Since the first reporton the pharmacological activity of curcuminin 1949, enormous amounts of research have reported diverse activitiesfor this natural polyphenol found in the dietary spice turmeric. However,curcumin has not yet been used for human application as an approveddrug. The clinical translation of curcumin has been hampered due toits low solubility and bioavailability. The improvement in bioavailabilityand solubility of curcumin can be achieved by its formulation usingdrug delivery systems. Hydrogels with their biocompatibility and lowtoxicity effects have shown a substantial impact on the successfulformulation of hydrophobic drugs for human clinical trials. This reviewfocuses on hydrogel-based delivery systems for curcumin and describesits applications as anti-cancer as well as wound healing agents. The most frequently investigated natural polymersused for hydrogel preparation are chitosan, dextrin, lignin, hyaluronicacid, carrageenan, tannic acid, alginic acid, and collagen.20 Synthetic and semisynthetic polymers can also be used for hydrogelpreparation.21 These polymers mainly includepolyethylene glycol (PEG), poly lactic acid, poly lactic coglycolicacid (PLGA), and poly vinyl alcohol (PVA) as well as carboxymethylcellulose (CMC), hydroxyethyl cellulose, hydroxypropyl cellulose,and hydroxypropyl methyl cellulose.29 Based on the monomers used for the preparation of the polymericnetwork, hydrogels may contain homo-polymeric, co-polymeric, or multi-polymericnetworks.31 While polymerization of a single type ofmonomers results in the formation of homo-polymeric networks, co-polymericor multi-polymeric networks result from the polymerization of twoor more types of monomers.33Hydrogels are three-dimensionalcross-linked polymeric networks with the ability to absorb water dueto the presence of hydrophilic functional groups in their structure.37 One of the major obstacles for drug development is the administrationof hydrophobic drugs with low solubility in aqueous media.38 Considering the unique properties of hydrogels,this delivery system can be used as a promising vehicle for formulationof low soluble hydrophobic therapeutics.40 This approach enables researchers and patientsto apply these therapeutic agents via oral, topical or parenteralroutes of administration.41The biocompatibility,swelling behavior, and low toxicity of hydrogels make them suitablecandidates for biomedical applications including drug delivery.1.2Curcuma longa species.42 From ancient days, it has been using in Asian food andtraditional medicine.43 As earlier defined,two chemical units (2 o-methoxy phenol) of the moleculeare connected by a seven-carbon linker with an α,β-unsaturateddiketone moiety.45 Curcumin acts as an electrondonor, and the π electron cloud stabilizes its chemical structure.The resonance structure exhibited inside the molecule is responsiblefor its contribution to many electron transfer reactions.47 Lampe first described the procedure for the synthesis of syntheticcurcumin in 1913.48 Later on, several scientistsdeveloped methods for high yield synthesis of curcumin which are inuse today.49 Curcumin is the most widelyused plant-based drug with various pharmacological benefits, includinganti-inflammatory, antioxidant, anti-viral, anti-bacterial, anti-fungal,anti-parasite, and anti-cancer.59 Numerous preclinical studies have proven curcumin to be effectivein several cancers because of curcumin’s capability to induceG2/M cell cycle arrest, trigger apoptosis, induce autophagy, disturbmolecular signaling, inhibit invasion and metastasis, and increasethe efficiency of existing chemotherapeutics.62 Theinflammatory response of curcumin is often steered by the radicalproduction of pro-inflammatory cytokines, including interleukin-6(IL-6), interleukin-1β (IL-1β), and tumor necrosis factor-α(TNF-α).64 Consequently, the downregulationof pro-inflammatory cytokines may effectively reduce the incidenceof inflammation.65 Curcumin is also involvedin other signaling pathways like it induces degranulation in humanneutrophils by increasing the cell surface expression of clustersof differentiation 35 (CD35), CD66b, and CD63.66 However, its clinical usage is limited due to its low bioavailabilityand poor water solubility.68 Further, curcumin utilization in food supplements and nutraceuticalproducts is challenging due to its high chemical instability.69 Curcumin shows a tendency to crystallize inaqueous acidic conditions and is unstable to chemical degradationin basic aqueous conditions, which is attributed to changes in themolecular structure of curcumin.70 Hence,scientists are developing various kinds of curcumin nanoformulationto overcome these main obstacles and improve its superior therapeuticefficacy.78Curcumin -1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6heptadiene-3,5-dione) is a bright yellow polyphenol extracted fromthe roots of 1.387 The biological activity of curcumin was first reported by Schraufstätterand Bernt, where they showed the anti-bacterial activity of the compoundagainst Staphylococcus aureus.88 Despite promising biological properties, low cost, andavailability in bulk, a limited number of human clinical trials havebeen reported for curcumin and its derivatives.89 The major challenge for human application of curcumin is its poorbioavailability resulting from low solubility and stability.62 Following the administration of substantialdoses of curcumin, its plasma level becomes negligible within hours.91 Considering the unique properties of hydrogels, they can improvethe bioavailability, solubility, and stability of curcumin for medicalapplications97 , preparation of diverse formulations for differentroutes of administration may facilitate their bench to bedside translation.105Various plant-derived natural products have been used for humanapplications as drugs or supplementary agents.ations97 1. This d2124 forexample, it inhibits vascular endothelial growth factor (VEGF) andits receptor VEGR-2 and fibroblast growing-factor 2 (FGF2) and downregulatesmatrix metalloproteinases (MMPs) 2 and 9  plays a significant role in cancer invasion and metastasisin epithelial-derived cancers, curcumin might represent a promisingtherapeutic agent in human hepatocellular carcinoma by inhibitingthe transforming growth factor-β1 (TGF-β1)-induced EMTin liver cells .aterials 2.134,1352.1138 The gelation mechanism of injectablehydrogels is further categorized into chemical and physical cross-linkedhydrogels.139 There are also several injectablecurcumin hydrogels developed for effective cancer therapy. For example,to promote the water solubility and bioavailability of curcumin, itis encapsulated in liposome (Cur-Lip). A curcumin liposome hydrogel(CSSH/Cur-Lip Gel) is formed through the biochemical cross-connectionof free thiol groups and thermoregulation during the addition of β-glycerolphosphate, which can be used as an injectable hydrogel  and curcumin, as a composite film, can be usedfor liver cancer.151 The curcumin-loadedfilm can be used as a promising delivery system against breast andliver cancers. Thanks to the incorporation of cellulose nanocrystals(CNCs), the objective was fulfilled by improving the encapsulationefficiency and maximizing the efficacy of loaded curcumin througha sustained release profile. Curcumin showed a slow release profileleading to improved bioavailability and prevented rapid metabolismand clearance from blood.152 Polyethyleneglycol d-α-tocopheryl succinate 1000 (TPGS) is a water-solubleformulation derived from vitamin E that acts as a surfactant. Thismaterial is able to form micellar NPs in water.153 Delivery of curcumin using this system resulted in a slowrelease of the drug and showed substantial effects on decreasing ROSconcentration and developing apoptosis of HT-29 colon cancer cells in vitro. In addition, it seems that the bioavailabilityof oral curcumin formulation by TPGS can be increased compared withfree curcumin.133Polymers such as poly have shown proper film-formingproperties due to the abundance of OH groups in their structure.In vitro and in vivo drug release and cytotoxicity studies showed long-term sustainedantitumor effects compared to the free drug or single drug-loadedhydrogel formulation.153A promising newapproach to overcome the hydrophobicity of curcumin is the applicationof injectable thixotropic hydrogels made from silk fibroin/hydroxylpropyl cellulose. 154 Encapsulated into polymeric micelles, hydrophobic drugsform stable water-based formulations used for intravenous or intraperitonealapplications. In vitro tests suggest that the hydrogelsystem can release curcumin in a controlled manner. In addition, curcuminhydrogels can significantly suppress tumor growth and metastasis ina mouse model of colorectal and peritoneal carcinomatosis. Furthermore,curcumin hydrogels suppressed proliferation, induced apoptosis, andreduced angiogenesis of tumors.155Polymericmicelles are commonly used as drug delivery systems toovercome the low solubility of hydrophobic drugs.156 glycyrrhetinic acid was employed to preparesupramolecular curcumin pro-gelator (GA-Cur). The targeted deliverysystem showed continuous release of the drug from the formulationthrough hydrolysis of the ester bond. GA-Cur is a promising targetedapproach for hepatic delivery of curcumin.157 Many other types of curcumin-loaded hydrogels are listed in In order to develop a liver-targeted delivery system for curcumin,3172 Curcumin represses the action of the inflammatory transcript factorNF-κB, which is responsible for controlling several genes involvedin the initial onset of the inflammatory reaction.173 Curcumin effectiveness for wound healing has been limitedin the clinical trials due to its poor solubility and fast metabolismas well as low uptake and poor pharmacokinetic properties and bioavailability.175 Various biopolymers for thedelivery of curcumin have been investigated, including chitosan, starch,zein, alginate, and silk.176 These systems haveshown various characteristics including biodegradability and biocompatibility,as well as a wide range of commercial applications, making them idealcandidates for drug delivery applications.177 These unique properties allow drugs or cells to be simply combinedinto aqueous polymer solutions via simply mixing and then adding theformulations into target tissue to form a gel in situ to act as adrug delivery system.178 Poor water solubilityof curcumin and its low bioavailability restrict its applications.To overcome these limitations, encapsulating curcumin in a hydrogelnetwork is a helpful strategy.179 The formationof an absorbent hydrogel is important for wound healing since it providesoxygen and stores large amounts of water.175 In addition, a thin layer of hydrogel on the wound surface protectsit from air damage.169Wound healingconsists of three phases including inflammation,proliferation plus the formation of granulation tissue, and mediumformation and conversion.180 Therefore,the humid environment provided by the hydrogel helps skin regeneration.In addition, hydrogels absorb growth factors and cytokines from plasmaor wound exudate and promote cell proliferation and relocation besidestheir angiogenesis to enhance wound healing  showed higher tightening efficiency, significanttransdermal permeability, enhanced drug release, and high cell viabilityfor transdermal application.186 The swellingbehavior of the dual loaded hydrogel was more than 1.2 times thatof the gelatin-free hydrogel. This system increases the water absorptionwith mixed gelatin.187 Since gelatin isdispersed throughout the polymer grid and acts as a cell glue183 and the antioxidant properties of curcumincan provide additional assistance for wound dressings,188 chitosan-P123 nano curcumin-gelatin showedenhanced wound dressing properties compared to single loaded hydrogels.This injectable, biocompatible, biodegradable, and thermo-reversiblehydrogel can be used as optimal promising biomaterial for controlleddrug delivery, tissue restoration, or other therapeutic applications.183Chitosan-pluronicP123-curcumin-gelatin is a promising candidate for the developmentof injectable hydrogels designed for wound healing.189 Loading curcumininto amphiphilic alkylated cerium oxide NPs improved its bioavailabilityand use at the wound location. These NPs show improved bioavailabilityand antioxidant properties. The hydrogel has multiple properties,illustrates continuous drug release (approximately 63% in 108 h),accelerates cell movement, and provides a remarkable antioxidant andanti-inflammatory activity in vivo (approximately39%). NPs and NP-loaded hydrogel systems showed potential antioxidanteffects and increased cell migration. Furthermore, the results ofthe protein reactivity of carrageenan as a model for inflammationshowed the development of effective anti-inflammatory effects of NPswhich can be considered as candidates for wound healing.190A curcumin and 2-hydroxypropyl gammaring-shaped cyclodextrin (Cur/HP-γ-CDS) complex in a Sacranhydrogel film (Sac-HGF) shows the highest wound healing capabilityin nude mice due to the increased solubility of curcumin and its antioxidantaction. Therefore, Sac-HGF can be used as a candidate biomaterialfor wound dressings.191 Wounds healed with micellar curcumin-loadedthermo-sensitive hydrogel (Cur-MH) showed significant dehydrationwith no sign of pathological fluid leakage commencing the wound. Insummary, compared to the normal saline-treated wound (NS treatment),the wound had no sign of inflammation or infection. In vitro experiments show that Cur-MH compound gel exchanges at about bodytemperature. Follow-up in vivo experiments foundCur-MH effective in rats. In full-thickness in line cut and removalwound models, it has a good effect.180 Researchersdeveloped a curcumin-loaded nanoemulgel (Cur-NEG) through high-energyultrasonic emulsification to improve wound healing in vivo. They used a minimum concentration of surfactant to prepare nanogels.192 As shown in The honey-curcumin hydrogel sponge canbe formulated by easy additionand in situ polymerization. Due to its high fluid absorption capacity,the hydrogel matrix provides a dry wound area. Chitosan and honeycontribute to faster wound healing. The prepared sponge is highlyflexible and has fine mechanical strength. It permits low penetrationof moisture and shows controlled diffusion of curcumin.3.2193 The nanocurcumin-loadedhydrogel can effectively improve wound healing by enhancing earlythe re-epithelialization process. Various properties of pro-curcuminand nanocurcumin hydrogels on wound healing can be elucidated by in vitro firmness examinations.194 In addition, collagen deposition in the wound was stained with Masson’sTrichrome, indicating that N,O-carboxymethyl chitosan/oxidized alginatehydrogel (CCS/OA) can effectively improve collagen deposition in granulationtissue. Nanocurcumin/CCS/OA hydrogel therapy significantly increasesDNA and protein content in the wound tissue, indicating frequent cellproliferation in the wound tissue as a result of promoted wound healing.195Curcumin nanomicelles can prevent curcumindegradation. Curcumin nanomicelles combined with dextran hydrogelare used to enhance continuous release of curcumin from hydrated dressingsin order to reduce inflammation, promote fibroblast proliferationand collagen synthesis, as well as promote full-thickness wound healingand improve angiogenesis 2.193 The196 In-situ-forming hydrogelscan completely cover the wound area.197 In an investigation, the curcumin-phospholipid complex (CPC) wasprepared by the interaction between the choline-containing phospholipids(CCPLs) and the hydroxyl groups of curcumin. In this study, in vitro tests showed suitable curcumin release. CPC-ISGsignificantly improved the epidermis healing, which is quite significantfor primitive skin formation.169In-situ-forming hydrogels (ISGs) areusually fluid liquids, and as they come into contact with physiologicalsurroundings such as ions, pH, and temperature, they suddenly turninto gels.187 This developsorganized deposition of collagen plus VEGF synthesis and expressionof Aquaporin-3 in diabetic wounds.198 Toextend the method for efficient and safe affixing wound healing indiabetics, a thermoresponsive hydrogel containing nanomedicine, loadedin gelatin microspheres (GMs), was designed and developed to transportcurcumin as a wound healing agent.187 Curcuminwas then joined to shape self-assembled nanoparticles (CNPs) usingreprecipitation to improve its solubility and stability. CNPs, coatedwith GMs, might be able to react with matrix metalloproteinase 9 (MMP-9),which is often overexpressed and persists at the site of non-healingskin lesions in diabetes.199 As a result,the skin recovery process demands an equilibrium between the collagenbreakdown and the production of new components of the extracellularmatrix,200 as determined by substrate MMPand tissue metalloproteinase inhibitors.187Recent studies show that curcumin NPs have a substantial effecton the formation of new blood vessels and endothelial cells, whichaccelerate wound healing.3.3201 As shown by the curcumin hydrogelformulation by PVA, the hydrogel has great antibacterial efficacycompared to the PVA hydrogel alone. The designed curcumin-PVA hydrogelhas shown increased antibacterial activity with increasing curcuminconcentration. This hydrogel formulation was tested against S. aureus and E. coli.202 Gelatin hydrogel dressings are made of ion-adapted self-organizedbacterial cellulose extracted from Glucanoacetobacter xylinus. The curcumin-loaded membranes were capable of diminishing the growthof Gram-positive and Gram-negative bacteria. This was also shown bythe morphological study of living and dead bacteria and by fluorescencecolor analysis.203 Several similar curcuminhydrogels are studied in wound healing applications.206 Various wound healing applications of curcumin-loaded hydrogelsare listed in The main cause of delayed wound healingis the infection of a wound due to bacterial growth around the wound.Therefore, it is essential to design hydrogels with high antibacterialactivity.457 It has been recognized asbeneficial in treating anti-inflammatory, anti-allergic, antioxidant,anti-hyperglycaemic, anti-cancer, antimicrobial, anti-atherosclerosis,and anti-hypertension properties. Curcumin’s ability to affecta large variety of molecular targets and its good safety profile establishedit to be a potential candidate for the treatment of many diseases.Over the decade curcumin received extensive consideration due to advancesin its potential therapeutic applications. Nevertheless, clinicalapplications of curcumin are minimal due to its poor solubility andbioavailability. To overcome these problems, the development of specificcurcumin-encapsulated nanocarriers (nanocurcumin) has tremendous interestand enhances its applications. With detailed literature investigation,nanocurcumins, including curcumin hydrogels, improve the pharmacokineticproperties of curcumin that offer better therapeutic value. So far,many hydrogels mediated nanocurcumin and other nanocurcumins, studiedfor the proof of concept only. Very few clinical studies of nanocurcuminshave been conducted, which showed favorable features such as bioavailabilityand retention time compared to curcumin alone and systemic safety.Still, there is a significant gap in the research field to assessthe safety and efficacy of nanocurcumin formulations in humans. Itrequires thoughtful and dedicated research efforts. We believe thatour present review article on hydrogel-based nanocurcumins will givedetailed information on recent updates to the audience and be helpfulfor future developments.Curcumin has an outstandingsafety profile among all the nutraceuticalswith numerous pleiotropic biological activities such as anti-inflammatory,antioxidant, and anti-cancer effects. It is a readily available, low-pricedcompound that can cross the BBB and is helpful for neurodegenerativediseases. Its pharmacological properties are becoming more interestingrecently as the applications of curcumin are a fast-growing, improving,and escalating enterprise, as evidenced by the studies. Curcumin exhibitsa variety of pharmacological activities as evidenced by its uses inmany diseases like cancer, diabetes, wound healing, arthritis, Alzheimer’s,Parkinson, inflammation, angiogenesis, atherosclerosis, hypertension,etc. Curcumin is enriched with many valuable phytoconstituents, whichare responsible for its efficacy and are proven experimentally andclinically.


Financial barriers in substance use disorder service systems have limited the widespread adoption—i.e., provider-level reach—of evidence-based practices (EBPs) for youth substance use disorders. Reach is essential to maximizing the population-level impact of EBPs. One promising, but rarely studied, type of implementation strategy for overcoming barriers to EBP reach is financing strategies, which direct financial resources in various ways to support implementation. We evaluated financing strategies for the Adolescent Community Reinforcement Approach (A-CRA) EBP by comparing two US federal grant mechanisms, organization-focused and state-focused grants, on organization-level A-CRA reach outcomes.A-CRA implementation took place through organization-focused and state-focused grantee cohorts from 2006 to 2021. We used a quasi-experimental, mixed-method design to compare reach between treatment organizations funded by organization-focused versus state-focused grants . Using administrative training records, we calculated reach as the per-organization proportion of trained individuals who received certification in A-CRA clinical delivery and/or supervision by the end of grant funding. We tested differences in certification rate by grant type using multivariable linear regression models that controlled for key covariates , and tested threats to internal validity from our quasi-experimental design through a series of sensitivity analyses. We also drew on interviews and surveys collected from the treatment organizations and (when relevant) interviews with state administrators to identify factors that influenced reach.p = .01). Sensitivity analyses suggested these findings were not explained by confounding temporal trends nor by organizational or state characteristics. We did not identify significant quantitative moderators of reach outcomes, but qualitative findings suggested certain facilitating factors were more influential for organization-focused grants  and certain barrier factors were more impactful for state-focused grants .The overall certification rates were 27 percentage points lower in state-focused versus organization-focused grants (As the first published comparison of EBP reach outcomes between financing strategies, our findings can help guide state and federal policy related to financing strategies for implementing EBPs that reduce youth substance use. Future work should explore contextual conditions under which different financing strategies can support the widespread implementation of EBPs for substance use disorder treatment.The online version contains supplementary material available at 10.1186/s13012-023-01305-z. Contributions to the literatureOrganization-focused grants were robustly associated with higher average organization-level certification rates in the Adolescent Community Reinforcement Approach, revealing a notable strength of that financing strategy relative to state-focused grants.Qualitative findings revealed key differences in facilitators and barriers by grant type, which help identify conditions under which each grant type could be most useful for maximizing provider adoption of evidence-based practices.These findings offer evidence to inform state and federal investments into policy efforts to implement evidence-based practices.Research has identified several evidence-based practices (EBPs) for the treatment of adolescent and young adult substance use disorders  . One exeDespite the strong evidence for EBPs like A-CRA, low rates of SUD service delivery (7.6% ages 12–17 and 4.4% ages 18–25)  suggest A recent review highlighted the importance of financing strategies , a type Federal agencies have the influence and resources to potentially increase EBP delivery nationwide through financing strategies , 20. UnfThe present study evaluated how well the two aforementioned SAMHSA grant strategies promoted A-CRA provider-level reach, one of the several key outcomes for evaluating the impact of implementation strategies , 22. ReaWe describe this study using the Standards for Reporting Implementation Studies  see Add. RelevanThe current study used a longitudinal, mixed-method  design tGiven that the state-focused financing strategy was designed by SAMHSA to improve implementation outcomes beyond the organization-focused initiative, for research question 1, we tested SAMHSA’s hypothesis that reach would be higher among organizations participating in the state-focused grants relative to the organization-focused grants. Our design used a natural experiment, without random assignment of treatment organizations to the different grant types, and we prioritized producing externally valid evidence on the impact of each grant strategy in large-scale initiatives. This design does not fully isolate the effects of grant type on outcomes; however, we also tested the robustness of our findings against major internal validity threats with a series of sensitivity analyses.We hypothesized for research question 2 that A-CRA reach outcomes would be moderated by two key factors shown to influence implementation: , 29 exteThe project design and hypotheses are grounded in the Exploration, Preparation, Implementation, Sustainment (EPIS) framework , 29, whiThrough its grant-funded initiatives, SAMHSA provided funding, oversight, and leadership for A-CRA implementation to (i) four cohorts of organization-focused grantees across 26 states, awarded between 2006 and 2010, followed by (ii) four cohorts of state-focused grantees across 22 states, awarded between 2012 and 2017. During the preparation and implementation phases, organizations or states applied for, received, and executed SAMHSA grants. All grants targeted inner context  factors at treatment organizations through training and certification for A-CRA clinicians and supervisors; leadership from treatment organizations selected which providers were available and interested to participate in the training. State-focused grants further targeted outer context  factors at the state level and bridging factors that connected state substance use agencies and treatment organizations; with these changes, SAMHSA aimed to maximize the number of providers offering A-CRA in their states and, thus, enhance reach outcomes relative to organization-focused grants. Reach outcomes are the culmination of the implementation phase, i.e., grant funding period, after which the sustainment phase begins. Next, we describe the two grant types, and Table SAMHSA awarded ~ $900,000 USD (across a 3-year period) to each treatment organization. These grants supported A-CRA implementation by paying for A-CRA delivery, A-CRA supervision, and other related activities. Our prior work showed that grantees commonly experienced initial success in implementing A-CRA with fidelity and reducing youth substance use, but difficulty sustaining the model post-funding –19.SAMHSA awarded ~ $3–4 million USD each  to state agencies that administered publicly funded SUD services. One-third of the grant funds paid for states to develop EBP-focused infrastructure, such as funding and training. Some grants required that state agencies propose “demonstration site” organizations, which established partnerships with the state agency and implemented A-CRA first before the state began broad dissemination to other organizations.We used CHS certification records, conducted semi-structured interviews, and collected survey data from clinicians and supervisors at treatment organizations that implemented A-CRA (inner context). For interviews and surveys, we sampled organizations within the 5-year period following the completion of grant funding. Individuals who were currently or recently employed as a clinician or clinical supervisor responsible for youth SUD treatment were eligible to participate; those knowledgeable about A-CRA implementation were preferred.χ2(1) = 1.45, p = .23) or turnover rates (χ2(1) = 0.82, p = .36), even after replacement.For organization-focused grantees, all 82 eligible organizations (in 27 states) were invited for interviews. Organizations that participated in state-focused grants were more numerous, so we created a comparable sample of 82 organizations (in 18 states) by randomly selecting up to five organizations per state, including up to three “demonstration sites.” To ensure a comparable sample size for state-focused grants, we re-selected 17 organizations across nine states ; CHS records showed that selected and not-selected organizations did not differ on A-CRA certification rates (The interviewed sample included 154 organizations (94% of the 164 selected), with 249 provider participants . For CHS records, data from all trained providers  at the selected organizations were included. For 11 organizations that had providers trained under both grant types, we used the first (organization-focused) observation in our primary analyses, but we explored other ways of handling those organizations in sensitivity analyses (described later).We also interviewed administrators from the state agencies (outer context) that received SAMHSA state-focused grants. Participants were currently or recently employed in a relevant leadership position and were eligible within the 5-year period after funding ended. Thirty-two administrators from 100% of the 18 eligible states participated; we conducted group interviews when multiple individuals from the same state agency participated.Table CHS created a database of certification outcomes for all sampled treatment organizations, which specified any certifications achieved by each clinician or supervisor trained in A-CRA during the grant period as well as relevant descriptive information.Interview protocols used a combination of open-ended questions and standardized probes . ProtocoFollowing each provider interview, respondents were sent a web-based survey that collected standardized measures  and other descriptive information , and fidelity scores for rated sessions resulting in passing scores. It was important to verify that each certified individual achieved fidelity , since fidelity is associated with client SUD outcomes , 38–43. n = 138). We used the funding stability subscale to capture external financial support for A-CRA and the organizational capacity subscale for internal leadership support of A-CRA.We proposed two prespecified moderators of A-CRA reach outcomes , but theWe collected other interview and survey data that measured constructs from EPIS. Innovation measures captured participants’ perceptions of and attitudes toward the A-CRA model. Inner context measures described the treatment organizations delivering A-CRA, outer context measures described extra-organizational factors that affected A-CRA reach, and bridging factors described factors that linked inner and outer contexts (including grant-funded activities). We asked about the impacts of the ongoing COVID-19 pandemic in the state-focused sample, but those data were only applicable to reach outcomes in three states with active state-focused grants in 2020.To compare reach rates between grant types, we fit a series of multivariable linear regression models with reach rate as the organization-level dependent variables. Each regression included grant type and several covariates: turnover rate, number of individuals trained, number of supervisors who pursued certification, grant end date, and quarters of grant funding. These covariates helped address threats to interpreting the “grant type” variable  and account for important factors identified in our qualitative analyses . Moderator variables  were entered at the organization level, along with an interaction term with grant type. Moderator variables were averaged for all ratings at that organization; if moderator items were missing, we imputed their mean value from other organizations with the same A-CRA sustainment status.ns = 34 for first-level TAY, 29 for full TAY), per our initial power calculations [p < .05 for statistical significance rather than controlling the experiment-wide error rate.The initial examination of reach rates revealed that both TAY outcomes had sample sizes too small for an adequately powered analysis , but the findings changed minimally so we report the basic models. We also conducted a series of sensitivity analyses that characterized threats to internal validity in our design. Specifically, we examined (a) observed secular trends using a non-equivalent dependent variable , 45, delWe analyzed interviews using conventional content analysis  to identTo analyze the state-focused interview transcripts, we developed a codebook in Microsoft Excel with code definitions and exemplars , organizFinally, we developed written summaries of the content of codes (with exemplar quotes), focusing on implementation-specific barriers and facilitators. The lead author and second author  incorpoTable p = .01), and the rates of first-level certification were 23 percentage points higher (p = .04), compared to state-focused grants. These models explained a large portion of the variance in outcomes (R2 = .33 and .22). There were no significant differences in full or supervisor certification rates.Table The detailed results of the sensitivity analysis regression models are reported in Additional file p = .18), and organizational capacity, 0.00 (p = .91). Interaction effects with grant type were as follows: funding stability, 0.01 (p = .88), and organizational capacity, 0.07 (p = .07).Neither tested moderator showed significant effects for the sub-sample of organizations (84%) with survey responses. Coefficients for the main effects on any certification were as follows: funding stability, 0.03 , and when relevant, experience providing substance use treatment services ranged from 1 to 25 years .Table From our coding of those interviews, we summarized the key barriers and facilitators to A-CRA reach. Findings are presented by EPIS domain . The written summaries give an overview of all findings but emphasize the key differences between contextual influences on organization-focused and state-focused grants. Table Respondents described numerous innovation factors as strengths of A-CRA, such as its content and procedures, structured format, and evidence-based status, and the training and certification support offered by CHS. These facilitators were often endorsed regardless of an individual’s certification status, though some respondents who stopped using A-CRA described its being overly structured and inflexible as a barrier. Respondents also broadly described the A-CRA certification process as complex and burdensome (whether they completed it or not), which was a major barrier to reach. A key difference between grant types was that state-focused grantees reported more frequent and serious barriers around fit of A-CRA with their client population compared to organization-focused grantees; across grant types, A-CRA being a good fit for the youth and families served was considered an important facilitator. Overall, providers had more detailed responses about A-CRA characteristics, though state administrators reported similar perceptions based on their discussions with providers.Respondents across roles identified organizational characteristics that facilitated reach of A-CRA, including leadership, supervisors, and clinicians who supported the model; “champions” who helped provide resources and encouraged certification; and organizational policies that aligned with A-CRA’s use . Conversely, the lack of support from key individuals (often related to the negative perceptions described under the innovation domain) and misalignment with organizational policies  created barriers to A-CRA reach. Turnover of providers and leaders was another major barrier, especially when it disrupted support for A-CRA or required the replacement of certified providers. Finally, one prominent facilitator uniquely described by organization-focused grantees was an engagement in strategic planning to promote the adoption and use of A-CRA. State-focused grantees described minimal strategic planning around A-CRA, so this facilitator was rarely present for them.State administrators provided rich perspectives on outer context factors that influenced A-CRA reach, though providers commented on similar factors. Across grant types, states’ policy contexts influenced A-CRA reach; for example, policies that encouraged the provision of EBPs facilitated A-CRA use, whereas a common barrier was limited reimbursement for non-service components of A-CRA . Among state-focused grantees, some respondents described state substance use agency administrators as helpful in guiding the implementation process, which also encouraged organizational buy-in. However, state administrators also described challenges to fulfilling that role, with the specific challenges varying from state to state . Finally, some state-focused grantees also described attitudes among caregivers, community members, and government agencies that increasingly de-prioritized addressing youth substance use —which decreased youth referrals and engagement in A-CRA.The most salient bridging factor for many respondents was their SAMHSA funding. For state-focused grants, state-led training activities and comprehensive funding for A-CRA training, certification, supervision, and delivery were identified as major facilitators of reach, bridging between outer context (state-focused grant) and inner context (enactment of grant activities at treatment organizations) through statewide coordination of activities. However, providers noted that some state administrators did not encourage the use of A-CRA or help identify referral sources, creating barriers to reach. A few states were able to extend efforts beyond training and funding into policy development  and broader implementation supports ; such activities promoted reach but were rare and challenging to execute due to aforementioned state policy barriers. Organization-focused grants provided similar benefits as state-focused grants  through a more direct mechanism, albeit to individual organizations only. Organization-focused grantees’ experiences differed in that they worked with SAMHSA directly, a collaboration often described as helpful but also as intensive and overwhelming . Treatment organizations involved in state-focused grants typically had fewer requirements and less interaction with grant administrators (state or SAMHSA). Finally, providers considered their partnerships with other organizations for referrals and on-site service delivery  as important facilitators  across grant types.For the three states where the COVID-19 pandemic overlapped with their state-focused grant, participants described several impacts on A-CRA implementation. Effects on A-CRA delivery (innovation domain) were mixed; despite the benefit of increased use of telehealth as an accessible treatment modality, providers noted that some clients experienced connectivity and equipment issues, making it harder to engage them in A-CRA via telehealth. Respondents also described how the pandemic exacerbated existing barriers like staff turnover (inner context) and difficulties working with partners for A-CRA referrals (bridging factors).Effective financing strategies are needed to support EBP implementation in SUD services. We compared two financing strategies from the SAMHSA on the implementation outcome of A-CRA provider-level reach . The shift from organization-focused to state-focused grants was associated with a 27 percentage point decrease in provider certification rates at treatment organizations, counter to our hypothesis. This finding appeared to be driven by differences in required first-level certification, with rates of the optional full certification < 1/3 in both grant types. We used a rigorous non-experimental design, with extensive sensitivity analyses supporting our interpretation that grant type was the major contributor to variation in reach, and confounding temporal, state, or organizational factors did not explain the findings.Interviews with providers and state administrators suggested that achieving A-CRA certification was challenging and resource-intensive regardless of the type of financing strategy. Barriers and facilitators spanned all EPIS domains, as well as the construct of interactions and relationships during the implementation process , which has rarely been reported in studies using EPIS , 29. ImpWe designed this mixed-methods study to provide an in-depth understanding of financing strategies and identify the practical implications for policymakers. The findings contradicted our hypothesis, which makes it even more imperative to use this research evidence to inform EBP financing strategy design. Indeed, SAMHSA created the state-focused grant mechanisms in response to concerns about organization-focused grants, which in our earlier work –19 had dState-focused grants might have additional benefits not identified in our organization-level analysis. Reach at the state level  could be higher in state-focused grants, which trained a much larger number of organizations than received organization-focused grants. Another important future direction is our planned state-level case study analyses , 49, in It is also possible that state-focused grants produced higher A-CRA sustainment rates, as SAMHSA intended, among certified providers—though this seems unlikely given that successful initial implementation  provides the necessary foundation for later sustainment . RespondOur study builds on a small but emerging area of research on provider-level reach rates as an implementation outcome. Two previous studies examined provider reach for SUD-specific treatments—contingency management at an opioid treatment program  and alcoResearchers hoping to conceptualize their own studies of financing strategies in behavioral health, medicine, public health, and/or prevention may find it instructive to consider recently published recommendations from a policy adaptation of the EPIS framework . Those rDespite its strengths, this study had several limitations. First, this work cannot directly inform efforts to improve the feasibility of the A-CRA certification process , which This study is the first published comparison of reach outcomes between alternate financing strategies to support EBP implementation. This work fills a critical gap in how implementation research findings can guide administrators’ and policymakers’ investments in EBPs for youth SUD. Having identified critical issues in A-CRA reach and sustainment outcomes for state-focused and organization-focused grant financing strategies, we are poised to begin collaboratively exploring policy solutions. We hope that this work will encourage researchers, policymakers, and system/organization leaders to collaboratively study optimal ways to implement diverse EBPs in financially sustainable strategies, including the use of novel analytic approaches . Continued efforts will be essential to understand how these strategies operate with service and administrative systems and organizations to ultimately improve the effectiveness of providers and the lives of clients.Additional file 1. StaRI reporting checklist.Additional file 2. All Wave 1 Interview Guides.Additional file 3. All Wave 1 Survey Items.Additional file 4. Supplemental Analyses for Quantitative Analysis of Provider-Level Reach Outcomes.Additional file 5. Expanded Summary of All Barriers and Facilitators to A-CRA Provider-Level Reach Outcomes.


Vertical transmission of SARS-CoV-2 from mother to fetus is widely accepted. Whereas most infected neonates present with mild symptoms or are asymptomatic, respiratory distress syndrome (RDS) and abnormal lung images are significantly more frequent in COVID-19 positive neonates than in non-infected newborns. Fatality is rare and discordant meta-analyses of case reports and series relating perinatal maternal COVID-19 status to neonatal disease severity complicate their extrapolation as prognostic indicators. A larger database of detailed case reports from more extreme cases will be required to establish therapeutic guidelines and allow informed decision making. Here we report an unusual case of a 28 weeks' gestation infant with perinatally acquired SARS-CoV-2, who developed severe protracted respiratory failure. Despite intensive care from birth with first line anti-viral and anti-inflammatory therapy, respiratory failure persisted, and death ensued at 5 months. Lung histopathology showed severe diffuse bronchopneumonia, and heart and lung immunohistochemistry confirmed macrophage infiltration, platelet activation and neutrophil extracellular trap formation consistent with late multisystem inflammation. To our knowledge, this is the first report of SARS CoV-2 pulmonary hyperinflammation in a preterm newborn with fatal outcome. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of COVID-19 disease, has broad clinical manifestations in adults that range from mild respiratory symptoms to acute respiratory failure. Maternal COVID-19 infection is associated with increased maternal death and serious morbidity –9. WhileIn adult patients, hyperinflammation contributes to the severity and mortality of COVID-19. Hyperinflammation is characterized by NF-kB activation and elevated systemic inflammatory markers, including cytokines IL-1β, IL-6, IL-8, IL-10, and TNF-α, mononuclear infiltrate with dysregulated macrophage activation and neutrophil activation with formation of neutrophil extracellular traps (NETs) that lead to further lung injury and microthrombi . Early cRoles for maternal age and COVID-19 severity in disease presentation and outcome of SARS-CoV-2 infected neonates is hotly disputed , 32, 33.A premature male was born at 28 weeks and five days gestation with a birth weight of 1,200 grams due to preterm labor. This was a dichorionic-diamniotic twin pregnancy. The co-twin was selectively terminated at an outside hospital at 21 weeks' gestation for multiple congenital anomalies secondary to amniotic band syndrome. The mother, a 23-year-old Hispanic with no reported history of serious disease tested positive for SARS-CoV-2 by polymerase chain reaction (PCR) tests 13 days before delivery and upon admission to the hospital. The mother was asymptomatic at presentation and reported no previous symptoms of COVID-19. She received antenatal steroids 29 days prior to delivery after she presented to the hospital with suspected rupture of membranes. Placental pathology of the co-twin showed acute chorioamnionitis with areas of fibrin deposition characteristic of intra-amniotic infection of an unidentified source.2O. The infant was admitted to the neonatal intensive care unit (NICU) on CPAP and FiO2 0.21. Chest x-ray showed mild diffuse hazy granular opacities consistent with respiratory distress syndrome  of 5cmHsyndrome . A nasop2 0.5) prompted treatment with dexamethasone  and remdesivir . Chest x-ray at this point showed diffuse bilateral hazy opacities with increased perihilar markings  and phospo-NF-kB-p65 expression in lung macrophages and cardiomyocytes, increased osteopontin (OPN) and intracellular adhesion molecule 1 (ICAM-1) in blood vessels in the lung and heart , and incIn this case study, vertical transmission of SARS CoV-2 is indicated by the presence of maternal infection during the third trimester and at birth, positive nasopharyngeal tests of the neonate at 24 h and 26 days following immediate separation from the mother to an NICU. The case timeline is outlined in Our first line use of dexamethasone, remdesivir, and anakinra is supported by previous successes in hospitalized COVID-19 positive adults , 38, preElevated inflammation markers at birth and chest x-rays depicting progressive bilateral interstitial infiltrates paralleled by the appearance of anti-SARS CoV-2 antibodies and exacerbated RDS are consistent with prenatal exposure to inflammation , ongoingThis is the first report to describe SARS CoV-2 pulmonary hyperinflammation in a preterm newborn with fatal outcome. Lung pathology revealed severe bronchopneumonia with obliteration of airspaces by inflammatory cells without evidence of active infection at death. Immunohistochemical analysis of the infant's lungs and heart revealed diffuse expression of LDLR, NF-kb, and OPN, suggesting hyperinflammation. Furthermore, expression of ICAM-1 suggested vascular dysfunction , and citThe unusual etiology of our case does not conform precisely with either MIS-N or PIMS-TS where intravenous dexamethasone with or without IgG are indicated , 57. Whe


Panel B). Although the PFA applications were intentionally delivered for PVI, the finding of extended PW lesions may be explained by the larger sized PFA catheter, the relatively shorter posterior inter-venal distance , but clearly also by the unique property of PF technology to obtain tissue lesions not necessarily through high catheter contact, but also in the proximity of the tissue–electrodes interface.We present the case of a 55-year-old patient with paroxysmal atrial fibrillation referred for pulmonary vein isolation (PVI) by means of pulsed-field ablation (PFA). Based on preprocedural imaging (computed tomography) showing a bilateral common ostium (CO) as PV variant, the larger penta-spline PFA catheter measuring 35 mm was selected for ablation. Bilaterally, acute PVI was observed during the first applications at the CO wiring the superior branches. Overall, eight application pairs were delivered with four pairs per CO . Pre- and post-ablation 3D ultra-high-density voltage map of the left atrium was acquired with a 64-pole basket mapping catheter (Orion). The post-ablation map confirmed acute PVI, but showed unexpected fused lesions on the lower posterior wall (PW) in addition (https://www.escardio.org/Education/E-Learning/Clinical-cases/Electrophysiology.The full-length version of this report can be viewed at:


Breast cancer is a highly heterogeneous disease that comprises multiple biological components. Owing its diversity, patients have different prognostic outcomes; hence, early diagnosis and accurate subtype prediction are critical for treatment. Standardized breast cancer subtyping systems, mainly based on single-omics datasets, have been developed to ensure proper treatment in a systematic manner. Recently, multi-omics data integration has attracted attention to provide a comprehensive view of patients but poses a challenge due to the high dimensionality. In recent years, deep learning-based approaches have been proposed, but they still present several limitations.In this study, we describe moBRCA-net, an interpretable deep learning-based breast cancer subtype classification framework that uses multi-omics datasets. Three omics datasets comprising gene expression, DNA methylation and microRNA expression data were integrated while considering the biological relationships among them, and a self-attention module was applied to each omics dataset to capture the relative importance of each feature. The features were then transformed to new representations considering the respective learned importance, allowing moBRCA-net to predict the subtype.https://github.com/cbi-bioinfo/moBRCA-net.Experimental results confirmed that moBRCA-net has a significantly enhanced performance compared with other methods, and the effectiveness of multi-omics integration and omics-level attention were identified. moBRCA-net is publicly available at The online version contains supplementary material available at 10.1186/s12859-023-05273-5. Breast cancer is one of the most frequently diagnosed cancers and is the second leading cause of death among women worldwide . Breast In recent years, several classification frameworks to predict the molecular subtypes of breast cancer have been reported. For example, Rhee et al. presented a hybrid approach integrating graph convolutional network and relation network using gene expression profiles and protein–protein interaction networks . Gao et However, classification approaches based solely on single-omics datasets only provide information on a single molecular level. These approaches fail to provide a comprehensive view of the biological processes involved in breast cancer and cannot capture correlations between molecules from different layers . To overRecently, deep learning-based multi-omics data integration has shown robust performance in several applications, including patient clustering, survival analysis , and bioIn the present study, we propose moBRCA-net, an omics-level attention-based breast cancer subtype classification framework that uses multi-omics datasets. Dataset integration was performed based on feature-selection modules that consider the biological relationship between the omics datasets . Moreover, for omics-level feature importance learning, a self-attention module was applied for each omics feature, and each feature was then transformed to the new representation incorporating its relative importance for the classification task. The representation of each omics dataset was concatenated and delivered to the fully connected layers to predict the breast cancer subtype of each patient.Our moBRCA-net framework consists of four modules: preprocessing, multi-omics data integration, omics-level feature importance learning, and classification. The workflow of our model is shown in Fig. The breast cancer (BRCA) cohort datasets were collected from The Cancer Genome Atlas (TCGA) . Three tFor gene expression and microRNA expression data, we first removed genes and microRNAs for which read counts were not available for all samples. After calculating size factors, the read counts were normalized by library size and were log-transformed using DESeq2 . For DNAp-value less than 0.01 was considered as differentially expressed genes (DEGs). We constructed the gene set composed of 1000 DEGs with the highest log(fold change) values and the lowest adjusted p-value.To prevent a possibility of overfitting and expensive computational costs to train the neural network model, we constructed a feature selection module to integrate the multi-omics datasets. First, the genes remaining after preprocessing were evaluated concerning their informativity as a breast cancer signature by performing differential analysis that compared them to matched normal samples using DESeq2. Genes with an absolute value of log (fold change) greater than 2 and an adjusted To integrate the multiple omics datasets based on their biological relationship, we identified the other omics features related to the selected DEGs. Studies have shown that promoter-associated CpGs play important roles in gene silencing, genomic imprinting, and cancerigenesis –30. Aften represents the dimension of the input data, we defined the k-dimensional embedding vector x was transformed to a new representation b is a bias term. To learn the features playing a key role for classifying the breast cancer subtypes and better understand the relative importance of those features, we implemented the omics-level self-attention module into our model. Given a set of original input data  follows :1\documeC represents the number of breast cancer subtypes, and y -based classifiers: Support vector machine (SVM) , Random Overall, moBRCA-net (denoted as ’moBRCA-net (omics-attn)’) significantly outperformed the other ML-based classifiers with an average accuracy of 0.891, F1-score of 0.887, and MCC of 0.831. SVM showed the second-best performance with 0.866, 0.861, and 0.808 for the average accuracy, F1-score, and MCC respectively -based data generation for certain minority classes in the imbalanced dataset improved the classification performance in various domain tasks such as respiratory disease classification , temporaNext, we investigated the performance improvement of moBRCA-net by the introduction of the omics-level attention modules for feature importance learning. We implemented two variants of moBRCA-net, in one a single-attention module was applied to all features at once, and in the other the attention module was removed to directly classify the breast cancer subtypes, which were denoted as ’moBRCA-net (single-attn)’ and ’moBRCA-net (no-attn)’, respectively. tenfold cross-validation was performed to measure performance. Without the attention module, the classification performance was comparable to that of the SVM, showing an average F1-score of 0.864   and RF. Furthermore, we investigated the impact of the CpG clusters by comparing the classification performance of moBRCA-net based on single CpG-based multi-omics integration using different feature selection methods Fig. . InteresTo validate whether utilizing multi-omics datasets could effectively improve the classification of breast cancer subtypes, we compared the performance of moBRCA-net when using different combinations of multi-omics datasets and a single-omics dataset. We denoted gene expression as ’gene’, DNA methylation as ’methyl’, and microRNA as ’microRNA’. In this experiment, moBRCA-net showed a relatively higher performance when trained based on multi-omics compared with single-omics data Table . Multi-op-value To understand how omics-level attention helped improve the performance of the model to accurately classify breast cancer subtypes, we interpreted the attention scores of moBRCA-net. First, to directly compare the abundance difference between the feature subtypes with the highest attention scores, we visualized the normalized gene expression values and beta values of those features obtained from samples of each breast cancer subtype. One-way analysis of variance  was perfStudies have shown that DNA methylation regulates gene expression in breast cancer, with hypermethylation leading to the silencing of regulatory genes, whereas hypomethylation activates genes required for metastasis, consequently resulting in uncontrolled growth and cancer progression . DNA met subtype . STAT5 e subtype . Indeed, subtype . Furthermentclass2pt{minimIn addition, we used the Molecular Signature Database (MSigDB) , 53 collIn this study, we describe moBRCA-net, a self-attention-based breast cancer subtype classification framework based on multi-omics integration. Gene expression, DNA methylation, and microRNA expression were integrated considering their biological relationships, and an omics-level attention module was implemented to identify and learn the important features critical for cancer classification. The model was then evaluated and compared with ML-based established methods. Overall, moBRCA-net outperformed the previously reported models in breast cancer subtype prediction. Moreover, we investigated the effectiveness of each module in moBRCA-net for classification improvement. Attention scores were interpreted to validate whether our model could concentrate on features highly relevant to cancer subtype classification.The major contribution of our study resides within the multi-omics data integration strategy. To maintain the biological relationship between the multi omics features while integration, feature selection module was constructed to identify the informative breast cancer signature genes and the relation between the identified genes and other two omics features were built based on the promoter and the target relationship. Moreover, we employed the omics-level attention module to first train the classifier to capture the relative importance of each feature for each omics data. Features were transformed to new representations considering the respective learned importance. These lead the breast cancer subtype classifier not to ignore the distribution difference of each omics dataset and to integrate multi-omics more efficiently by reducing the dimensionality of each omics. This is also supported by the experiments performed in our study Fig. ” SectionmoBRCA-net showed a high prediction performance, but there are still some bottlenecks that could be further improved. For example, due to the limitation of the computational resources, feature selection was performed to reduce the number of features for training our model. Also, if the model could learn the dependency between the omics features directly via graph network, it could potentially be able to extract useful relations between the features of different omics datasets. In the future, we plan to extend our moBRCA-net platform to utilize graph neural networks.Additional file 1: S1: Table 1. Hyperparmeter optimization results based on the average accuracy for classifying breast cancer subtypes. Table 2. Classification accuracy results with the different number of selected features in multi-omics data integration. S2: Optimization results of the baseline methods with the different combination of the parameters. S3: Accuracy and weighted F1-score results of moBRCA-net, its variants, and the machine learning-based classifiers from the performance evaluation based on 10-fold cross validation in Fig. S4: Average subtype-wise accuracy and weighted F1-score results of moBRCA-net (omics-attn) from the performance evaluation based on 10-fold cross validation in Fig. S5: Average overall and subtype-wise performance results of moBRCA-net with/without data augmentation based on 10-fold cross validation. S6: Accuracy and weighted F1-score results of moBRCA-net using different feature selection methods in the multiomics data integration based on 10-fold cross validation in Fig.  S7: Normalized values of features showing top 3 to 5 average attention scores from each omics. S8: Top 200 features showing the highest average attention scores across patients for each omics dataset. S9: 53 overlapped genes with the gene set collection from MSigDB and PAM50, amont the top 200 genes showing the highest avearge attention scores across patients.


Cellvibrio japonicus and found that it has sophisticated regulation that includes substrate sensing and growth rate components for genes that encode starch-degrading enzymes. Among the 22 genes predicted to encode starch-active enzymes, C. japonicus has 10 alpha-amylases, 4 alpha-glucosidases, 2 pullulnases, and 2 cyclomaltodextrin glucanotransferases, 15 of which were up-regulated during exponential growth on starch and 8 up-regulated in stationary phase. Growth analyses with an enzyme secretion deficient mutant of C. japonicus suggested that secreted amylases are essential for this bacterium to degrade starch. Our approach of coupling a physiological growth assay with transcriptomic data provides a platform to identify targets for further genetic or biochemical analysis that can be broadly applied to other starch-utilizing bacteria.Bacterial utilization of starch is increasingly of interest as the importance and contributions of animal gut microbiomes become more defined. Consequently, identifying and characterizing the bacterial enzymes responsible for the degradation, transport, and metabolism of starch will enable developments in pharmaceutical, biotechnological, and culinary industries searching for novel prebiotics, carrier molecules, and low glycemic index sweeteners. The current challenge is that bacteria proficient at starch utilization often have hundreds of carbohydrate active enzymes, and it is unclear which are essential for starch utilization using only homology-based bioinformatics or computational methods. Complementary experimental data are also needed, especially to understand the regulation of bacterial starch utilization. We have completed an RNAseq analysis of the Gram-negative bacterium C. japonicus regulates carbohydrate active enzymes, or how this bacterium compares to gut microbiome constituents in terms of starch utilization potential.Understanding the bacterial metabolism of starch is important as this polysaccharide is a ubiquitous ingredient in foods, supplements, and medicines, all of which influence gut microbiome composition and health. Our RNAseq and growth data set provides a valuable resource to those who want to better understand the regulation of starch utilization in Gram-negative bacteria. These data are also useful as they provide an example of how to approach studying a starch-utilizing bacterium that has many putative amylases by coupling transcriptomic data with growth assays to overcome the potential challenges of functional redundancy. The RNAseq data can also be used as a part of larger meta-analyses to compare how  Cellvibrio japonicus has 17 predicted genes that encode GH13 family enzymes  and Klebsiella oxytoca (12 genes)  Carbohydrate Active enZymes (CAZymes), can degrade the amylose and amylopectin components of starch . These e enzymes . With th2 genes) , 7. To caomicron  genes anucosides , we perfC. japonicus strains in MOPS-defined media  with glucose, maltose, or starch as the sole carbon source (gsp), which has been previously shown to be essential for secreting CAZymes in C. japonicus  as the experimental substrate because it is comprised of α−1,4 glycosidic bonds , which is a native substrate for GH13 CAZymes. We grew n source  prior toaponicus . This stlization . For thi maltose –15. On16-100 asC. japonicus grown using glucose (GSE90955) were used for comparative analysis  up-regulated (fold change >2.0) that could be broadly grouped by gene ontology (GO)   as havinogy (GO) , which iere used , 15, 17.C. japonicus cells versus glucose-grown cells during stationary phase, 289 genes were significantly up-regulated and could be grouped by the GO terms carbohydrate metabolic processes (GO:0005975), signal transduction (GO:0007165), chemotaxis (GO:0006935), and phosphorelay signal transduction (0000160). Among the up-regulated genes were 31 CAZyme-encoding genes, including 3 predicted α-amylase-encoding genes, 3 alpha-glucosidase-encoding genes, 1 pullulnase-encoding gene, and 1 cyclomaltodextrin glucanotransferase-encoding gene. Overall, the CAZyme-encoding genes up-regulated during stationary phase were previously shown to encode proteins possessing a wide array of carbohydrate-degrading activities , bacterial-type flagellum assembly (GO:0044780), organization (GO:0044781), and motility (GO:0071973), signal transduction (GO:0000160 and GO:0007165), transcription regulation (GO:0006355), and carbohydrate metabolism (GO:0005975). There was a total of 15 CAZyme-encoding genes that were up-regulated, which included only two predicted α-amylase-encoding genes but no alpha-glucosidase-encoding genes, pullulnase-encoding genes, or cyclomaltodextrin glucanotransferase-encoding genes. As only two amylase genes  were regulated (but not exclusively) by growth rate, we hypothesize that the major regulatory mechanism that C. japonicus employs to control starch-degradative enzymes is through substrate detection. This mechanism is markedly different than some Gram-positive bacteria, which have been shown to exclusively control amylase-encoding genes, and more generally those involved in carbohydrate degradation, via growth rate or catabolite repression (Comparing gene expression analysis of starch-grown pression –23.C. japonicus during starch utilization that can serve as resource for future meta-analyses of polysaccharide degradation and metabolism in bacteria. Given the problems associated with defining physiological function based solely on bioinformatic predictions (C. japonicus and related bacteria due to the abundance of genes that encode carbohydrate active enzymes.We have reported here an RNAseq dataset for the Gram-negative saprophyte dictions , integra


This cohort study uses hospitalization and 30-day mortality risks to create a temporal profile of the severity of COVID-19 in Massachusetts from July 2021 to December 2022. In this retrospective cohort study, while accounting for a comprehensive set of confounders including demographic characteristics, immunity, therapeutics, and comorbidities, we constructed a temporal profile of SARS-CoV-2’s severity in Massachusetts, measured through hospitalization and 30-day mortality risks, and hypothesized that severity was diminishing.Recent SARS-CoV-2 variants have been associated with lower severity.STROBE reporting guideline.We constructed an incident-level database of SARS-CoV-2 cases recorded within Mass General Brigham between July 1, 2021, and December 31, 2022. The use of data was approved by the Mass General Brigham institutional review board, with a waiver of informed consent for the use of deidentified data. Our study followed the 6 Cases were categorized based on the infection month, with November 2021 as the comparison benchmark. Covariates including the patient demographic characteristics, therapeutics, comorbidity score, vaccination status, and previous SARS-CoV-2 infection were adjusted using entropy balancing to reduce confounding bias. Outcomes were hospitalization and in-hospital death within 30 days of a positive SARS-CoV-2 test result. We computed weighted odds ratios and 95% CIs between November 2021 and other months with a survey-weighted logistic regression model. Further, we stratified our cases by vaccination status (unvaccinated vs ≥2 vaccines) and calculated similar treatment outcomes between unvaccinated individuals in November 2021 and vaccinated and unvaccinated individuals in all other months. Last, we calculated the adjusted absolute risk difference and ratio of hospitalization and mortality between the vaccinated and unvaccinated groups , 19% were unvaccinated . By selfThis cohort study shows that SARS-CoV-2’s severity waned between July 2021 and December 2022 after adjusting for confounders regardless of vaccination status. Overall, vaccination is still associated with a decrease in risk of severe SARS-CoV-2 outcomes. Use of entropy balancing provides an advantage over standard matching, as unmatched individuals are not discarded, which increases effect estimates’ precision and findings’ generalizability. With the SARS-CoV-2 variants demonstrating milder severity over recent months, it is reasonable to presume that our findings are broadly generalizable. Our models can be easily deployed across varying locations to assess temporal severity of SARS-CoV-2.Study limitations include vaccinated and unvaccinated populations not being balanced when examining severity over time, not accounting for time from vaccination, unobserved SARS-CoV-2 infections not detected by a positive test result or electronic health record flag, and unobserved vaccination at another facility. Mortality is underestimated by only including in-hospital deaths within 30 days; however, this is evenly distributed across months. Other unmeasured confounders may remain.


Recently, a great effort has been devoted to studying attenuated and subunit vaccine development against SARS-CoV-2 since its outbreak in December 2019. It is known that diverse virus-like particles (VLPs) are extensively employed as carriers to display various antigenic and immunostimulatory cargo modules for vaccine development. Single or multiple antigens or antigenic domains such as the spike or nucleocapsid protein or their variants from SARS-CoV-2 could also be incorporated into VLPs via either a genetic or chemical display approach. Such antigen display platforms would help screen safer and more effective vaccine candidates capable of generating a strong immune response with or without adjuvant. This review aims to provide valuable insights for the future development of SARS-CoV-2 VLP vaccines by summarizing the latest updates and perspectives on the vaccine development of VLP platforms for genetic and chemical displaying antigens from SARS-CoV-2. Th. ThStrepS. pneumoniae), was introduced. This system has the ability to combine with the SpyCatcher-SpyTag and generate multiple components of protein fusions across various protein production platforms, encompassing prokaryotic and eukaryotic cells, plants, and cell-free systems [The versatility of the SpyTag/SpyCatcher plug-and-display technology is not exclusively reliant on antigen display, but has also been observed in the investigation of antigen stability. Sharma et al.  demonstr systems ,75,76,77 systems . Adding  systems . Unfortu systems ,81,82.Helicobacter pylori (H. pylori) non-haem 24-mer ferritin to the receptor binding domain (RBD) and/or heptad repeat (HR) subunits of the SARS-CoV-2 spike (S) protein. Nanoparticle vaccines elicited stronger neutralizing antibodies and cellular immune responses than monomer vaccines. RBD and RBD-HR nanoparticle-vaccinated hACE2 transgenic mice with RBD and/or RBD-HR nanoparticles had lower viral loads in their lungs after the SARS-CoV-2 challenge. Neutralizing antibodies and cellular immune responses against other coronaviruses were also stimulated with RBD-HR nanoparticle vaccines. The lumazine synthase (LuS) scaffold protein from Aquifex aeolicus, which is fused with N-linked glycan and a Spy tag at the N-terminus (SpyTag-LUS/N-linked glycan), can give a solution to a problem caused by inefficiencies in nanoparticle vaccine production [Streptococcus dysgalactiae) and Gram-negative  bacteria. This system demonstrated enhanced productivity and efficacy, as well as eliciting stronger immune reactions in mice when compared to the original SpyTag/SpyCatcher system. The engineered protein modification for self-assembly may differ between envelope, non-envelope, and scaffold proteins. A study from Ma et al.  demonstroduction . The Spyoduction . The fusoduction ,86,87,88oduction . In theioduction . Followioduction ,90,91. Goduction ,86. A production  devised As of 30 March 2023, the World Health Organization (WHO) reported that there are 184 vaccines currently undergoing clinical development, with approximately 199 vaccines in the preclinical phase, in order to support global vaccine development endeavors. Additionally, the list of platforms employed for vaccine development efforts includes seven VLP-based vaccines . OngoingCurrently, it is widely acknowledged that the production and commercialization of VLP vaccines encounter many technical and analytical obstacles, even though the routine processes and strict industry regulations are followed. The separation and purification of artificially generated extracellular vesicles and VLPs from diverse bioreactors remain particularly challenging. To enhance the purity and efficacy of the desired VLP products, additional investigations are required to explore applicable procedures for eliminating extracellular vesicles and viruses (such as baculoviruses or lentiviruses) generated from utilized bioreactors. For instance, the utilization of fluorescent labelling and fluorescence-activated cell sorting (FACS) techniques can prove advantageous in the monitoring and optimization of both upstream and downstream processes involved in the production of virus-like particles (VLPs) ,94,95.


Virtual-assisted lung mapping (VAL-MAP) is a preoperative bronchoscopic multispot dye-marking technique that has demonstrated efficacy and good outcomes over the last 10 years. However, conventional VAL-MAP has certain limitations. Several new techniques have been developed to address the problems associated with conventional VAL-MAP and have shown good results. Here, we summarize not only the history and outcomes of VAL-MAP, but also the latest update of this preoperative marking technique using bronchoscopy.Virtual-assisted lung mapping (VAL-MAP) is a preoperative bronchoscopic multispot dye-marking technique used in sublobar lung resection of barely palpable lung nodules. This review summarizes the history and outcomes of the VAL-MAP procedure. VAL-MAP was developed in 2012, and long-term outcomes of lung resection using VAL-MAP have recently been verified. Problems associated with conventional VAL-MAP include a prerequisite of post-mapping computed tomography (CT), occasional inability to see dye marks during surgery, and infrequent resection failure due to deep resection margins; various techniques have been developed to address these issues. VAL-MAP using electromagnetic navigation bronchoscopy with on-site adjustment can omit post-mapping CT. The use of indocyanine green in VAL-MAP has increased the success rate of marking detection during surgery without causing additional complications. VAL-MAP 2.0—a three-dimensional mapping technique that involves the intrabronchial placement of a microcoil—has increased the accuracy of sublobar resection, particularly for deeply located tumors. Although these promising new techniques have some limitations, they are beneficial for sublobar lung resection. Lung cancer is a leading cause of cancer-related death, accounting for nearly 2.2 million cases in 2020 . The devWhile the importance of sublobar resection for small lung cancer, which is often detected via a low-dose CT scan, it has been notified recently that local recurrence has been a significant issue to overcome. The JCOG0802/WJCOG4607L trial showed that significantly more locoregional relapses occurred in patients who had segmentectomy (11%) than in those who had lobectomy (5%). The CALGB 140,503 trial also showed that locoregional recurrence was significantly higher in sublobar resections (13.4%) than in lobectomies (10.0%). Furthermore, the development of minimally invasive surgeries, such as video-assisted thoracic surgery and robot-assisted thoracic surgery, has made it more challenging to palpate and detect small pulmonary nodules intraoperatively . These rVirtual-assisted lung mapping (VAL-MAP) is a preoperative bronchoscopic multispot dye-marking technique used to identify barely identifiable pulmonary nodules and draw appropriate resection lines in sublobar lung resection . This unThe steps of VAL-MAP 1.0 are summarized in Virtual-assisted lung mapping is designed using the three-dimensional reconstruction of computed tomography (CT) data and virtual bronchoscopic images. Bronchoscopic dye injection is performed on the day of surgery or 1 day before surgery via a metal-tipped catheter preloaded with 1 mL of indigo carmine (black arrow). Post-mapping CT images are taken and developed into three-dimensional images for surgery to confirm the actual locations of the marks (arrowhead) and the tumor. Finally, the operation is conducted with the help of dye marks, which can be readily recognized as blue spots on the surface of the lung (white arrow). This figure is reprinted from the reference .There are many different localization methods for hardly palpable lung nodules, and two major image-guided localization techniques are CT-guided localization and bronchoscopic marking. CT-guided localization techniques are widely used for preoperative markings with various localized materials, including hook-wires, microcoils, fiducial markers, dyes, and radiotracers. The advantages and disadvantages of each material are shown in The advantages and disadvantages of bronchoscopic marking techniques are also shown in p = 0.071) } based on CT. The study demonstrated that the risk of resection failure reached 10% when the resection line was deeper than 30 mm from the lung surface. To overcome the problems arising from VAL-MAP 1.0, several new techniques have been developed.Several problems with VAL-MAP 1.0 have been reported . First,  = 0.02) . This stn margin . The reqThe steps of ENB VAL-MAP 1.0 are summarized in Moreover, on-site adjustment with ENB VAL-MAP eliminates the post-mapping CT scan requirement . On-siteRecently, we have shown that successful sublobar lung resection using VAL-MAP requires the creation of “three” intraoperative visible marks per established lung nodule . Two hunIn contrast, an increase in the number of VAL-MAP marks may cause bronchoscopic complications more often, leading to patient burden. Thus, strategies that can improve the success rate of marking detection during the surgery are indispensable. One of such strategies is the use of indocyanine green (ICG). ICG is a widely used fluorescent dye that is visualized with a near-infrared (NIR) thoracoscope . The vivAlthough VAL-MAP using ICG is likely to be quite useful and safe in increasing the detectability of markings, the use of ICG in VAL-MAP has certain limitations. One is that VAL-MAP using ICG is available only in institutions where an NIR scope is installed. Moreover, ICG is not applicable in patients with iodine hypersensitivity.The required resection margin was an independent risk factor for the successful resection of the targeted nodules in a multicenter prospective study . Since VVAL-MAP has demonstrated efficacy and good outcomes over the last 10 years, and several new techniques have been developed to address the problems associated with conventional VAL-MAP. They are promising and beneficial for sublobar lung resection.