the patient fully returned to practice, her Quick Disabilities of the Arm, Shoulder, and Hand (DASH) resulted in 0% dysfunction with 0% disability in the Sports module, and 6/68 on Tampa Scale for kinesiophobia (lower is better for each). The patient and her guardian were satisfied with the outcome. One-month follow-up included an interview with the patient and her guardian regarding her current symptoms during tennis practice. The patient stated that her shoulder returned to prior level of function without any limitations. , education such as form correction, or intensity tapering became less efficient and may discourage patient participation . This neDespite the limitations presented in this case, it demonstrated that TH is an achievable, cost-efficient, and repeatable procedure for delivering physical therapy care. Future TH studies should focus on exploring the validity of the examining process and objective testing.3.In conclusion, this unique international telehealth case study demonstrated a successful episode of remote rehabilitation for a high-level teenage athlete with RTC muscle strain. This case is also a pioneering example of exploring the potential of TH in PT evaluations and treatments and improving patients’ accessibility to PT in underserved populations.4.The patient was contacted again one month following discharge. She reported that she had returned fully to tennis practice and continued with her independent HEP to maintain shoulder health.


No increased risk was observed in patients with resolved preformed DSAs . Patients with resolved preformed DSAs have similar graft prognoses as patients without DSAs, therefore, the persistence of preformed DSAs and development of de novo DSAs are associated with inferior long-term allograft outcomes.The post-transplant evolution of antihuman leukocyte antigen donor-specific antibodies (anti-HLA DSAs) includes three clinical patterns: resolved preformed DSAs, persistent preformed DSAs, and de novo DSAs. The aim of this retrospective study was to analyze the impact of resolved preformed, persistent preformed, and de novo anti-HLA-A, -B, and -DR DSAs in kidney transplant recipients on long-term renal allograft outcomes. This is a post hoc analysis of the study conducted in our transplant center. One hundred eight kidney transplant recipients were included in the study. Patients were followed for a minimum of 24 months after allograft biopsy, which was performed 3 to 24 months after kidney transplantation. The identification of persistent preformed DSAs at the time of biopsy was the most significant predictor of the combined endpoint of the study (>30% decline in estimated glomerular filtration rate or death-censored graft loss; HR = 5.96, 95% CI 2.041–17.431,  The presence of donor-specific antibodies (DSAs) directed against human leukocyte antigens (HLA) is a risk factor for antibody-mediated rejection (ABMR) and allograft loss in kidney transplant (KTx) recipients . Over thDSAs may occur before transplantation (preformed) or arise de novo after transplantation. Alloimmunization before KTx can be caused by pregnancy, blood transfusion, or prior transplant ,6. The dIn solid-organ transplantation, the monitoring of anti-HLA DSAs is currently under investigation. Recently, clinical recommendations for the post-transplant assessment of anti-HLA DSAs were published . NeverthThe aim of this retrospective study was to analyze the impact of resolved preformed, persistent preformed, and de novo anti-HLA-A, -B, and -DR DSAs in kidney transplant recipients with long-term renal allograft outcomes.This is a post hoc analysis of the study conducted in our transplant center . All 108Patients were divided post hoc into four groups according to their DSA status at the time of biopsy: no anti-HLA DSAs, resolved preformed anti-HLA DSAs, persistent preformed anti-HLA DSAs, and de novo anti-HLA DSAs.All transplantations were performed with negative complement-dependent cytotoxicity crossmatch. Donor–recipient pairs were ABO blood group compatible. All patients were of white ethnicity. None of the patients received a desensitization protocol before KTx. The decision of whether and what induction therapy (basiliximab or anti-thymocyte globulin (ATG)) to administer was made individually for each patient after the assessment of their immunological risk. Mostly, patients with a high immunological risk received ATG while patients with an intermediate immunological risk received basiliximab. In all patients, the maintenance immunosuppressive regimen consisted of the standard of care agents: a calcineurin inhibitor (tacrolimus or cyclosporine) with mycophenolate mofetil and prednisone. Only two patients were treated with cyclosporine due to tacrolimus intolerance. Biopsies were assessed using Banff classification criteria .At the time of biopsy, serum samples were obtained from all the patients and stored for further analysis. Sera were tested for the presence of anti-HLA DSAs and their characteristics including class, specificity, MFI, C1q-binding capacity, and IgG subclass according to the protocol described previously . ClinicaThe primary outcome of the study was defined as a permanent 30% decline (two consecutive laboratory assessments ≥ 3 months apart) in the estimated glomerular filtration rate (eGFR) compared to the baseline at the time of biopsy or death-censored graft loss . The Chronic Kidney Disease Epidemiology Collaboration 2009 (CKD-EPI) creatinine equation was used to determine the eGFR.Approval of the study was obtained from the Medical University of Warsaw medical ethics committee . The study was performed according to the principles expressed in the Declaration of Helsinki. All patients provided informed written consent.t-test, Mann–Whitney U test, and one-way analysis of variance , depending on the number and distribution of the compared variables. The Chi-square test was used to compare categorical variables. The Yates continuity correction was used when the frequency of events was low (<5). Logistic regression was performed to identify variables associated with DSA’s status. Event-free survival was assessed using the Kaplan–Meier method and log-rank test. Univariate and multivariate Cox proportional hazards models were used to quantify the hazard ratios for the study outcome. A backward stepwise elimination method was applied. A p-value < 0.05 was considered statistically significant.Statistical analysis was conducted using R software. The Shapiro–Wilk test was used to assess the normality of distribution. Continuous variables were presented as the means with standard deviation (SD) or medians with quartiles 1 and 3 (Q1–Q3) as appropriate. Categorical variables are presented as frequencies and percentages. Differences between continuous variables were assessed using the A total of 108 consecutive patients who received kidney transplants from brain-dead deceased donors and underwent renal allograft biopsy 3 to 24 months post-KTx were included in the study. Based on the DSA status at the time of biopsy, four groups were identified: no anti-HLA DSAs (N = 80), resolved preformed anti-HLA DSAs (N = 9), persistent preformed anti-HLA DSAs (N = 9), and de novo anti-HLA DSAs (N = 10). The demographic and clinical characteristics of the groups are provided in p < 0.0001). Two patients (22.2%) with persistent preformed DSAs experienced graft loss. As many as 42.9% of patients with persistent preformed DSAs had proteinuria at the end of follow-up, ≥50 mg/dL (p = 0.0001). No statistical difference between the groups was identified regarding the time of follow-up after biopsy, median proteinuria at the end of follow-up, time from biopsy to the primary outcome, and mortality.All patients were followed for a minimum of 24 months after the biopsy procedure. The clinical outcomes are shown in p < 0.0001 and p = 0.0090, respectively). However, there was no statistical difference regarding the combined endpoint survival between patients with resolved preformed anti-HLA DSAs after transplantation and patients without anti-HLA DSAs (p = 0.918).The event-free survival according to DSA status is shown in p = 0.0011), followed by the occurrence of de novo anti-HLA DSAs , proteinuria at biopsy ≥ 50 mg/dL , and increased donor’s age, with borderline statistical significance .The multivariate Cox regression model for the risk of the combined endpoint is depicted in Multivariate logistic regression analysis revealed several clinical, independent variables significant with DSA status . The useIn the study, the clinical impact of the evolution of anti-HLA DSAs in kidney transplant recipients was assessed. In recent years, transplant matching has evolved due to the development of new technologies. Moreover, the characteristics of donors and recipients have changed, which alters KTx outcomes . CurrentIn the cohort, 50% of patients with preformed anti-HLA DSAs cleared their antibodies after transplantation. None of the patients received desensitization treatment. This is directly in line with previous findings by Sanev et al. In their large study, which included 924 kidney transplant recipients, they also demonstrated that the persistence of DSAs is associated with higher MFI values and antibodies directed against HLA class II . HoweverOur results present that KTx recipients with resolved preformed anti-HLA DSAs have similar renal allograft outcomes regarding the eGFR and graft survival compared to patients without DSAs. Overall, these findings are in accordance with the results reported by other researchers ,24,25. TThe detection of persistent preformed anti-HLA DSAs at the time of biopsy and the identification of de novo anti-HLA DSAs are the main independent predictors of worse graft outcomes, defined as a 30% sustained decline of the GFR or graft failure. No significant difference between the characteristics of these antibodies was found. This might imply that current, circulating DSAs are more essential in the prediction of the renal allograft compared to resolved DSAs. A similar conclusion was reached in studies involving not only adult renal transplant recipients but also patients after other solid organ transplantation such as heart, lung, or intestine ,28,29,30Our findings highlight the clinical impact of the clearance of anti-HLA DSAs after renal transplantation. This is particularly important within the context of emerging new desensitization therapies ,34. HoweOne of the limitations of the present study is the post hoc, retrospective nature of the analysis. In addition, a relatively modest number of patients had been identified with DSAs in this single-center investigation. The characteristics of pre-transplant antibodies such as C1q-binding capacity and IgG subclasses were not assessed. Donor typing did not include HLA-C, -DP, and -DQ, therefore, antibodies against these antigens were not analyzed and the ABMR risk could not be assessed precisely. The cross-reactive epitope groups were not considered. Patients were tested for post-transplant DSAs only at the time of kidney allograft biopsy, which was performed 3 to 24 months after the transplant. Follow-up biopsies are not available. The use of different agents in the induction of immunosuppressive therapy was inevitable.To conclude, kidney transplant recipients with resolved preformed anti-HLA DSAs have similar graft prognoses as patients without DSAs. The persistence of preformed anti-HLA DSAs after kidney transplantation and the occurrence of de novo anti-HLA DSAs are independent predictors for inferior long-term allograft outcomes. Candidates for kidney retransplantation should undergo a cautious immunological risk assessment. These findings are valuable in light of the development of new desensitization protocols. Although transplant immunology evolves rapidly, there are many gaps in the current knowledge and further studies are needed to answer clinical questions.