Vancomycin is the most frequently used antibiotic, accounting for up to 35% of hospitalized patients with infection, because of its optimal bactericidal effectiveness and relatively low price. Vancomycin-associated AKI (VA-AKI) is a clinically relevant but not yet clearly understood entity in critically ill patients. The current review comprehensively summarizes the pathophysiological mechanisms of, biomarkers for, preventive strategies for, and some crucial issues with VA-AKI. The pathological manifestations of VA-AKI include acute tubular necrosis, acute tubulointerstitial nephritis (ATIN), and intratubular crystal obstruction. The proposed pathological mechanisms of VA-AKI include oxidative stress and allergic reactions induced by vancomycin and vancomycin-associated tubular casts. Concomitant administration with other nephrotoxic antibiotics, such as piperacillin–tazobactam, high vancomycin doses, and intermittent infusion strategies compared to the continuous infusion are associated with a higher risk of VA-AKI. Several biomarkers could be applied to predict and diagnose VA-AKI. To date, no promising therapy is available. Oral steroids could be considered for patients with ATIN, whereas hemodialysis might be applied to remove vancomycin from the patient. In the future, disclosing more promising biomarkers that could precisely identify populations susceptible to VA-AKI and detect VA-AKI occurrence early on, and developing pharmacological agents that could prevent or treat VA-AKI, are the keys to improve the prognoses of patients with severe infection who probably need vancomycin therapy. Acute kidney injury (AKI) is a common complicated disorder affecting about 2–7% of hospitalized patients and 13–78% of critically ill patients . An esseStreptococcus sp., Enterococcus sp., Actinomyces sp., Clostridium sp., and Eubacterium sp. x[IGFBP-7]/Cr between 144 and 192 hrs  was a predictor of non-recovery of VA-AKI [In a human study, Pang et al.  conducte< 0.001) . A prospf VA-AKI . In addif VA-AKI  found thf VA-AKI .As mentioned earlier, a high 5-HIAA/5-HT ratio in serum (AUC 0.88 (0.88–0.96)) was proposed as a potential surrogate biomarker for VA-AKI in humans . FurtherTo date, no promising therapy is available to treat VA-AKI. The only exception is that several case series suggest that the treatment of biopsy-proven ATIN with oral steroids for four weeks  may accelerate the rate of recovery . Since sSince the conventional treatment for VA-AKI is still limited, prevention is currently the most crucial strategy. The first step of the preventive strategy for VA-AKI is risk stratification according to the known risk factors listed in For those who have to receive vancomycin therapy, the preventive management includes: (1) adequate hydration before and during the vancomycin treatment course for the patients with intravascular volume depletion; (2) treatment and correction of the concurrent acute illness of the patients; (3) avoiding the concomitant use of other potentially nephrotoxic agents if other choices of drugs exist; (4) using the continuous infusion method rather than the intermittent infusion method; (5) avoiding treatment durations in excess of seven days if possible; (6) avoiding excessive vancomycin exposure by therapeutic drug monitoring and dosage adjustment (the updated guidelines recommend an AUC/MIC ratio of 400–600 (assuming a vancomycin MIC of 1 mg/L) as the target for patients with severe MRSA infections for considering both clinical efficacy and patient safety ); (7) coIn conclusion, VA-AKI is a clinically relevant but not yet clearly understood entity among critically ill patients. An increasing body of evidence regarding its pathophysiological mechanism, risk, and preventive strategies has been continuously published up to now. The pathological mechanisms of VA-AKI include oxidative stress induced by vancomycin, allergic reactions to vancomycin, and VTCs. All these mechanisms cause ATN, ATIN, and VTCs. In the future, disclosing more promising biomarkers that could precisely identify susceptible populations for VA-AKI and detect VA-AKI occurrence early, and developing pharmacological agents that could prevent or treat VA-AKI, are the keys to improve the prognoses of patients with severe MRSA infection who probably need vancomycin therapy.


Hundreds of tRNA genes and pseudogenes are encoded by the human genome. tRNAs are the second most abundant type of RNA in the cell. Advancement in deep-sequencing technologies have revealed the presence of abundant expression of functional tRNA-derived RNA fragments (tRFs). They are either generated from precursor (pre-)tRNA or mature tRNA. They have been found to play crucial regulatory roles during different pathological conditions. Herein, we briefly summarize the discovery and recent advances in deciphering the regulatory role played by tRFs in the pathophysiology of different human diseases.  Insights into the potential of RNA therapeutics have led to the extensive research progress in the arena of non-coding RNAs. This review provides insight about the tRNA-derived RNA fragments and summarizes their various functional implications in the pathophysiology of various human diseases. Coding transcripts are mainly mRNAs that are translated into proteins. Non-coding transcripts do not code for any protein, rather they play a significant role in the regulation of most protein-coding genes. Non-coding transcripts primarily comprise rRNAs, tRNAs, small nucleolar RNAs (snoRNAs), long non-coding RNAs (lncRNAs), and small non-coding RNAs (sncRNAs). With the advancements in next-generation sequencing technologies, extensive research has been performed to explore the roles of various sncRNAs , small interfering RNAs (siRNAs), and piwiRNAs, among others. sncRNAs derived from tRNAs, called tRNA-derived small RNAs (tsRNAs), comprise tRNA-derived RNA fragments (tRFs) and tRNA halves (tiRNAs) and are an emerging class that has recently come into consideration. These tsRNAs are 14–40 nt in length, which originates from both precursor tRNA as well as mature tRNA by the action of different ribonucleases (RNases), including DICER, ELAC2, angiogenin (ANG), and RNase P, among others. These are predominantly generated during various kind of stress conditions.,,,The human genome contains more than 500 tRNA-encoding genes, although a significant number among them are either not expressed or are expressed in a highly regulated cellular- or tissue-specific manner.E. coli, generated in response to bacteriophage infection.in vitro.,The first information of tRNA cleavage was noticed in ,,Pro 5ʹ half. They demonstrated that upon ribosome binding, the tRNAPro 5ʹ half inhibits protein biosynthesis with the concomitant appearance of a low-molecular-weight translation product. Functionally, tRNA halves (5′-tiRNAs and 3′-tiRNAs) and tRFs  seem to follow different mechanisms for their biological activity. Indeed, many investigations have indicated that only a fraction of cytosolic tRNAs is cleaved into tRNA halves during the stress response, and the levels of adaptor tRNA do not reduce critically enough to stop protein synthesis.,,,ANG-mediated tRNA cleavage commonly occurs at the anticodon loop of mature tRNA, leading to the accumulation of 5′- and 3′-tiRNAs in response to stress.,,,Recent studies by various groups have indicated diversified functional roles for tRFs as regulatory factors in human pathophysiological conditions, including virus infections .5,6,65,6Leishmania donovani and Leishmania braziliensis and reported that parasites secrete exosomes containing mRNA and sncRNA cargos. These RNAs were capable of modulating recipient cell phenotypes to support chronic infection. These exosomes deliver their ncRNA cargos to the macrophages, which support host-parasite interactions for seeking entry into the cell cytoplasm. RNA sequencing (RNA-seq) analysis of exosomal RNAs indicated that most of the sequences were derived from ncRNA species. Further in-depth analysis reveals the presence of tRFs with regulatory functions. Identified tRNA fragments were derived from the 5′ end or mid-5′ end (anticodon loop) and the 3′ end of specific tRNAs. Those identified tRFs and tiRNAs are tRNAAsp, tRNAGln, tRNAGlu, and tRNALeu, in which tsRNAAsp and tsRNAGln were found to be most abundant.Trypanosoma cruzi that show that extracellular vesicles released by T. cruzi contained tRNA-derived halves along with other ncRNAs that are crucial for promoting its replication cycles and to confer infection in the susceptible cells.Mycobacterium and tRNA or its derived fragments. Obregón-Henao et al.UGC that include the 3′ terminal CCA were shown to function as efficient adjuvants stimulating TLR3 and inducing the generation of T helper 1 (Th1) and cytotoxic T lymphocytes (CTLs) against a co-administered subunit vaccine antigen.GUG half in activating the TLR7 which is produced in response to infection by Mycobacterium tuberculosis.Various studies have shown that many pathogens secrete exosomes that contain different ncRNA-derived along with tRNA-derived fragments that are known to induce interleukins (ILs) and interferon (IFN) production.CTC. They further demonstrated that production of cytosolic 5′-tiRNA-GluCTC is independent of Dicer or Drosha and specifically produced in an ANG-dependent manner upon RSV infection. These 5′-tiRNAs enhance viral replication by inhibiting the translation of APOER2 by interacting with the 3′ end of the cognate mRNA.,CCC, tRF5-LysCTT, and tRF5-CysGCA) derived from the 5′ end of mature tRNAs. They found that, out of these three 5′-tRFs, tRF5-GlyCCC and tRF5-LysCTT were functionally competent in inducing the production of pro-inflammatory cytokines such as IL-6, IL-8, and RANTES.pro (tRF-3019) was shown to be capable of priming reverse transcription of human T cell leukemia virus-1 (HTLV1) genome and in fact be packaged into viral particles.TGA-derived 3′-tRF trailer interacts with La/SSB protein, which acts as a RNA chaperon to provide stability to the nascent transcripts produced by RNA Pol III. La/SSB also plays a significant role in HCV internal ribosome entry site (IRES)-mediated translation. The interaction between tRF and La/SSB leads to limited availability of cytoplasmic La/SSB, which inhibits HCV IRES-mediated translation.Different studies have also discussed the tRFs 5′- and 3′-tiRNA generation and their functional role during viral infections. Generation of tRFs and their related roles are very well studied in respiratory syncytial virus (RSV) infection. Wang et al.These studies suggest that tRFs play a significant role in host-pathogen interactions. They regulate host-pathogen interaction by various mechanisms such as gene targeting at the posttranscriptional level, intercellular and intracellular communications, and immune regulations during infectious diseases. Further characterization of tRFs function in infectious diseases will thus answer the possibility of therapeutic applications of these tRFs.,Cancer is a multifactorial disease characterized by uncontrolled proliferation of cells and their invasion into either surrounding tissues or metastasis to distant sites within the organism. Deregulation of cell proliferation mostly results from defects in signaling pathways and/or gene expression patterns.,,Asp, tRNAGlu, tRNATyr, and tRNAGly, which interact with YBX1 protein. YBX1 (Y-box binding protein 1) is known to bind oncogene transcripts in breast cancer and enhance their stability, which further leads to an increase of cell proliferation and promotes cancer. Contrarily, tRFs produced during cancer compete with the oncogene transcripts to bind with YBX1 and thus suppress cancer cell growth.CAG, a 3′-tsRNA, functionally interact with mRNA of ribosomal proteins RPS28 and RPS15. This interaction results in enhanced translation, which leads to the proliferation of hepatocellular carcinoma cells.,,Ser-derived ts-4521 leads to cell proliferation.tRFs are known to regulate protein translation by interacting with ribosomes and aminoacyl tRNA synthetases.TGA by ELAC2. Overexpression of this tRF was seen in different cancer cell lines with specific correlation with proliferation of prostate cancer cells.c and directly contribute to ANG-mediated angiogenesis and cancer cell proliferation.tRNA fragments in cancer can also be produced as a result of induction of specific endonucleases. 3′-tRFs  are produced by the cleavage of the cytoplasmic pre-tRNA-SerTAC-derived tRF-3017A promotes metastasis in gastric cancer by regulating the NELL2 tumor suppressor gene.,CUN, which was considered as a valuable molecular target in colorectal cancer.CAG is shown to promote cancerous cell survival in lung cancer.A direct correlation of tRNA fragments with metastasis and cell proliferation has been shown recently. Tong et al.CAC-2-1 with oral squamous cell carcinoma (OSCC), tiRNA-5034-GluTTC-2 with gastric carcinoma, and 5′-tRNA-GluCUC in prostate cancer.,CTC-003, tRF-GlyCCC-007, tRF-GlyCCC-008, tRF-LeuCAA-003, tRF-SerTGA-001, and tRF-SerTGA-002) derived from the 5′ end of five different tRNAs were shown to have significantly decreased expression in cancerous tissue when compared to control samples.,Since the prognosis of cancer improves dramatically with an earlier diagnosis, a lot of attention is being given to the discovery of early biomarkers.,Defects in tRNA metabolism or associated processing enzymes often leads to various neurological disorders. ANG mutations are reported in patients suffering from different neurological disorders, suggesting a direct involvement in pathways that lead to motor and dopaminergic neuron degeneration. The modulated RNase activity of mutant ANG has shown its implications in the pathogenesis of amyotrophic lateral sclerosis (ALS) and Parkinson’s disease (PD).A study by Blanco et al.CLP1 mutation was reported. CLP1 is a kinase protein that is primarily associated with pre-tRNA processing and generates mature tRNAs. Mutated CLP1 results in accumulation of unspliced pre-tRNAs and depleted mature tRNA levels of different isoacceptor tRNAs such as tyrosine, leucine, and isoleucine. Due to mutated CLP1 protein, spliced tRNA could not get ligated, which results in accumulation of tiRNAs, which leads to reduced protein translation and cell death.,,In pontocerebellar hypoplasia (PCH), GCC, tRNA-ValCAC, tRNA-GlnCTG, tRNA-GluTTC, and tRNA-LysTTT. Functional studies of these tRFs revealed that they inhibit pluripotent genes and lead cell differentiation in a lineage-specific manner.,Stem cells carry the potential for extensive division and differentiation, which helps in embryonic development and tissue regeneration. Particularly, mouse embryonic stem cells (mESCs) have the capacity for infinite self-renewal and differentiation into any cell type.,,,,,Accumulating evidence has suggested that biogenesis of tRFs occurs in response to various stress conditions.,5C) and N2-methylguanosine (m2G), were found to be significantly upregulated in HFD group sperm.in vitro fertilization (IVF) and tested the altered expression of metabolic genes. They found that IVF-derived offspring of male mice (fed on a low-protein diet) exhibited significant hepatic upregulation of the certain gene encoding the cholesterol biosynthesis enzyme squalene epoxidase in comparison to control IVF offspring. They suggest that tRF-GlyGCC is regulated by a low-protein diet and it targets a MERVL endogenous retroelement. tRF-GlyGCC-mediated repression of MERVL-regulated genes may affect placentation and finally culminate in altered development of the offspring.GCC has been shown to have the characteristic of dimerization. This property protects tRFs against the action of cellular nucleases.,,Increasing lines of evidence from animal studies and human famines suggest that parental environmental exposure can affect the germline and influence their progeny through epigenetic mechanisms.,AAA were enriched in both cellular and cell-derived vesicles (shuttle RNA), whereas an enriched concentration of tRFs derived from tRNA-AspGAY was present in immune cell cytoplasm.GTA was found to be upregulated, whose function is yet to be discovered.Recent studies have suggested the roles of tRNA fragments in eliciting an immune response. Dendritic cells (DCs), when interacting with T cells, release multi-vesicular bodies (MVBs) containing shuttle RNAs, including tRFs, which mediate communication between these cells to ensure the immune response.Rickettsia infection leads to ANG upregulation in mouse endothelial tissue. This upregulated ANG induces tRNA fragments such as tRNA-ValGTG and tRNA-GlyGCC, which are probable mediators of the immune response and autophagy.CGC induces IL-8 production by stimulating transcription factor p65.Gong et al.CTC induce chemokine and cytokine production.CTG, tRNA-HisGTG, and tRNA-GlnTTG, which induces an antibody response and thus can be categorized as potential biomarkers.Different viruses also generate tRFs, which mediate some type of immune response. During RSV infection, 5′ halves derived from tRNA-GluImmune cells have pools of 3′- and 5′-tRFs in their cytoplasm, while only 5′-tRFs are enriched in immune cell-derived vesicles.Aging is a constitutive change in all parts of the body, including the brain, which commonly shrinks in volume, and is followed by large size of sulci.,,GCA. Further analysis confirms that the overexpression of tRF-3003a leads to inhibition of JAK3 expression. Inhibition of JAK3 significantly reduces IL-6, which has been shown to be associated with OA pathogenesis and progression.CTC and tiRNA-5031-GluCTC-1 tiRNAs were downregulated in both high-grade OA human cartilage and old equine chondrocytes. The results showed that aged equine samples have changes in the expression of specific tRNAs and tRFs when compared to young equine samples and found the induction of tiRNA-GluCTC and tiRNA-HisGTG in old compared to young equine chondrocytes and in high-grade compared to low-grade cartilage. Along with these tiRNAs, the parent tRNA-CysGCA was also found to be increased in both aged equine chondrocytes and high-grade OA human cartilage samples.GTT, tRNA-IleAAT, and tRNA-AspGTC are downregulated. Intriguingly, increased concentrations of tRFs derived from naturally occurring suppressor tRNAs were noticed in the endogenous plasma of RA patients.Osteoarthritis (OA), rheumatoid arthritis (RA), and aging are independent processes; however, aging-related events lead to OA and RA. Studies have revealed that stress, a key hallmark of aging, promotes OA through cartilage degradation.tRNA-derived RNA fragments are the new entrant in the field of small ncRNAs. These fragments are generated from different regions of a tRNA molecule. Their generation depends on personal attributes, cell and tissue specificity, environmental (stress) conditions, and tRNA modifications. They have a functional role in various pathophysiologies.Earlier these fragments were considered to be junk byproducts. Their specificity in several biological processes suggests that these fragments can regulate various cellular functions, including fine modulation of gene expression, through either transcriptional, post-transcriptional, or both mechanisms, in response to internal and external stress.GCC as a result of ANG induction. These tiRNAs promote virus replication and thus can potentially be targeted to control RSV infection. tRNA-derived fragment mimics or anti-sense oligonucleotides (ASOs) against ideal tRF candidates can be used to explore the therapeutic application of tRFs.This review highlights some exciting aspects of fragmentation of tRNAs and their role in various pathophysiologies. Furthermore, we explore herein the possibilities to use tRNA fragments as biomarkers for the severity of disease or therapeutic potential. These tRNA fragments interact with TLRs and mediate the immune response. Wang et al.CAG is also known to promote cancerous cell survival in lung cancer. ASOs can thus potentially be targeted against these tRNA fragments. ASOs can work as inhibitors to inhibit the roles of the tRNA fragments that support cancerous cell growth and metastasis. tRFs involved in aging-related processes such as neurodegeneration, OA, and RA can be used as biomarkers. Along with these highlighted roles of tRFs in human diseases, they also serve as conduits for the transfer of information across progeny. During early embryogenesis and inheritance, 5′-tiRNA pools have been shown to be localized in the cauda region of sperm, which might get transferred to oocytes during fertilization.YBX1 is a protein that stabilizes oncogene transcripts and accelerates cell proliferation in breast cancer. Some tRNA fragments were found to interact with YBX1, which can suppress cell proliferation. Similarly, tRNA-Ser-derived ts-4521 also suppresses cell proliferation in lung cancer. These tRNA fragments can thus potentially be used as mimetic tRNA therapy against breast and lung cancer. In another case, a study on gastric cancer has shown that tRF-3017A promotes metastasis. tRF-LeuThe field of tRNA-derived RNA fragments is still at its early stage. There are many unanswered questions related to their biogenesis and functional roles. Advancements in deep-sequencing technology are needed to overcome various technical challenges. First, the tRNA structure is highly prone to modifications, which makes it difficult for sequencing and further characterization. Second, it is hard to discriminate among the reads generated by pre-tRNA or mature tRNA. Third, tsRNA generation particularly depends on multiple variables such as tissues specificity, disease conditions, cellular stresses, and presence of various modifications in tRNAs, etc. These factors complicate the identification of tRNA-derived RNA fragments generated from isodecoder tRNAs, which share sequence similarity but have different chromosomal locations. Technical advancement and more comprehensive studies are needed to discover the potential of tRNA-derived fragments as putative biomarkers or prognostic markers and also to discover the strategies for prospective therapeutics either as a stand-alone therapy or as an adjunct therapy.


Health care aides provide direct care for older residents with advanced dementia in long-term care facilities. This study aims to understand care aides’ perceptions of what is ‘good’ care, what is person-centered care, and how to provide person-centered care for older residents with advanced dementia, as preparatory work of the WE-THRIVE consortium’s efforts to develop internationally-relevant common data elements of person-centered dementia care and launch comparative research in LMICs. Semi-structured interviews were conducted with health care aides (N=35) from 2 government-owned and 2 private long-term care facilities in urban China. Directed and conventional content analysis were used, drawing upon core constructs of person-centered dementia care and Nolan’s (2006) senses framework. We found that although care aides were not trained in person-centered care, they did incorporate person-centeredness in their work by tailoring their care to the needs of older residents and facilitating interactions with residents and their peers through communication cues.


In vitro and in vivo experiments demonstrated that ICG-IBA-RGD exhibits excellent photothermal conversion capability and high tumor ablation efficiency.In this work, we present a novel photothermal agent ICG-IBA-RGD based on albumin-binding strategy for enhanced tumor targeting imaging and photothermal therapy.  In this work, we present a novel photothermal agent ICG-IBA-RGD based on albumin-binding strategy for enhanced tumor targeting imaging and photothermal therapy. Surgery treatment cannot remove all tumor cells, and sometimes even leads to the spread of tumor cells. Due to the lack of tumor specificity, both radiotherapy and chemotherapy will induce severe local or systemic side effects during tumor therapy.2 Besides, tumor tissue or cells will also become resistant to chemotherapeutic drugs and radiation during chemotherapy irradiation processes. Based on the limitations of the above therapeutic methods, photothermal therapy (PTT), which is a novel non-invasive cancer treatment strategy, has attracted extensive attention because of its high efficiency, easy operation, negligible side effects and good bioavailability.3,4 The core of PTT is photothermal agents (PTAs) that can convert near-infrared (NIR) light into cytotoxic heat for killing tumor cells.5 Importantly, PTT can ignore the effects of cellular resistance because it induces cell death via physical mechanisms such as protein denaturation and membrane rupture.6 Additionally, it can realize high precision since thermal effect is produced only when NIR light and PTAs are combined.Nowadays, cancer has been becoming a leading cause of human death with the rapid population growth and aging around the world.7,8 sulfide nanoparticles,9,10 and carbon-based materials11,12 exhibit favorable absorbance feature, excellent photothermal conversion efficiency and good photo-stability. However, obstacle of metabolism and potential long-term toxicity may restrict their future clinical translation.13 It has been shown that gold nanoparticles can retain in the mononuclear phagocyte system for over 6 months and induce adverse effects including DNA damage, oxidative stress and inflammation.14–16 Organic small molecule-based PTAs with good biocompatibility and biodegradability offer distinct advantages with regard to potential clinical application,17 which have attracted increasing attention as potential alternatives to nanomaterials in the field of PTT. Indocyanine green (ICG) is a NIR dye for FL and PA imaging approved by the U.S. Food and Drug Administration (FDA). Furthermore, ICG shows its potential as photothermal agent as its satisfactory NIR optical absorption and photothermal conversion capabilities. Therefore, ICG has been employed as a kind of theranostic platform in many studies.18–20Extensive efforts have been made to develop numerous types of inorganic and organic PTAs. Inorganic nanomaterials including gold nanoparticles,in vivo of ICG hamper its clinical use as photothermal agent.21 To overcome these limitations, ICG loaded nanoparticles which can improve the stability of ICG against light and heat and enhance specific tumor targeting ability should be developed. Human serum albumin (HSA) is the most abundant protein in plasma with multiple ligand binding sites, cellular receptor engagement, and a long circulatory half-life (19 days) because of interaction with the recycling neonatal Fc receptor.22–25 The utilization of these properties promotes albumin to be an attractive candidate for half-life extension and targeted intracellular delivery of drugs attached by covalent conjugation, genetic fusions, association or ligand-mediated association.26 As the long circulating protein in the blood, albumin has been used to extend blood half-life and reduce renal clearance of both imaging probes and therapeutic drugs.27 In addition to the prolonged half-life, albumin has also been found to specifically target tumor regions due to its enhanced permeability and retention (EPR) effect, abnormal nutritional needs, albumin receptor binding and SPARC-inducing effect,28–30 which is a unique advantage as the tumor-targeted drug delivery carrier.31,32 Dumelin et al.33 reported a class of albumin binders with stable non-covalent binding to HSA, of which 4-(p-iodophenyl)butyric acid (4-IBA) was one of the promising structures and aroused great attention in the field of molecular imaging and cancer radioligand therapies. The category of 4-IBA derivatives was first identified by Neri's group. These binders exhibited stable non-covalent binding with albumin and dissociation constants (Kd) in the low micromolar range. In addition, since the binding of 4-IBA to albumin is reversible, the albumin–drug complex can be used as a drug reservoir to enhance the drug biodistribution and bioavailability.26 It has been proved that 4-IBA-modified radio pharmaceuticals have extremely high affinity to HSA, and improve the biological half-life and tissue distribution in vivo.34–36 However, the application of 4-IBA as the albumin binder to improve the half-life and therapeutic efficacy of PTAs has rarely been reported.However, the instability in aqueous medium and nonspecific biodistribution vβ3-integrin overexpressed on tumor angiogenic endothelium; (3) 4-IBA for albumin binding. ICG-IBA-RGD exhibited good biocompatibility, excellent photothermal conversion capability upon single 808 nm laser irradiation. In vitro and in vivo experiments demonstrated that ICG-IBA-RGD could efficiently ablate tumor cells and completely eliminate tumor through photothermal effect when irradiated with an 808 nm laser. Notably, the tumor did not recur during the experiment. In summary, this work presents a biocompatible multifunctional albumin-binding photothermal agent ICG-IBA-RGD with great potential for image-guided photothermal therapy.In this work, we report a novel photothermal agent termed as ICG-IBA-RGD for efficacy NIR imaging and image-guided therapy . ICG-IBAThe synthetic route of ICG-IBA-RGD was shown in −2. As shown in −1) were exposed to 808 nm laser (1.0 W cm−2) irradiation for 10 min. As shown in −1 suspension of ICG-IBA-RGD + HSA remarkably raised from approximately 20.5 °C to 63 °C after 10 min of irradiation, which is high enough to kill tumor cells via hyperthermia. T) of ICG-IBA-RGD + HSA suspensions at various concentrations.In order to study the photothermal performance of ICG-IBA-RGD, different solutions, including ICG-IBA-RGD + HSA, ICG-RGD + HSA, ICG-IBA-RGD, ICG-RGD, ICG and deionized water, were irradiated for 10 min by an 808 nm NIR laser at a power density of 1.0 W cm−1  was observed on 4T1 cells even at the highest concentration of 400 μg mL−1 . The res−1, the 4T1 cells were obviously dead, and at 200 μg mL−1, there were no living cells in the field of view. The results suggest that the high photothermal conversion efficiency of ICG-IBA-RGD could effectively kill tumor cells in vitro.To further verify cell-killing effects of ICG-IBA-RGD in conjunction with NIR laser irradiation, 4T1 cells treated with different conditions were stained with a fluorescein diacetate  and propidium iodide  to visualize live and dead cells. As shown in in vivo, intravenous injections of ICG-IBA-RGD and ICG-RGD through the tail vein to the mice bearing 4T1 tumors were performed, and then the intensity of the fluorescence emitted by the live animal was monitored by the in vivo imaging system. As shown in ex vivo FL images of major organs and tumors harvested from mice at the time of 48 h. As expected, for the ICG-RGD group, few fluorescence was detected in the tumor, while higher fluorescence intensity was detected in the liver, which is the major metabolic organ of ICG-RGD. In the ICG-IBA-RGD group, a greater amount of fluorescence was detected in the tumor. Moreover, the weak fluorescence intensity of major organs, such as heart, liver, spleen, lung and kidney, further validated the tumor oriented accumulation of ICG-IBA-RGD. The fluorescence in liver was mainly concentrated in the gallbladder area. The above results indicated that ICG-IBA-RGD can effectively reveal its distribution and confirm tumor location to realize efficient PTT.To investigate the tumor-targeting ability of ICG-IBA-RGDin vivo are then performed on 4T1 tumor model. As mentioned before, the mice were divided randomly into 4 groups: (1) PBS only (2) laser only (3) ICG-IBA-RGD (1 mg mL−1) (4) ICG-IBA-RGD (1 mg mL−1) + laser. Throughout a 14 day treatment period, the size of the tumors was measured every 2 days and the growth curve of tumors was drawn. As shown in Encouraged by above results, the PTT experiments To further evaluate the efficacy of ICG-IBA-RGD on tumor PTT, the histopathological sections of tumor of the mice in different groups were analyzed 14 days after treatment. Histological images of H&E stained tumor sections showed that there was no obvious malignant necrosis in the groups of mice treated with the PBS, laser and ICG-IBA-RGD only, while apparent nucleus dissociation and necrosis were observed in mice treated with ICG-IBA-RGD plus laser. In addition, the TUNEL staining results showed that the apoptotic cells of tumor tissues had significantly increased in ICG-IBA-RGD treated mice with 808 nm laser irradiation, but not in the other three groups. PCNA staining results indicated that the proliferation capacity of tumor tissue was obviously reduced in ICG-IBA-RGD treated mice after laser irradiation as compared to those in the other three groups . These rIn summary, we successfully designed and constructed a superior albumin-binding photothermal agent ICG-IBA-RGD for targeted tumor imaging and photothermal therapy. ICG-IBA-RGD shows enhanced photothermal conversion efficiency and low toxicity both at the cellular and animal levels. The increased and long-lasting accumulation in tumor of ICG-IBA-RGD was observed in the imaging experiment, which will benefit for targeted photothermal therapy. Actually, the phototherapy study indeed demonstrated that ICG-IBA-RGD could efficiently eliminate tumor without recurrence within 14 d. Collectively, we report here a superior albumin-binding photothermal agent ICG-IBA-RGD and bring a new insight into the development of excellent phototheranostics for practical cancer theranostics.All animal procedures were performed in accordance with the Guidelines for Care and Use of Laboratory Animals of Central South University and approved by the Animal Ethics Committee, The Second Xiangya Hospital, Central South University, China.There are no conflicts to declare.RA-011-D0RA09653A-s001


Clinical presentations of COVID-19 are highly variable, yet the precise mechanisms that govern the pathophysiology of different disease courses remain poorly defined. Across the spectrum of disease severity, COVID-19 impairs both innate and adaptive host immune responses by activating innate immune cell recruitment, while resulting in low lymphocyte counts. Recently, several reports have shown that patients with severe COVID-19 exhibit a dysregulated myeloid cell compartment, with increased myeloid-derived suppressor cells (MDSCs) correlating with disease severity. MDSCs, in turn, promote virus survival by suppressing T-cell responses and driving a highly pro-inflammatory state through the secretion of various mediators of immune activation. Here, we summarize the evidence on MDSCs and myeloid cell dysregulation in COVID-19 infection and discuss the potential of MDSCs as biomarkers and therapeutic targets in COVID-19 pneumonia and associated disease. MDSCs aWhile MDSCs were discovered in malignant diseases and the majority of studies assessed MDSCs in cancer conditions, emerging evidence shows that MDSCs are more diverse and are involved in inflammation, autoimmunity and infection . The facvs. harmful role of MDSCs is rather complex and depends on the animal species/model system used, the stage of disease, and the ratio/balance between pathogens, T cells and MDSC  have evolved several mechanisms to inhibit IFN-I induction and signaling, e.g. suppression of MAV signaling , 34. PatThere is increasing evidence to suggest that distinct innate immune responses, specifically, underlie the different clinical trajectories of COVID-19 patients and that the hyperinflammatory syndrome in severe COVID-19 results from a dysregulated host innate immune response . TranscrhiCD11chi inflammatory monocytes with an interferon-stimulated gene signature, indicative of terminally differentiated monocytes, whereas severe COVID-19 was characterized by a lack of type I IFNs, high levels of HLA-DRlow classical monocytes and CD10lowCD101-CXCR4+/- neutrophils with an immunosuppressive profile in blood and lungs of severe cases, suggestive of emergency myelopoiesis  adaptive immune responses. MDSC activity can be enhanced by malignant or infectious triggers as described for a variety of viral, bacterial and fungal infections. Several recent reports have shown that patients with severe COVID-19 exhibit a dysregulated myeloid cell compartment, with increased MDSC levels and activity correlating with disease severity. In mild COVID-19 disease course, studies have reported an increase in HLA-DRopoiesis , 47–50.+CD16+ neutrophils, decreased granulocytic expression of integrin CD11b, and Th2-related CRTH2 downregulation in eosinophils and basophils established a COVID-19 signature. Severity was also associated with emergence of PD-L1 checkpoint expression in basophils and eosinophils  dependent mechanism. Furthermore, these patients showed increased Arg-1 and IL-6 plasma levels. COVID-19 patients had fewer T cells, and displayed downregulated expression of the CD3zeta chain  and VDAC, were present in the PBMC of the acute COVID-19 patients and correlated with severity of disease . Furtherta chain . In relaentation .Single-cell RNA sequencing (scRNAseq) data from bronchoalveolar lavage (BAL) also revealed the presence of neutrophils and macrophages as a hallmark of severe COVID-19 . Among tScRNAseq profiling was further used to characterize the PBMC compartment of uninfected controls and COVID-19 patients and cells in paired broncho-alveolar lavage fluid (BALF) . A closeImmature myeloid subsets and bronchoalveolar cells of critically-ill COVID-19 patients have been found to express HIF1alpha, a critical regulator of the differentiation and function of MDSCs, and transcriptional targets related to inflammation ; virus sensing, (TLRs); and metabolism  . The up-Calprotectin (S100A8/9) plasma level and a routine flow cytometry assay detecting decreased frequencies of non-classical monocytes have been shown to discriminate patients who develop a severe form of COVID-19, suggesting a prognostic value that deserves prospective evaluation . ElevateIn summary, various observational retrospective investigations suggest that MDSCs, their subsets or MDSC-related markers and signatures Figure 1via the DP2 receptor signaling in group 2 innate lymphoid cells (ILC2). Targeting PGD2/DP2 signaling using a receptor antagonist such as ramatroban could be used in immunotherapy for immune dysfunction and lymphopenia in COVID-19 disease  disease .Alarmin S100A8 was robustly induced in SARS-CoV-2-infected animal models as well as in COVID-19 patients. Paquinimod, a specific inhibitor of S100A8/A9, could rescue the pneumonia with substantial reduction of viral loads in SARS-CoV-2-infected mice . A groupTherapeutic strategies targeting the migration/recruitment of myeloid cells from bone marrow as mentioned above could be considered for the treatment of COVID-19 induced hyper-inflammation and immune dysregulation . InhibitGiven the expression of inhibitory receptor upregulation observed in a variety of cell subsets during the progression of COVID-19 , targetiRegarding potential targeting of MDSCs in COVID-19, it is essential to define the disease stage and disease severity level where such a therapeutic approach might have the greatest potential and would be beneficial rather than harmful to disease outcome. This consideration is key, as, in analogy to other viral infections –26. MDSCCOVID-19 activates the innate immune system and suppresses adaptive T cell responses. MDSCs are key cellular players connecting innate and adaptive immunity. Both M-MDSCs and PMN-MDSCs accumulate in patients with COVID-19 and reflect disease outcome, but what does this mean for the future of COVID-19 diagnosis, monitoring and treatment? Currently, inflammation, cell-death- and coagulation-associated serum proteins such as CrP, LDH and IL-6 as well as D-Dimers are used to characterize COVID-19 severity and disease progression clinically; it remains to be assessed how MDSC frequencies in peripheral blood and/or airway fluids relate to these clinical serum markers and whether combined/composite biomarker scores composed of both serum proteins and cells (PMN-MDSCs and/or M-MDSCs) could be superior than clinical serum markers alone to monitor and predict the outcome and treatment response in COVID-19. Targeting MDSCs as future therapeutic approach in COVID-19 is farer away, yet could add substantial value, particularly in combination with other immunomodulatory drugs, such as cytokine blockers.MR contributed to manuscript research and writing. FS contributed to manuscript research and writing. DH contributed to manuscript research and writing. All authors contributed to the article and approved the submitted version.Authors MR, FS and DH were employed by the company Novartis.


High-dose chemotherapy followed by autologous hematopoietic stem cell transplantation (Auto-HSCT) is a standard frontline treatment for fit mantle cell lymphoma (MCL) patients. As there is a need for predictive factors to identify patients unlikely to benefit from this therapy, we investigated the prognostic impact of lymphoma cell contamination of autologous stem cell grafts. Analyzing a cohort of 36 MCL patients, we show that lymphoma cell contamination of stem cell grafts is associated with poor outcomes after Auto-HSCT. Its analysis might thus improve risk assessment and enable risk-stratified treatment strategies for MCL patients.Novel predictive factors are needed to identify mantle cell lymphoma (MCL) patients at increased risk for relapse after high-dose chemotherapy and autologous hematopoietic stem cell transplantation (HDCT/Auto-HSCT). Although bone marrow and peripheral blood involvement is commonly observed in MCL and lymphoma cell contamination of autologous stem cell grafts might facilitate relapse after Auto-HSCT, prevalence and prognostic significance of residual MCL cells in autologous grafts are unknown. We therefore performed a multiparameter flow cytometry (MFC)-based measurable residual disease (MRD) assessment in autologous stem cell grafts and analyzed its association with clinical outcome in an unselected retrospective cohort of 36 MCL patients. MRD was detectable in four (11%) autologous grafts, with MRD levels ranging from 0.002% to 0.2%. Positive graft-MRD was associated with a significantly shorter progression-free and overall survival when compared to graft-MRD negative patients  and predicted early relapse after Auto-HSCT (median time to relapse 9 vs. 44 months). As a predictor of outcome after HDCT/Auto-HSCT, MFC-based assessment of graft-MRD might improve risk stratification and support clinical decision making for risk-oriented treatment strategies in MCL. High-dose chemotherapy (HDCT) followed by autologous hematopoietic stem cell transplantation (Auto-HSCT) is standard of care in the front-line management of transplant-eligible mantle cell lymphoma (MCL) patients achieving remission to induction chemoimmunotherapy ,2. EarlyMore recent studies in MCL on the other hand demonstrated that polymerase chain reaction (PCR)-based MRD assessment in PB and BM samples after induction chemoimmunotherapy predicts outcome after Auto-HSCT ,15,16,17As multiple studies have demonstrated that response to induction chemoimmunotherapy is of prognostic significance in MCL patients ,43,44,45n = 32) or at relapse (n = 4). Response assessment was performed by CT or positron emission tomography (PET)-CT at the end of induction or salvage therapy prior to HDCT/Auto-HSCT based on respective current International Working Group (IWG) criteria , MIPI risk group and remission status at Auto-HSCT the hazard ratios of graft-MRD for PFS and OS were 2.5 [95% CI 0.4–14.8] and 2.1 [95% CI 0.3–14.2], respectively  and OS (HR 4.0 [95% CI 1.1–14.9]) after Auto-HSCT . Graft-Mectively .Identifying MCL patients at increased risk for relapse after HDCT/Auto-HSCT is of growing importance as novel therapeutic options, including bruton tyrosine kinase (BTK) inhibitors, immunomodulatory agents and chimeric antigen receptor (CAR) T cell therapies have improved the prognosis of MCL patients relapsing after HDCT/Auto-HSCT and might even challenge the status of HDCT as standard frontline treatment in the future ,35,36,41−4 [0.01%]) here showed true-positive and true-negative rates of 80% and 92% with respect to PCR, respectively. In the study at hand, acquisition of high numbers of events/lymphocytes [−5 (0.001%). One downside of the reliance on a lack of polyclonal B cell background, however, is the lacking suitability of our MFC panel for long-term disease monitoring  after Auto-HSCT. On the other hand, the MFC panel used in our study is based on cell surface markers generally assessed at diagnosis and is thus more widely applicable than MFC panels relying on the detection of markers not necessary assessed in clinical routine flow cytometry to discriminate MCL cells from polyclonal B cell background  CT ,43 are wphocytes  and the nd CD11a ). Additind CD11a ,64,65.Overall, PFS and OS after Auto-HSCT in our MCL patient cohort were comparable with previous reports of MCL patients undergoing intensive frontline therapy ,8,17,45.Of note, the prevalence of graft-MRD was lower than reported in PCR-based MRD studies using PB and/or BM samples ,15,46. IWhile also detectable in one patient in CR, graft-MRD was primarily observed in patients not achieving a complete response to induction chemoimmunotherapy. Of note, graft-MRD was associated with a shorter PFS in this subgroup and predicted early relapse after Auto-HSCT. Based on these observations, assessment of graft-MRD might be of particular relevance in patients not achieving a CR prior to Auto-HSCT.It remains to be elucidated whether autografted lymphoma cells actually contribute to an unfavorable disease course or graft-MRD rather reflects aggressive disease biology and residual in vivo disease as an indicator of a particularly poor response to induction therapy. As such and due to its association with early relapse, detection of graft-MRD raises the question whether HDCT/Auto-HSCT remains the adequate treatment strategy in these poor-responding patients. Both BTK inhibitor-based and CAR T cell therapy, despite a lack of data in this setting, here represents promising and legitimate alternatives ,36,41. WWe here show for the first time that MRD assessment in autologous stem cell grafts by MFC, a non-invasive, inexpensive and well-established method, predicts outcome after high-dose chemotherapy. As a predictor of early relapse and poor outcome after HDCT/Auto-HSCT, graft-MRD assessment might contribute to pretransplant risk assessment and supports clinical decision making for risk-oriented treatment strategies in MCL patients.


Missing scores complicate analysis of the Patient-Reported Outcomes version of the Common Terminology Criteria for Adverse Events (PRO-CTCAE) because patients with and without missing scores may systematically differ. We focus on optimal analysis methods for incomplete PRO-CTCAE items, with application to two randomized, double-blind, placebo-controlled, phase III trials.N = 64 and 107, respectively). For each trial, we conducted between-arm comparisons on the PRO-CTCAE via complete-case two-sample t-tests, mixed modeling with contrast, and multiple imputation followed by two-sample t-tests. Because interest lies in whether CTCAE grades can inform missing PRO-CTCAE scores, we performed multiple imputation with and without CTCAE grades as auxiliary variables to assess the added benefit of including them in the imputation model relative to only including PRO-CTCAE scores across all cycles.In Alliance A091105 and COMET-2, patients completed PRO-CTCAE items before randomization and several times post-randomization ; NCT01522443 (COMET-2). Safety and treatment tolerability have historically been assessed solely by clinicians via the Common Terminology Criteria for Adverse Events (CTCAE). However, clinician reports under-detect symptoms relative to patient reports. Therefore, the National Cancer Institute contracted development of the Patient-Reported Outcomes version of the CTCAE (PRO-CTCAE) item library, which is now widely implemented in cancer clinical trials. However, missing PRO-CTCAE responses from patients complicate reporting and analysis. As with other patient-completed questionnaires, patients with and without missing PRO-CTCAE responses may systematically differ, thus jeopardizing the validity and generalizability of clinical trial results. For example, patients with more severe symptoms might miss PRO-CTCAE assessments more often than patients with less severe symptoms, potentially leading to invalid conclusions being drawn from analyses based on only the remaining patients’ PRO-CTCAE responses. This paper focuses on analysis of the PRO-CTCAE when some patients’ responses are missing (Alliance A151912), with application to 2 randomized, double-blind, placebo-controlled, phase III trials: Alliance A091105 and COMET-2. In each trial, we applied various methods for comparing patient-reported symptoms across treatment arms while addressing missing PRO-CTCAE responses. We found that optimal methods use patients’ responses to the PRO-CTCAE at other time points to provide information about their would-be responses to the PRO-CTCAE at the time point of interest. Clinicians’ CTCAE grades did not provide useful information about patients’ missing PRO-CTCAE responses.http://healthcaredelivery.cancer.gov/pro-ctcae). Patients complete a subset of items based on symptomatic adverse events most relevant to the treatment(s) under investigation. The National Cancer Institute recommends reporting patients’ PRO-CTCAE scores in conjunction with clinicians’ CTCAE grades to improve the evaluation of symptomatic adverse events in cancer clinical trials [To incorporate the patient perspective into assessments of symptomatic adverse events, the National Cancer Institute contracted development of the Patient-Reported Outcomes version of the Common Terminology Criteria for Adverse Events (PRO-CTCAE) . The PROl trials . Relativl trials , 4. FurtHowever, missing scores complicate reporting and analysis of the PRO-CTCAE. Patients with and without missing scores may systematically differ, thus jeopardizing the validity and generalizability of clinical trial results. For example, patients with more severe symptomatic adverse events might drop out or feel too unwell to complete PRO-CTCAE assessments, such that these patients miss PRO-CTCAE assessments more often than patients with less severe symptomatic adverse events. Analyses based on only the remaining patients’ PRO-CTCAE scores may result in biased parameter estimates and invalid conclusions.t-test, mixed modeling with contrast, and multiple imputation followed by a two-sample t-test. Because interest lies in whether CTCAE grades can inform missing PRO-CTCAE scores, we then perform multiple imputation with and without CTCAE grades as auxiliary variables to assess the added benefit of including CTCAE grades in the imputation model relative to only including PRO-CTCAE scores across all cycles.This paper focuses on optimal analysis methods for incomplete PRO-CTCAE items (Alliance A151912), with application to 2 randomized, double-blind, placebo-controlled, phase III trials: Alliance A091105 and COMET-2. We conduct between-arm comparisons within each trial while comparing the following strategies for addressing missing PRO-CTCAE scores: complete-case two-sample In A091105, patients with desmoid tumors or deep fibromatosis were randomized 2:1 to receive sorafenib or placebo by mouth once daily. Upon confirmation of progression, patients assigned to the placebo arm were allowed to cross over to open-label use of sorafenib. The primary endpoint was progression-free survival . The primary endpoint was pain response at week 6 confirmed at week 12  Consortium , the PROt-test, mixed modeling with contrast, and multiple imputation followed by a two-sample t-test. The complete-case two-sample t-test assumes a missing completely at random mechanism, meaning the probability of missingness is unrelated to the observed and missing scores. Excluding patients with missing scores yields unbiased parameter estimates under a missing completely at random mechanism because the observed scores can be regarded as a random subsample of the hypothetical complete scores. Mixed modeling and multiple imputation assume a missing at random mechanism, meaning the probability of missingness is unrelated to the missing scores after conditioning on the observed scores. The missing at random mechanism is a much more plausible assumption than the missing completely at random mechanism. Mixed modeling uses maximum likelihood estimation and includes patients’ responses to a PRO-CTCAE item at all available cycles, such that patients with any observed scores contribute to the mixed model. In doing so, mixed modeling accounts for missing PRO-CTCAE scores using patients’ responses to the same PRO-CTCAE item at different cycles (but not patients’ responses to other PRO-CTCAE items or clinicians’ CTCAE grades). Multiple imputation involves creating multiple copies of the dataset with different imputed scores (imputation phase), analyzing the imputed datasets as though they were complete datasets , and pooling the parameter estimates and standard errors across the imputed datasets to yield a single set of results (pooling phase). Multiple imputation quantifies uncertainty due to the missing scores and inflates the standard errors accordingly based on changes in the imputed scores across the imputed datasets.Between-arm comparisons on the PRO-CTCAE at week 12Multiple imputation was performed with and without CTCAE grades as auxiliary variables to assess the added benefit of including CTCAE grades in the imputation model relative to only including PRO-CTCAE scores across all cycles. To impute the missing scores, multiple imputation uses available information about patients’ would-be responses to the incomplete PRO-CTCAE items. This information may include patients’ responses to the same PRO-CTCAE item at different cycles, patients’ responses to other PRO-CTCAE items, and/or clinicians’ CTCAE grades . For example, a patient who reports no pain at Cycle 2 may be more likely to report no pain at Cycle 3. A patient who reports severe insomnia at Cycle 3 may be more likely to report severe fatigue at Cycle 3. If a clinician reports grade 1+ vomiting at Cycle 3, then the patient may be more likely to report severe vomiting at Cycle 3. This information helps multiple imputation make plausible guesses about what patients’ missing scores would have been had they completed the PRO-CTCAE assessment. In the imputation model, auxiliary variables correlate with an incomplete PRO-CTCAE item and/or missingness. Auxiliary variables that correlate with an incomplete PRO-CTCAE item improve power by providing information about the missing scores; auxiliary variables that correlate with an incomplete PRO-CTCAE item and its missingness indicator  reduce bias . In genet-test was conducted using the TTEST procedure ; complete-case logistic regression was conducted using the LOGISTIC procedure (for the PRO-CTCAE item assessing presence). Mixed modeling was performed using the MIXED procedure  or GLIMMIX procedure (for the PRO-CTCAE item assessing presence). Each mixed model included a fixed intercept; fixed effects for time, arm, and arm by time interaction; and autoregressive (lag 1) residual covariance matrix that accounts for repeated PRO-CTCAE assessments within patients. Time was treated as nominal, such that the mixed models did not make assumptions about the trajectory of patients’ PRO-CTCAE scores over time. A logit link function was specified within the mixed model for the PRO-CTCAE item assessing presence. Multiple imputation was conducted twice for each PRO-CTCAE item using the MI procedure. In the first imputation model, each PRO-CTCAE item was imputed based on patients’ responses to that PRO-CTCAE item at all available cycles. In the second imputation model, each PRO-CTCAE item was imputed based on patients’ responses to that PRO-CTCAE item at all available cycles plus relevant CTCAE grades at week 12  or LOGISTIC procedure (for the PRO-CTCAE item assessing presence), and results were pooled across the imputed datasets using the MIANALYZE procedure.All analyses were performed in SAS 9.4 . The complete-case two-sample sive lag  residualTo assess similarity of results, estimates of average treatment effects at week 12 and statistical efficiency were compared across strategies for addressing missing PRO-CTCAE scores. Statistical efficiency was measured by confidence interval width, with narrower confidence intervals indicating greater statistical efficiency.Of the 87 patients enrolled in A091105, 64 consented to participate in the correlative study. Of these 64 patients, 36 (56.3%) were randomized to the sorafenib arm and 28 (43.8%) were randomized to the placebo arm. The randomization ratio was not 2:1 (sorafenib:placebo) as specified in the protocol due to a randomization algorithm error that was detected and corrected partway through enrollment [Of the 119 patients enrolled in COMET-2, 107 completed the baseline PRO-CTCAE assessment and at least one post-baseline PRO-CTCAE assessment and were thus included in this analysis. Of these 107 patients, 53 (49.5%) were randomized to the cabozantinib arm and 54 (50.5%) were randomized to the mitoxantrone-prednisone arm. Median age was 65 years (range = 44 to 80 years). Baseline ECOG performance status was 0 or 1 for 94 (87.9%) patients. All patients had undergone at least two prior lines of systemic treatment for castration-resistant metastatic prostate cancer.n = 7), not having a clinic visit (n = 2), staff not administering the questionnaire booklet (n = 2), or unspecified reason (n = 2). Of the 51 patients who completed the PRO-CTCAE assessment at week 12, 50 completed all 19 items and 1 missed 5 of the 19 items due to skipping a page of the questionnaire booklet.In A091105, 100.0 to 52.8% and 100.0 to 39.3% of those randomized to the sorafenib and placebo arms, respectively, completed the PRO-CTCAE assessment between baseline and week 32  for mixed modeling and multiple imputation, whereas the sample size used for analysis was 50 or 51  for the complete-case two-sample st Table . Mixed mr = 0.627), though this correlation was inflated due to observing very few nonzero CTCAE grades for mucositis oral at week 12 , patient-reported frequency of nausea and clinician-reported nausea (r = 0.447), patient-reported severity of vomiting and clinician-reported vomiting (r = 0.441), and patient-reported interference of fatigue and clinician-reported fatigue (r = 0.436). The weakest correlations occurred between patient-reported interference of pain and clinician-reported arthralgia (r = 0.014), patient-reported severity of pain and clinician-reported myalgia (r = 0.059), and patient-reported frequency of diarrhea and clinician-reported diarrhea (r = 0.096).Table r = 0.427 and 0.445, respectively). Clinician-reported insomnia, fatigue, decreased appetite, and peripheral neuropathy did not strongly correlate with their patient-reported counterparts (range = − 0.102 to 0.046; Table Relative to A091105, COMET-2 had a much higher proportion of nonzero CTCAE grades due to targeting a more advanced cancer patient population and administering a more toxic chemotherapy regimen (Table In both trials, between-arm comparisons on the PRO-CTCAE at week 12 were similar regardless of whether CTCAE grades were included in the imputation model Tables  and 4. It-test, mixed modeling with contrast, and multiple imputation followed by a two-sample t-test. In both trials, mixed modeling and multiple imputation provided the most similar estimates of the average treatment effect. These results are unsurprising because, unlike a complete-case two-sample t-test, mixed modeling and multiple imputation provide unbiased parameter estimates under a missing at random mechanism and do not exclude patients with missing scores—highly desirable features per the SISAQOL Consortium [Properly handling missing PRO-CTCAE scores supports more accurate and generalizable causal inferences regarding treatment tolerability. In this paper, we conducted between-arm comparisons while applying the following strategies for addressing missing PRO-CTCAE scores in A091105 and COMET-2: complete-case two-sample nsortium .We also performed multiple imputation with and without CTCAE grades as auxiliary variables to assess the added benefit of including CTCAE grades in the imputation model relative to only including PRO-CTCAE scores across all cycles. Our results suggest that CTCAE grades can inform missing PRO-CTCAE scores for any adverse events that show strong agreement between clinician and patient reports, though model simplicity and computational ease may warrant use of other strategies. In A091105 and COMET-2, inclusion of CTCAE grades in the imputation model was not worthwhile because the strongest correlations occurred among the same PRO-CTCAE item at different cycles. These results make sense because the information provided by patients often differs from the information provided by clinicians. Thus, we recommend using patients’ PRO-CTCAE scores for the same symptom at different cycles to inform patients’ missing PRO-CTCAE scores. This can be accomplished via mixed modeling or multiple imputation. Although multiple imputation is widely available in statistical software packages, multiple imputation is much more demanding procedurally than mixed modeling. The user must examine convergence diagnostics; ensure the imputation model includes all the analysis variables; create, manage, and analyze multiple imputed datasets; and pool the results. The extent to which statistical software packages automate this procedure varies. Conducting mixed modeling for PRO-CTCAE items assessing frequency, severity, or interference is consistent with recommendations outlined by the SISAQOL Consortium . Howevert-test, mixed modeling with contrast, and multiple imputation followed by a two-sample t-test. Because we did not simulate the data, we cannot calculate bias of the parameter estimates. However, our results can serve as the basis for a future simulation study evaluating strategies for addressing missing PRO-CTCAE scores.Limitations of this work include our focus on two phase III trials with modest sample sizes. Examining other phase II and phase III trials as well as other patient populations may improve the generalizability of these results. However, the A091105 and COMET-2 sample sizes are not atypical for plausible applications of these strategies for addressing missing PRO-CTCAE scores. Another limitation is our focus on the similarity of parameter estimates and differences in statistical efficiency across the complete-case two-sample In summary, using patients’ PRO-CTCAE scores for the same symptom at different cycles to inform patients’ missing PRO-CTCAE scores can mitigate problems associated with missing scores. Accurately evaluating patients’ PRO-CTCAE scores promotes the safety and tolerability of treatments as well as improves the implementation and interpretation of cancer clinical trials.


This study compared the performance of four serology assays for Coronavirus Disease 2019 (COVID-19) and investigated whether COVID-19 disease history correlates with assay performance. Samples were tested at Northshore using the Elecsys Anti-SARS-CoV-2 (Roche Diagnostics), Access SARS-CoV-2 IgG anti-RBD (Beckman Coulter), and LIAISON SARS-CoV-2 S1/S2 IgG (DiaSorin) as well as at Genalyte using Maverick Multi-Antigen Serology Panel. The study included one hundred clinical samples collected before December 2019 and ninety-seven samples collected from convalescent plasma donors originally diagnosed with COVID-19 by PCR. COVID-19 disease history was self-reported by the plasma donors. There was no difference in specificity between the assays tested. Clinical sensitivity of these four tests was 98% , 96% (Roche), 92% (DiaSorin), and 87% (Beckman). The only statistically significant differences in clinical sensitivity was between the Beckman assay and both Genalyte and Roche assays. Convalescent plasma donor characteristics and disease symptoms did not correlate with false negative results from the Beckman and DiaSorin assays. All four tests showed high specificity (100%) and varying sensitivities (89–98%). No correlations between disease history and serology results were observed. The Genalyte Multiplex assay showed as good or better sensitivity to three other previously validated assays with FDA Emergency Use Authorizations. The Coronavirus Disease 2019 (COVID-19) pandemic created an urgent need for serology tests against a novel viral pathogen. Multiple companies rapidly developed assays to detect antibodies against the causative agent, SARS-CoV-2. These in vitro diagnostic tests were then submitted to the United States Food and Drug Administration (FDA) for Emergency Use Authorization (EUA), which permitted their marketing. Alternatively, in-house developed assays were validated as laboratory developed tests (LDTs).Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is known to bind to the angiotensin-converting enzyme II (ACE2) receptor in order to gain entry to target cells. Similar to the first severe acute respiratory syndrome coronavirus (SARS-CoV), it appears that a region of the coronavirus spike protein known as the receptor binding domain (RBD) is primarily responsible for the SARS-CoV-2-ACE2 interaction. Interestingly, the spike protein is “primed” for ACE2 binding via an interaction with another human protein called TMPRSS2 . TherefoDiaSorin LIAISON® SARS-CoV-2 [Beckman Coulter Access SARS-CoV-2 [Roche Elecsys® Anti-SARS-CoV-2 [SARS-CoV-2 are composed of 16 nonstructural proteins and 4 structural proteins; spike (S), envelope (E), membrane (M), and nucleocapsid (N). The RS-CoV-2  is an inRS-CoV-2  is a chePrimarily due to the novel nature of COVID-19, the correlation between antibody reactivity and immunity to relapse or re-infection is unknown . NeverthThe NorthShore COVID-19 Convalescent Plasma Collection Study was approved by the NorthShore Institutional Review Board (EH20-170). Written informed consent was obtained from convalescent plasma donors. Control samples collected prior to December 2019 were residual anonymized specimens derived from clinical samples, for which Western IRB granted an exception to informed consent for research use (WIRB#20161322).Convalescent plasma samples used in this study were collected from donors who had been recruited in April and May 2020. Recruitment efforts included messages sent to physicians within the NorthShore University HealthSystem, messages posted on internal websites, and listing on a national website for convalescent plasma collection centers. Participants were required to be greater than 18 years old, have recovered from COVID-19 greater than two weeks prior to consent, provide documentation of a positive SARS-CoV-2 PCR result confirming their diagnosis, and test negative by repeat nasopharyngeal SARS-CoV-2 PCR prior to sample collection. Potential donors were directed to call or email the NorthShore Blood Bank.One hundred frozen clinical samples collected prior to December 2019 by Genalyte were used as negative controls. These serum samples were collected in serum separator tubes, centrifuged for 15 minutes at 4,000 RPM, then stored in 1 ml aliquots at -80 C. All transported samples were shipped on dry ice. Samples from individuals who were confirmed to have COVID-19 were taken from convalescent plasma donors who volunteered for the NorthShore University HealthSystem COVID-19 Convalescent plasma collection study. NorthShore plasma donor samples were collected in serum separator tubes (BD vacutainer SST 367986), spun at 4,500 RPM for 7 minutes , and aliquots were frozen at -80 C.Samples were collected from convalescent plasma donors in BD serum separator tubes . The tubes were spun, ~1ml aliquots were made, and the aliquots were frozen at -80°C. An aliquot from each donor was shipped overnight on dry ice to Genalyte for testing; another aliquot was thawed and tested on the three other assays at NorthShore.Four serology assays were used to analyze the specimens. Three of these are assays with current Emergency Use Authorizations (EUA) from the Food and Drug Administration (FDA) and adapted for use on automated high-throughput chemistry analyzers. Each of these assays targets a single antigen of the SARS-CoV-2 virus and testing was performed in a CLIA accredited laboratory at NorthShore University Health System . The final of these is a novel assay manufactured by Genalyte  which probes for antibodies against multiple SARS-CoV-2 antigens and other viral antigens. Specimens are then determined to be positive or negative for SARS-CoV-2 antibodies through the use of a multivariate machine learning model. This testing was performed in San Diego at Genalyte’s headquarters.Aliquots tested at NorthShore were run on three automated assays: Elecsys Anti-SARS-CoV-2 (Roche Diagnostics), Access SARS-CoV-2 IgG (Beckman Coulter), and LIAISON SARS-CoV-2 S1/S2 IgG (DiaSorin). Each sample was tested on the same day using all three platforms. Each of these assays targets a different epitope of the SARS-CoV-2 virus and employs a different immunoassay format. The DiaSorin assay detects IgG antibodies against the S1 and S2 spike proteins of SARS-CoV-2 virus in an indirect chemiluminescent format. The Beckman assay detects IgG antibodies against the Receptor Binding Domain (RBD) of the S1 protein in a two-step immunoassay format. The Roche assay detects all antibody subtypes that are able to bind nucleocapsid viral antigens in a chemiluminescent sandwich immunoassay format. The Roche, Beckman, and DiaSorin assays are run on automated instruments with high specimen throughput capability, but relatively large physical footprint and startup costs. Conversely, the Genalyte assay is run on a non-automated device with relatively lower throughput, but also lower up-front acquisition costs and smaller physical footprint.The Genalyte assay employs multiplex detection technology based on silicon photonics that uses ring resonance to measure binding of macromolecules to sensors on a miniature silicon chip. The Maverick Diagnostic System detects changes in resonance wavelength as macromolecules, such as antibodies, bind to their respective antigens that are bound to the chip. Maverick™ SARS-CoV-2 Multi-Antigen Serology Panel  is designed to detect IgG and IgM class of antibodies to five SARS-CoV-2 antigens within a multiplex format based on photonic ring resonance technology , 11. A mh}, k = 1,…n where the {Θk} are independent identically distributed random vectors and each tree casts a unit vote for the most popular class at input X. In particular, each new training set is drawn, with replacement, from the original training set. Then a tree is grown on the new training set using random feature selection. The generalization error of a forest of tree classifiers depends on the strength of the individual trees in the forest and the correlation between them. In the random forest model, the number of features to consider when looking for the best split at each node is set to Genalyte’s Multi-Antigen Analysis Algorithm (MAAA) employs the well-known Random Forest Ensemble method with 3000 decision “trees”. Random forest is a classifier consisting of a collection of decision trees {f splits .Convalescent plasma donor COVID-19 disease history was obtained at the time of participant consent via a donor history screening form and blood donor screening questions see . Free-teGenalyte assay data provided target-specific quantitative response broken down into IgG and IgM-based reactivity. The target-specific data from the 5 SARS-Cov2 targets was compared with the qualitative results of both the Beckman and DiaSorin assays to determine whether the results of the Genalyte targets correlated with the likelihood of the Beckman and DiaSorin assays being positive. P values were calculated with two-tailed Fisher’s exact test on a contingency table of true positives versus false negatives. The Haldane-Anscombe correction was used to calculate real-valued odds ratios for instances when a zero-valued denominator would have otherwise been present .Due to a lack of a priori knowledge of the expected range of serology test results for antibodies against a SARS-CoV-2, a power calculation was not performed prior to initiation of this study. The significance of pair-wise differences in sensitivity between the four assays tested was determined using a two-tailed Fischer’s exact test on a 2x2 table of true positives and false negatives. Target-specific analysis of the samples producing a false-negative result on the Beckman and DiaSorin assays was performed using the Fischer’s exact test on the qualitative results of both anti-spike protein assays along with the target-specific results of the Genalyte assay.The Maverick TM SARS-CoV-2 Multi-Antigen Serology Panel v2 utilizes a multi-analyte analysisalgorithm (MAAA) to make the determination on patient samples of positive or negative orindeterminate for antibodies to the SARS-CoV-2 virus. The algorithm employed is a well-knownensemble method called Random Forests Classification. Random Forests contain a number ofdecision trees constructed from randomly chosen features that each make predictions on the data set, the aggregation of which gives the final result. These models are capable of fitting complex datasets and are resistant to overfitting.Our implementation of the method uses 3000 such decision trees sampled randomly from training data and validated against test data. The model was also cross validated using five-fold cross validation. Three models were trained, and the combined IgG and IgM model proved to be the most robust, and is the model carried forward to call samples positive or negative or indeterminate for antibodies to SARS-CoV-2.There was no statistically significant difference in donor gender, age, symptom duration, time from symptom resolution to sample collection, or difference in the presence or absence of any of the symptoms measured see  between The total antibody assays  showed higher sensitivity compared with the IgG assays (Beckman and DiaSorin), see Among the samples from patient’s with PCR-proven COVID-19, the Genalyte assay detected anti-S1-F IgG most often and anti-NC IgM the least often see . SamplesWhen SARS-CoV-2 serology assays were initially developed, there was concern that false-positive rates in these assays would be high due to cross reactivity with antibodies formed against non-SARS-CoV-2 coronaviruses . While tThe false negative results detected here were most common with the assay that uses the fewest number of SARS-CoV-2 epitopes as its target, in particular the RBD domain of the spike protein. Although this domain is clearly important for viral infectivity, it is likely that antibodies against other viral epitopes may also diminish viral infectivity. This could occur by inducing or preventing conformational changes in viral proteins that are necessary for fusion with the cell membrane as well as by promoting viral clearance through opsonization. However, it is important to note that binding antibody strength does not necessarily correlate with neutralizing antibody titer.Finally, this study tested the hypothesis that false negative results on serology assays may correlate with certain patient characteristics  or COVID-19 disease symptoms. Such a correlation was not found for false negative results from the two anti-spike assays. This study included insufficient numbers of false negative results from the other two assays tested to draw a meaningful conclusion.Given the lack of COVID-19 PCR testing in the United States during the initial weeks of the SARS-CoV-2 pandemic, the high rate of very mild or asymptomatic infections, and the real possibility of false negative PCR assays, serology testing will likely remain a critical component of COVID-19 clinical diagnostics in addition to epidemiologic study. Although it remains unclear whether antibodies against certain SARS-CoV-2 epitopes can confer immunity to reinfection, it is apparent that the serologic response to COVID-19 is heterogeneous .S1 Fig(TIF)Click here for additional data file.


Recently, mesenchymal stem/stromal cells (MSCs) due to their pro-angiogenic, anti-apoptotic, and immunoregulatory competencies along with fewer ethical issues are presented as a rational strategy for regenerative medicine. Current reports have signified that the pleiotropic effects of MSCs are not related to their differentiation potentials, but rather are exerted through the release of soluble paracrine molecules. Being nano-sized, non-toxic, biocompatible, barely immunogenic, and owning targeting capability and organotropism, exosomes are considered nanocarriers for their possible use in diagnosis and therapy. Exosomes convey functional molecules such as long non-coding RNAs (lncRNAs) and micro-RNAs (miRNAs), proteins , and lipids from MSCs to the target cells. They participate in intercellular interaction procedures and enable the repair of damaged or diseased tissues and organs. Findings have evidenced that exosomes alone are liable for the beneficial influences of MSCs in a myriad of experimental models, suggesting that MSC- exosomes can be utilized to establish a novel cell-free therapeutic strategy for the treatment of varied human disorders, encompassing myocardial infarction (MI), CNS-related disorders, musculoskeletal disorders (e.g. arthritis), kidney diseases, liver diseases, lung diseases, as well as cutaneous wounds. Importantly, compared with MSCs, MSC- exosomes serve more steady entities and reduced safety risks concerning the injection of live cells, such as microvasculature occlusion risk. In the current review, we will discuss the therapeutic potential of MSC- exosomes as an innovative approach in the context of regenerative medicine and highlight the recent knowledge on MSC- exosomes in translational medicine, focusing on in vivo researches. Mesenchymal stem/stromal cells (MSCs) can be obtained from a variety of human tissues, ranging from bone marrow (BM) to umbilical cord (UC), typically showing multipotent differentiation competencies –4. BecauConcerning the definition of the International Society for Extracellular Vesicles (ISEV), EVs are lipid bilayer particles which are naturally secreted from the cells, and do not show replication capacity. The ISEVs have also cited that the two classes (classes 1 and 2) of EV markers and one class of non-EV markers (class 3) are essential to be inspected for demonstration of the existence of EVs . Class 1Generally, EVs are categorized into 3 subclasses regarding sizes and biogenesis procedures, surrounding exosomes 50–150 nm), microvesicles (MVs) (100–1000 nm), and apoptotic bodies (ApoBDs) (500–5000 nm) . Exosome0 nm, micIn the 1960s, Friedenstein and his colleagues for the first time found that the osteogenic capability, as demonstrated through heterotopic transplantation of BM cells, was dependent on a BM cell's minor subpopulation . They foDespites of the expression of similar levels of the surface antigen by MSCs derived from diverse sources, some reports have evidenced differences between them. In this regard, UC-MSCs demonstrated higher proliferation rate, and lower expression of p53, p21, and p16 against BM-MSCs and A-MSCs . BesidesIn 1983, exosomes were firstly identified by Johnstone et al. as vesicles that contribute to mammalian reticulocyte differentiation and maturing . ExosomeDespite their inherent biological activities, exosomes are recently introduced as encouraging drug carriers that rely on their small size, supreme biocompatibility, and aptitude to load particular and different therapeutic ingredients such as proteins, nucleic acids, and small molecules . HoweverThe exosome involves numerous cytoplasmic and membrane proteins, including receptors, enzymes, transcription factors, extracellular matrix (ECM) proteins, nucleic acids , and also lipids . ExaminaEVs proteome studies are novel strategies and are part of the rising interest in proteomics investigations. To date, a large number of proteomics studies have been carried out on EVs to clarify their varied roles. Today, mass spectrometry (MS) is a major method for the recognition and characterization of the protein content of EVs. As pronounced, the EVs can be generated by a spectrum of various cell types and they can be obtained from biological fluids and also conditioned medium (CM) , 65. A dAlthough MSC-EVs derived from different sources mainly show remarkable therapeutic influences, they have various physiological functions, which can affect their therapeutic applications. For instance, EVs obtained from hBMMSCs show superior potential to treat cartilage defects or osteoarthritis which are in association with bone diseases , 68. AlsRendering literature, there are some challenges in the clinical application of exosomes, especially in regenerative medicine. Between various utilized isolation and purification strategies, there is no general approach for exosomes separation from other micro-particles and also several exosomes subset’s separations , 74. UntAs described, MSC- exosomes could serve several therapeutic benefits such as lower immunogenicity and upgraded safety profiles compared with MSC, and thereby are considered a rational substitute to whole-cell therapy to treat a wide spectrum of human disorders by delivering functional molecules to the target cells. In exosomes, proteomic examinations have led to the recognition of hundreds of proteins participating in central biological events, including Exo biogenesis, cellular construction, tissue recovery, and also inflammatory response . As wellVarious preclinical studies have been carried out to address the therapeutic efficacy of the MSC- exosomes in lung inflammatory disorders Table . InvestiOn the other hand, investigation of the therapeutic effects of exosomes in a neonatal mouse model of bronchopulmonary dysplasia (BPD) verified beneficial effects of human UCMSC- exosomes therapy which were closely linked to tumor necrosis factor alpha-stimulated gene-6 (TSG-6) . The TSGIn humans, a prospective nonrandomized open-label cohort study was carried out in April 2020, and verified the safety and efficacy of exosomes (ExoFlo™) derived from allogeneic BM-MSCs for treatment of 24 patients with severe 2019 coronavirus disease (COVID-19). Accordingly, a single intravenous injection of ExoFlo resulted in alleviated patient’s clinical condition and oxygenation in the absence of any unwanted events during 2 weeks follow-up. Robust improvements in neutrophil count and reduction in acute phase reactants, such as C-reactive protein (CRP), ferritin, and D-dimer, described MSCs- exosomes as a capable therapeutic candidate for severe COVID-19 .Recent findings have suggested that MSCs- exosomes could transfer certain bioactive molecules and sustain repair and recovery of damaged liver. Correspondingly, investigation of the therapeutic capacity and underlying molecular mechanism for human BMMSCs- exosomes in carbon tetrachloride (CCl4)-induced rat liver fibrosis indicated that Exo therapy effectively improved liver fibrosis, as shown by a lessening in collagen assembly, improved liver activities, suppressed inflammation, and also promotion of the hepatocyte regeneration. Moreover, exosome supported a decrease in MDA, IL-1, and IL-6 serum levels and also suppressed the Wnt/β-catenin pathway axis in hepatic stellate cells (HSC), verifying that BMMSCs- exosomes could improve CCl4-induced liver fibrosis . ImportaOn the other hand, injection of UCMSCs- exosomes led to a reduction in serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and pro-inflammatory cytokines levels and also inhibited NLR family pyrin domain containing 3 (NLRP3) activation in ALF animal models. Based on observations, exosomal miRNA-299-3p was responsible for the inhibition of inflammatory responses and also NLRP3 stimulation in transplanted models, enabling liver tissue repair . SimilarBesides, human menstrual blood-derived stem cells (MenSCs)-derived exosomes could express cytokines, such as intercellular adhesion molecule-1 (ICAM-1), angiopoietin-2, tyrosine kinase Axl, angiogenin, insulin-like growth factor-binding protein 6 (IGFBP-6), osteoprotegerin, IL-6, and IL-8, improving liver function and abrogating liver cell’s apoptosis in mice model . On the MSCs- exosomes have been described to recover kidney function, reduce kidney damage, and inhibit chronic kidney fibrosis by various mechanisms, such as inhibition of the pro-fibrotic genes TGF-β1. Evaluation of whether miRNAs deregulation constrains the regenerative competencies of MSCs- exosomes in a model of glycerol-induced acute kidney injury (AKI) in severe combined immunodeficient mice showed that kidney genes deregulated following damage were alleviated via MSCs- exosomes treatment but not via Drosha-knockdown vesicles, thus signifying a crucial role of miRNAs in recovery after AKI . FurtherC) supported by the upregulation of miR-4516 [C can bind and sustain mitochondrial kinase PTEN-induced kinase 1 (PINK1), playing important role in the protection of mitochondrial against oxidative stress-stimulated apoptosis and averting mitochondrial stress [Molecular analysis has demonstrated that miR-4516 expression commonly is down-regulated in the kidney cortex of chronic kidney disease (CKD) mice models and leads to the triggered cytoskeleton reorganization and mitochondrial dysfunction, and stimulates kidney fibrosis. Therefore, there is a hypothesis that improved expression of miR-4516 by melatonin or other molecules could decrease ROS formation and finally ameliorate mitochondrial function . CollectmiR-4516 . PrPC cal stress . In vitrl stress .Recently, several studies in animal models have evidenced the potent capacities of the MSC- exosomes therapy in CNS-related disorders, ranging from neurodegenerative diseases to spinal cord injury (SCI) Table . RenderiInvestigations of MSC- exosomes therapy in musculoskeletal disorders have resulted in promising consequences in pre-clinical models Table . It seemCurrent reports have shown that the exosomes derived from MSCs could sustain chondrocyte homeostasis and improve the pathological sternness of OA in vivo, making them a potent strategy to treat OA. Investigation of the possible beneficial effects of the infrapatellar fat pad (IPFP) MSCs-derived exosomes (MSCIPFP- exosomes) on OA revealed that Exo could alleviate the OA severity by inhibition of chondrocyte cell apoptosis, improvement of matrix production, and a reduction in the presentation of a catabolic factor in vitro. Also, MSCIPFP- exosomes boosted autophagy rates in chondrocytes by negative regulation of mechanistic target of rapamycin (mTOR) because of the existence of high-levels of miR-100-5p in MSCIPFP- exosomes. In OA murine model, MSCIPFP- exosomes restored articular cartilage’s damages and gait abnormality by sustaining cartilage homeostasis, which relied on miR100-5p-mediated modulation of the mTOR-autophagy pathway . SimilarPrevious studies have shown that Wnt5a exists in human OA cartilage and could boost chondrocyte catabolic functions by non-canonical Wnt signaling, which proposes a possible role in OA progress by various mechanisms, such as the promotion of MMP1 and MMP13 expression . AccordiMSCs- exosomes also could exert functional recovery in RA murine models by induction of inhibitory effect on effector immune cells. For instance, MSCs-derived exosomes in collagen-induced arthritis (CIA) models inhibited T lymphocyte proliferation and also reduced the frequencies of CD4-positive and CD8-positive T cell subsets.On the other hand, MSCs-exosomes augmented Treg cell percentage, and decreased plasmablast differentiation, thus documenting the positive effects of MSCs- exosomes in RA by suppressing immune responses . Besides−/− mice quickly expired because of the systemic inflammation elicited by spontaneously activated NF-κB [Recently, studies have demonstrated that tumor necrosis factor-α-induced-protein 3 (TNFAIP3) gene polymorphisms (rs2230926 and rs5029937) are allied with the promoted risk of RA , and alsed NF-κB .Respecting recent studies, MSCs-exosomes are known to contribute to myocardial repair following myocardial infarction (MI), while the underlying mechanism is not entirely elucidated. Sphingosine-1-phosphate (S1P) is an important regulator of the immune-inflammatory response; thereby its role in MI has been widely confirmed. Owing to this fact, it has been found that MSCs derived from adipose tissue (AMSCs) and their exosomes may stimulate a recovery in MI by repressing cardiac dysfunction, apoptosis, and fibrosis, accompanied with inhibition of inflammatory reactions in vitro and in vivo. Despite the macrophage M2 polarization induced by theses Exo, it seems that S1P/ sphingosine kinase 1 (SK1)/sphingosine-1-phosphate receptor-1 (S1PR1) axis is responsible for AMSCs- exosomes -mediated myocardial recovery because S1PR1 silencing mitigated the suppressive influences of Exo on MI-stimulated cardiac apoptosis and fibrosis in vitro. Therefore, observations indicated that AMSCs-Exo could alleviate MI mechanistically through activating S1P/SK1/S1PR1 signaling and promoting M1- to M2-phenotype shift . In thisBesides, Exo derived from macrophage migration inhibitory factor (MIF)-overexpressed MSCs restored heart function, decreased heart remodeling, supported mitochondrial functions, and suppressed ROS generation in MI murine models . These fChronic wounds are mutual and endure to be the one cause of morbidity and mortality, while therapeutic options for these conditions are missing and often unsuccessful. Collecting proofs have showed the efficacy of MSCs- exosomes to repair and regenerate the damaged area, and thereby sustain cutaneous wound healing. MSCs- exosomes activates numerous axis contributing to wound healing, such as AKT, ERK, and STAT3, and also trigger the expression of hepatocyte growth factor (HGF), IGF1, nerve growth factor (NGF), and stromal-derived growth factor-1 (SDF1), providing an opportunity for wound healing cell-free based therapies . Also, pMSCs- exosomes are now being broadly applied to advance innovative regenerative approaches for several disorders as they serve most of the therapeutic attributes of MSCs. Exosomes provide a chance for cell-free therapy, which diminishes safety issues concerning the use of viable cells . During


Transcranial magnetic stimulation (TMS) is a non-invasive brain stimulation tool designed to probe the strength of inhibitory and excitatory neurotransmission in the cortex. Combined with electromyography, paired-pulse TMS paradigms have revealed a deficit in inhibition mediated by GABA-A receptors in patients with schizophrenia. Combined TMS-electroencephalography (TMS-EEG) provides a more detailed examination of cortical excitability and may shed more light into the pathophysiology of schizophrenia. Of the various peaks of the TMS-evoked EEG signal, responses at 45 (N45) and 100 ms (N100) likely reflect GABA-A and GABA-B receptor-mediated inhibition, respectively. Responses at 25 ms (P25) are affected by voltage-gated channel ligands, whereas glutamatergic processes may be related to the P70 component. We here aim to systematically investigate the role of these neural processes in patients with schizophrenia by using TMS-EEG.TMS-evoked EEG potentials (TEPs) were recorded in patients with schizophrenia (n = 19) and in age-matched healthy controls (n = 17). 150 TMS pulses at 90% of resting motor threshold were applied over the left primary motor cortex during EEG recording. Differences in TEPs between the two groups were analysed for all electrodes and for time windows corresponding to each TEP  by applying multiple independent sample t-tests. Further, a cluster-based permutation analysis approach was implemented to correct for multiple comparisons.Compared to controls, patients showed amplitude reduction for the P25 , N45  and P70 component .There results extend on previous literature about impairment of GABA-A receptor mediated inhibition in schizophrenia, as demonstrated by the N45 amplitude reduction, whereas no significant differences in GABA-B index  were revealed. Our results also showed that, although specific mechanisms underlying P25 and P70 have not been fully elucidated yet, excitatory neurotransmission is altered in this clinical population. To conclude, TMS-EEG may provide a more comprehensive view of the inhibitory and excitatory mechanisms involved in the pathophysiology of schizophrenia.