Raman spectroscopic analysis is a promising and powerful optical investigative tool to interrogate tissue and/or biofluids for in vitro and in vivo applications. Fast-acquisition Raman imaging techniques support real-time, objective evaluation of molecular tissue composition in prostate cancer. Within the capability of non-destructive and non-invasive imaging, Raman spectroscopy-based techniques offer a reliable tool for cancer detection. There is a push for intraoperative use and/or multimodal imaging approaches that utilise the strength of each spectroscopic modality to improve prostate cancer detection and treatment.Optical techniques are widely used tools in the visualisation of biological species within complex matrices, including biopsies, tissue resections and biofluids. Raman spectroscopy is an emerging analytical approach that probes the molecular signature of endogenous cellular biomolecules under biocompatible conditions with high spatial resolution. Applications of Raman spectroscopy in prostate cancer include biopsy analysis, assessment of surgical margins and monitoring of treatment efficacy. The advent of advanced Raman imaging techniques, such as stimulated Raman scattering, is creating opportunities for real-time in situ evaluation of prostate cancer. This review provides a focus on the recent preclinical and clinical achievements in implementing Raman-based techniques, highlighting remaining challenges for clinical applications. The research and clinical results achieved through in vivo and ex vivo Raman spectroscopy illustrate areas where these evolving technologies can be best translated into clinical practice. Prostate cancer is the second most frequent malignancy in men and the fifth leading cause of death worldwide . The pri−1). Alternatively, the light may be scattered at a range of different wavelengths, and hence frequencies, in a process referred to as Raman scattering (or inelastic scattering). Raman scattering is a rare phenomenon, with only ~1 in 108 incident photons undergoing inelastic scattering. The inelastic scattered photons will have either higher energy (anti-Stokes Raman scattering) or lower energy (Stokes Raman scattering) due to the interaction of incident photons with vibrational modes associated with chemical bonds within the target sample. The energy difference between the incident photons and the Raman scattered photons characterises the chemical bonds excited by that energy and is measurable as a difference in wavelength of light compared to the excitation wavelength. As such, a Raman spectrum is a plot of the photon intensity in arbitrary units (a.u. or arb. units) as a function of Raman shift, which is plotted in wavenumber, cm−1 (1/wavelength). A Raman spectrum, therefore, characterises the various vibrational modes associated with chemical bonds that comprise the sample. The intensity of vibrations produced in Raman spectroscopy is dependent on the Raman cross-section of the molecule being interrogated. Water has a low Raman cross-section, meaning that aqueous samples are ideal for analysis by Raman spectroscopy due to the low background from the water; however, sensitivity can be an issue and is discussed later in this review.Raman spectroscopy is a powerful analytical methodology that can characterise biological specimens at high spatial resolution in a non-invasive manner. To do so, Raman spectroscopy uses monochromatic light, typically generated from a laser source in the mid-ultraviolet , visible (400–700 nm) or near-infrared  regions. Upon excitation of the sample by an incident laser, two scattering phenomena can occur: Rayleigh scattering (elastic scattering) and Raman scattering (inelastic scattering) A. Of theCompensating for the low probability associated with Raman scattering, high-efficiency laser sources, high-throughput optics and low-noise detectors have enabled Raman spectroscopy to be developed into a powerful imaging technique capable of reporting on the underlying biology of the sample. Interfacing Raman spectroscopy with an optical microscope has enabled the development of Raman imaging using high-magnification lenses to deliver subcellular spatial resolution B. The sp4. In practice, a laser with a short wavelength will have a greater intensity of Raman scattering and hence sensitivity than a laser with a long wavelength. However, the lower photon energy associated with the increasing laser wavelength can be advantageous for biological imaging, due to reduced sample phototoxicity which can result in sample overheating and burning. In addition, background absorption and fluorescence from tissue samples are generally reduced when NIR lasers are used for imaging, which reduces the background autofluorescence from the cell/tissue in the resultant spectra and images. The use of pulsed laser sources can also promote a similar effect. For most biological applications, the laser power focused onto the sample is typically <50 milliwatts (mW) for mid-UV and visible laser sources and <200 mW for NIR laser sources.The sensitivity of Raman scattering is dependent on the excitation wavelength, where the Raman scattering intensity is proportional to 1/λ−1) associated with proteins, lipids, DNA and carbohydrates . WSORS makes use of an applied spatial offset between the points of excitation and collection in a Raman measurement to collect photons that have been scattered by the subsurface medium. Comparisons between the spectra collected with no offset (surface measurements) and those collected with a lateral offset (subsurface measurements) enable the delineation of the spectral differences in composition at depth A 15]. S. S15]. S3 vibration signifying the presence of proteins (those containing branches amino acids) within live cells , where the frequency difference between the two beams is matched to a vibration of interest in the sample. The pump beam is usually a tuneable laser , whilst the Stokes beam is a fixed-wavelength laser . Retuning the pump laser to a different wavelength enables the detection of a different vibrational mode within the sample . Two prove cells . Retunin species . The res−1 of the Raman spectrum (highlighted as the peak for amide-I (−1) and CH2 deformation (1296 and 1444 cm−1), respectively . I. I−1 of ectively . Sample ectively . A set oIn order to perform optimal SERS measurements, consideration must be made with respect to sample preparation and nanoparticle composition. Silver colloids are commonly used nanostructures. The Leopold and Lendl method is typically used to produce silver salts using a reducing agent such as hydroxylamine hydrochloride and sodium hydroxide  . AlternaThe generated Raman spectrum includes peaks from all the Raman-active species within the sample, and therefore the technique can rapidly generate a large and information-rich dataset. When interfaced with a microscope system, the subsequent Raman images contain a full or partial Raman spectrum at each pixel within the image. As such, the extraction of chemical information may require multivariate analysis techniques to extricate the underlying chemical, biological or structural information. The analysis of subtle changes within the spectral profiles may be associated with disease, biological processes or the direct influence of external agents, for example, drugs. Salient techniques used in post-processing of Raman spectra include K-Means clustering analysis (KMCA), principal component analysis (PCA) and partial least squares regression (PLSR). In brief, KMCA aims to partition data into clusters based on the similarity of spectral features , whilst Raman spectroscopy has been evaluated as a diagnostic tool to differentiate between malignant and non-malignant prostatic tissues. Aubertin et al. interrogated 32 fresh prostatectomy specimens using Raman spectroscopy to identify and grade prostate cancer : 20–50 Rn = 4) as well as ex vivo spectroscopic analysis  to differentiate between cancerous and non-cancerous prostate tissue [Intraoperative surgical guidance is an area of active research for clinical implementation of Raman spectroscopy. While restricted to small cohort studies, researchers have developed biopsy-needle-compatible probes to support real-time assessment of the tumour margins. Pinto et al. integrated Raman spectroscopy to the normal workflow of robotic-assisted radical prostatectomy for in vivo . Analysis of the Raman spectra was performed by machine learning algorithms and achieved a sensitivity of 88.2% and specificity of 87.9% in detecting CRPC [Over the last decade, novel inhibitors of the androgen receptor pathway have significantly improved the outcomes of patients with incurable prostate cancer . Despiteing CRPC .Pelvic lymph node dissection (PLND) is the procedure whereby pelvic lymph nodes at risk of harbouring metastatic deposits are surgically removed. Accurate detection of lymphatic metastasis is essential to guide treatment decisions. The standard of care nomograms to determine whether PLND will be performed suffer from both false negatives  and false positives (resulting in overtreatment) ,46. OccuUsing a similar approach, Raman spectroscopy can be used to detect bony metastasis in vivo. Bone metastasis occurs in 8–35% of newly diagnosed prostate cancers ,52,53. Rp < 0.001) [Understanding the mineralisation of bone is equally important in detecting metastasis. In a severe combined immunodeficient mouse model, bone metastasis was introduced by direct injection of human prostate cancer (LNCaP or its derived C4–2b subclone) cells into the tibia of the mice. Raman spectroscopy-based analysis revealed significant changes to bone mineralisation in the cortex of tumour-bearing tibia. The ratio of carbonate:matrix of bone is an important parameter associated with tissue maturity and disease. In bones affected by prostate cancer, the carbonate:matrix ratio significantly decreases, suggesting elevated bone turnover (< 0.001) . The comRecent advances in metabolism show that prostate cancer cells are capable of reprogramming their mitochondria to increase the synthesis of lipids . Raman sJamieson et al. investigated the effects of propranolol, a beta-blocker, on lipid metabolism in prostate cancer cells. Treatment with propranolol resulted in lipid accumulation in PC-3 cells compared to non-cancerous PNT2 cells . The adrIn the earlier sections of this review, we have focused on the application of Raman spectroscopy techniques on disease diagnosis in tissues. However, clinically relevant diagnostics have also been investigated for samples removed from the body, in particular pertaining to blood analytes. Blood analytes include cell-free DNA (cfDNA), circulating tumour cells (CTCs) and extracellular vesicles (EVs).Medipally et al. compared mean Raman spectra of plasma from 43 patients with prostate cancer and 33 healthy volunteers. An increase in the bands associated with nucleic acids from cell-free DNA was observed in the Raman spectra of plasma from prostate cancer patients , wherebyRaman spectroscopy has also been employed to generate spectral fingerprints from extracellular vesicles (EVs) released by prostate cancer cells. EVs are small, lipid-bound particles containing nucleic acid and protein, which are released by metabolically active cells into the tumour microenvironment, with some EVs found in the circulation. EVs function to support communications between tumour cells and their local environments . Lee et Raman spectroscopy is highly versatile and can be used to analyse urine samples in addition to blood. Del Mistro et al. carried out a proof-of-concept SERS analysis on urine samples from 18 patients . Using a machine learning algorithm, the spectral data were classified into two classes . Only one sample was misclassified by this model, leading to a sensitivity of 100% and a specificity of 89% . EquallyOne of the most common clinical tests to identify men at risk of prostate cancer is measurement of the serum level of prostate-specific antigen (PSA) protein. Unfortunately, the PSA test has its limitations, commonly associated with type I errors . It can Raman imaging techniques are optical methods which are minimally invasive and typically non-destructive when conducted under optimised conditions. Given that Raman imaging techniques provide chemically specific detection, often without the requirement for contrast agents, there remains a huge potential for in vivo applications across the remit of disease diagnosis, monitoring therapeutic outcomes and real-time surgical guidance. With appropriate design, SERS nanoparticles have shown efficient sensitivity and specificity for disease diagnosis in a variety of biofluids.Technological advances in lasers and optical components have been realised over the past decade, and to that end, relatively inexpensive, off-the-shelf infrared Raman devices are already used to differentiate between malignant and benign tissue in breast cancer . FurtherConsiderable potential exists for the use of fast-acquisition SRS imaging for surgical applications. As a label-free optical imaging method, based on differences in the Raman spectral profiles, SRS has been successfully applied to differentiate tumours against non-neoplastic tissue in a glioblastoma murine model , along wFor diagnostic applications, SERS-based techniques have shown considerable utility within prostate cancer disease. However, point-of-care SERS devices are currently limited by the requirement for trained personnel to conduct the analysis, the need for specialised equipment and limits of detection that fall below the required safety standards for applications in medical settings . HoweverDespite its advantages and potential, Raman spectroscopy is yet to be widely implemented in clinical practice . This maConsidering the technological advances that have been realised in Raman microscopy systems over the past ten years, there remain challenges regarding microscope design and subsequent regulatory approval before systems can be deployed in the clinic. For example, many coherent Raman imaging systems are either a bespoke creation built by research groups or are formed by combining relevant, commercially available components. As such, these systems are not conducive for clinical application. Commercial systems which offer the complete multiphoton imaging capability with built-in coherent Raman imaging have recently been launched to the market  and may Within the capability of non-destructive, non-invasive and highly sensitive analysis of biomolecules, Raman spectroscopy-based techniques offer a reliable tool for cancer detection. The spectral information acquired with Raman spectroscopy is beyond the reach of other techniques, especially if applied under in vivo conditions. Testing the potential of Raman spectroscopy for clinical benefit will be a major scope for the next 10 years. In the future, it is foreseeable that there will be a major push for intraoperative use and/or multimodal imaging approaches that utilise the strength of each spectroscopic modality to improve prostate cancer detection and treatment.


Drug-induced liver injury (DILI) has become a serious public health problem. For the management of DILI, discontinuation of suspicious drug or medicine is the first step, but the treatments including drugs and supporting approaches are needed. Reference to clinical patterns and disease severity grades of DILI, the treatment drugs were considered to summarize into hepatoprotective drugs , anticholestatic drug , immunosuppressants (Glucocorticoids) and specific treatment agents . The current article reviewed the accumulated literature with evidence-based medicine researches for DILI in clinical practice. Also the drawbacks of the clinical studies involved in the article, unmet needs and prospective development for DILI therapy were discussed. With the extensive use of drugs and the continuous development and application of new drugs, the safety of drug use is widely concerned. Drug-induced liver injury (DILI) is one of the most common and serious adverse drug reactions (ADRs) . DILI caThe most definitive treatment for DILI is to suspend the offending drug(s) and to avoid re-exposure . After dShown as It required appropriate pharmacotherapy to reduce liver damage after stopping use the offending drug(s), when serum transaminase of patients could not normalize and bilirubin remained abnormal within a long time. Reference to the clinical patterns of DILI, pharmacotherapy was considered to classify as follows: 1) hepatoprotective drugs, 2) anticholestatic drug, 3) immunosuppressants, 4) specific treatment. And their efficacy for DILI based on clinical practice evidence was summarized in Hepatoprotective drugs refer to drugs that can improve liver function, promote liver cell regeneration and/or enhance liver detoxification, and there is no unified understanding in regard to its classification. According to the different mechanism of action, it can be roughly divided into detoxification drugs (eg. NAC and GSH), anti-inflammatory drugs (eg. Glycyrrhizic acid preparation), hepatocyte membrane protector (eg. PPC), antioxidant drugs  .NAC is an N-acetylated derivative of L-cysteine (L-Cys) and a precursor of Glutathione (GSH), and has the pharmacological effects of anti-oxidation, anti-inflammation, dilatation of microvessels and protection of DNA molecules . It is tin vivo by glucuronidase to glucuronic acid and glycyrrhizin acid. At present, Glycyrrhizin acid preparation have been commonly used in the treatment of DILI in clinical application around the world, mainly including compound glycyrrhizin tablets, magnesium isoglycyrrhizinate (MgIG) and other drugs  is the main active ingredient. It can provide endogenous phospholipids to repair damaged liver cell and organelle membrane, so as to restore membrane function and increase the fluidity and stability of cell membrane, which protects from liver diseases caused by a variety of factors . Lei X fBicyclol  is the first chemical new drug independently developed by China and used to treat inflammatory liver injury. It has been recommended by Russian physicians’ clinical guidelines for the treatment of DILI . Its phaSilymarin is a flavonoid isolated for the first time in 1968 from the seed extract of milk thistle plant which is mainly a mixture of lignin-derived flavonols, including silybin, silydianin, silychristin, and isosilybin  with silMultiple drugs have been reported to cause bile duct injury, which mostly affects the biliary epithelium of the interlobular ducts, thereby causing an inflammatory response directed at cholangiocytes. Cholestatic and mixed cholestatic and hepatocellular injury are two of the most severe manifestations of drug induced liver disease (DILD) . There in = 22) although due to uncontrolled design of this study a firm conclusion cannot be drawn  have been widely used in clinic practice, due to its anti-inflammatory, immunosuppressive, anti-allergic and anti-shock effects. GCs was recommended for severe DILI patients in cases where the patients’ serum TBIL levels are exacerbated under regular therapy, regardless of whether serum ALT level decreases or not . In addiIn addition to hepatoprotective drugs, anticholestatic drugs, immunosuppressants, there are some specific drugs that play an important role in DILI treatment. For example, L-carnitine was used for liver injury caused by 3-keto-valproic acid (VPA). Anticoagulants such as low-molecular-weight heparins and defibrotide were used for sinusoidal obstruction syndrome (SOS)/hepatic veno-occlusive disease (VOD).LC is a vitamin-like amino acid derivative, mainly derived from daily diet, especially red meat and dairy products. It plays a key role in physiological metabolism include promoting the transport of long-chain fatty acids, preventing the accumulation of acetyl-CoA in mitochondria, and thus promoting the β oxidation of fatty acids, which further are widely used in weight-loss products . In 1982Drug-induced liver vascular injury is relatively rare with unknown pathogenesis, in which the target cells can be hepatic sinus endothelial cells, hepatic venules, as well as the hepatic vein and the portal vein, and the clinical types include sinusoidal obstruction syndrome (SOS)/hepatic veno-occlusive disease (VOD) , peliosiThe mixed phenotype of DILI was often associated with multiple manifestations of cholestasis and hepatocyte injury, with significantly increased all TBIL, ALP, ALT and AST. Despite the lack of adequate evidence-based medical evidence, it is not uncommon for clinicians to choose two even more hepatoprotective drugs for DILI treatment in clinical practice. A clinical study involving 40 Chinese patients with DILI during chemotherapy after radical gastritis showed that PPC combined with bicyclol significantly reduced serum ALT, AST and ALP levels compared with PPC treatment . A cliniMost cases of DILI are self-limiting and resolve with prompt identification and cessation of the offending agent. The patients with abnormal liver function after drug withdrawal and chronic DILI   are ofteAcute DILI had been reported to occur in 5–10% of patients hospitalized for jaundice , and morin vitro mechanical, chemical or biological devices, AELS is a temporary or partial replacement of liver function to assist in the treatment of liver dysfunction or related diseases, mainly including biological artificial liver (BAL), no-biological artificial liver (NBAL) and their mixture. NBAL plays an important role in clinical practice, but its curative effect remains to be discussed. An open randomized controlled trial in regard to High-volume plasma exchange (HVP) in patients with ALF showed that treatment with HVP can increase liver transplant-free survival (58.7 vs. 47.8%), and the incidence of severe adverse events was similar to control group  set up by DILIN in 2012 and HepaTox website (http://www.hepatox.org) by Chinese Medical Association drug-induced liver disease Group in 2014 records information on more than 1,000 common drugs for liver damage. It can provide a basis for avoiding the use of potential hepatotoxic drugs in clinical practice.DILI is one of the most common and serious adverse drug reactions (ADRs), leading to acute liver failure (ALF) and even death in severe cases . Up to nFor pharmacological therapy of DILI, there is a lack of normative guidance or guidelines. There is usually a complete or near complete resolution of DILI within a matter of days to weeks after offending drug withdrawal. The starting point of protective drugs intervention is still difficult to choose. Therefore, dynamic monitoring of liver biochemical parameters is recommended by all clinical practice guidelines. Based on the type of target cells damaged, patients tend to show different clinical phenotypes of DILI with a distinction in liver biochemical indicators. In this paper, we reviewed a large amount of evidence-based medical studies and summarized the commonly used drugs for DILI in clinical practice. NAC, MgIG, PPC, bicyclol, silymarin are potential hepatoprotective drugs for DILI. In anticholestatic drugs, UDCA, SAMe, cholestyramine showed therapeutic value for DILI. LC is recommended for liver injury caused by VPA. GCs is suitable for patients with obvious hypersensitivity or autoimmune symptoms and no obvious improvement in liver biochemical indicators after withdrawal. Anticoagulants, such as low-molecular-weight heparins and defibrotide, play irreplaceable roles in some special types of DILI . They could provide reference for the selection and combined application of protective drugs for different clinical phenotypes of DILI. Nevertheless, the definite therapeutic effects of these drugs are still to be confirmed by prospective randomized and controlled studies.in vitro and in vivo  in APAP-induced liver injury was reported crucial recently. Despite the lack of clinical trials, hepatocyte apoptosis of APAP-treated mice decreased following administration of a CXCL9 neutralizing antibody or CXCR3 gene knockout . In huma in vivo . Attempt in vivo . Excitin in vivo .


Background: The association between abnormal invasive fractional flow reserve (FFR) and the fat attenuation index (FAI) of lesion-specific peri-coronary adipose tissue (PCAT) is unclear.Method: Data of patients who underwent coronary computed tomography angiography (CTA) and subsequent invasive coronary angiography (ICA) and FFR measurement within 1 week were retrospectively included. Lesion-specific FAI (FAIlesion), lesion-free FAI , epicardial adipose tissue (EAT) volume and attenuation was collected, along with stenosis severity and plaque characteristics. Lesions with FFR <0.8 were considered functionally significant. The association between FFR and each parameter was analyzed by logistic regression or receiver operating characteristic curve.Result: A total of 227 patients from seven centers were included. EAT volume or attenuation, traditional risk factors, and FAInormal  were not significantly different in patients with or without abnormal FFR. In contrast, lesions causing functional ischemia presented more severe stenosis, greater plaque volume, and higher FAIlesion . Additionally, the CTA-assessed stenosis severity  and FAIlesion  were determined to be independent factors that could predict ischemia. The combination model of these two CTA parameters exhibited a diagnostic value similar to the invasive coronary angiography (ICA)-assessed stenosis severity .Conclusion: It was FAIlesion, not general EAT parameters, that was independently associated with abnormal FFR and the diagnostic performance of CTA-assessed stenosis severity for functional ischemia was significantly improved in combination with FAIlesion. CT) had been proved to significantly improve diagnostic accuracy for the detection of ischemia-causing lesions and is recommended to evaluate the functional significance of intermediate stenosis (30–90%) to help guide invasive coronary angiography (ICA) referral and revascularization treatment planning  and stenosis severity did not improve the predictive ability of CTA on FFR abnormal (The presence of coronary artery disease (CAD), specifically the presence of a flow-limiting lesion, is a primary reason for the onset of myocardial ischemia. Invasive fractional flow reserve (FFR) is highly recommended to determine the drop in flow around a coronary lesion to guide decision-making for coronary revascularization , 2, but stenosis 0–90% to planning . Howeverabnormal . Meanwhiabnormal –9, but tabnormal –12, and abnormal –16.In the past few years, peri-coronary adipose tissue (PCAT) had been shown to be an updated image biomarker of CAD –21. Fat This study was conducted to investigate the association between lesion-specific FAI and abnormal FFR, and to assess whether lesion-specific FAI could enhance the diagnostic ability for functional ischemia in combination with other coronary CTA parameters to facilitate clinical decision making.This retrospective cross-sectional study included data of patients scanned between April 2017 and October 2019 at seven centers . The inclusion criteria were as follows: (1) patients with suspicion of CAD and underwent coronary CTA as part of the clinical routine; (2) an invasive coronary angiography (ICA) in combination with FFR measurement was performed within 1 week in a non-emergent setting. The exclusion criteria were as follows: (1) poor image quality (such as significant artifacts in the interested segment) or too limited or minimal PCAT to be analyzed (the thickness of PCAT was less than the diameter of the adjacent artery); (2) patients with anatomic variations in the heart or coronary arteries ; (3) patients with previous CAD history, such as previous percutaneous coronary intervention, coronary artery bypass graft surgery or myocardial infarction; (4) patients with total occlusion lesion/s; (5) patients with malignancy or cardiomyopathies.This retrospective study was approved by the institutional review board of Shengjing Hospital of China Medical University (No. 2021PS689K). Because it was a retrospective study and the examination was necessary for the clinical diagnosis of the subjects, the exemption of written informed consent was approved by the institutional review board.All coronary CTA data were reviewed by a single experienced observer (with over 10 years experience) who was blinded to clinical data of the patients. All patients were divided into functional ischemia group (Group A) and non-functional ischemia group (Group B) based on the invasive FFR measurements. The study design and grouping scheme were shown in All CTA scans carried out by each center were in accordance with the guidelines recommended by the Society of Cardiovascular Computed Tomography . All of Peri-coronary adipose tissue is a part of EAT and is defined as the adipose tissue voxels [−190 to −30 Hounsfield units (HU)] located within a distance from the outer vessel wall equal to the diameter of the respective vessel (lesion) and lesion-free FAI , respectively. Lesion-free PCAT was defined as adipose tissue surrounding coronary arteries without atherosclerosis in the proximal segment of the Right Coronary Artery. If the RCA was atherosclerotic, PCAT of the left anterior descending (LAD) artery or the left circumflex (LCX) artery was used to measure FAInormal.To conduct the analysis of FAI according to the lesion status of a vessel, we measured the lesion-specific FAI  for FAIlesion measurement.To assess the repeatability of FAIEpicardial adipose tissue volume and attenuation measurement was performed on an offline workstation . EAT was defined as adipose tissue contained within the visceral pericardium. The visceral pericardium was identified and traced manually on axial CT images between the levels of pulmonary trunk bifurcation and cardiac apex. EAT volume was automatically calculated (in ml) using contiguous voxels with threshold attenuation of −190 to −30 HU  as the rThe plaque characterization was performed and a series of quantified plaque features were measured with a dedicated plaque analysis software . Maximum diameter stenosis (DS), total plaque volume (TPV), and remodeling index (RI) were recorded using a method described previously . PlaquesSelectively invasive coronary angiography (ICA), stenosis assessment, and FFR measurement were performed in each center by an experienced interventional cardiologist who was blinded to the CTA results. Standard quantitative coronary analysis (QCA) was used to quantify the ICA stenosis severity with the method as previously described. FFR was performed with a coronary pressure wire  subsequently in vessels with intermediate stenosis between 30 and 90% , 16, 26.Data were analyzed using commercially available software . The continuous variables were described by mean ± standard deviation or median (interquartile range). The categorical variable was expressed as a percentage.t-test, Mann-Whitney U-test, and chi-square test were applied to compare the difference of EAT parameters, FAIlesion, FAInormal, plaque characteristics, and traditional CAD risk factors between patients (or vessels) with and without FFR abnormity. Paired t-test was performed to compare the intra-patient difference of FAInormal and FAIlesion.Independent-sample lesion, plaque characteristics, ICA stenosis severity, and functional ischemia. The receiver-operating characteristic (ROC) curve was used to analyze the discriminatory power of each parameter for predicting coronary functional ischemia. To assess the repeatability of FAIlesion measurement, inter-observer variability was evaluated using the intraclass correlation coefficient (ICC). A p < 0.05 was considered statistically significant.Univariate and multivariate logistic regression analyses were applied to evaluate the association between FAIThere were 249 patients that underwent coronary CTA scanning, subsequent invasive angiography, and FFR testing. Twenty-two of them were excluded for various reasons (details of which are shown in p = 0.07) and attenuation  between the two groups of patients  patients were identified as having coronary functional ischemia. Three of these patients had ischemic lesions in two separated coronary main branches . Thus, a total of 91 vessels were identified as flow limiting. To evaluate the association between EAT volume/attenuation and functional ischemia, we analyzed these parameters in patients with (group A) or without (group B) functional ischemia. The results indicated that there were no significant differences in the EAT volume , the result showed that FAIlesion was much higher than FAInormal  mm3, p < 0.01, p < 0.01) and CTA  testing. The differences of each plaque component ratios and high-risk plaque features between lesions with or without functional ischemia were non-significant , we found that severity of stenosis and FAIlesion were independently associated with abnormal FFR  mmnificant . With unrmal FFR .lesion for diagnosing functional ischemia. When integrating CTA stenosis and FAIlesion (Model 1), the AUC of these combined non-invasive parameters increased and presented diagnostic performance similar to ICA stenosis . Combining FAIlesion and ICA stenosis (Model 2), AUC was further improved beyond ICA stenosis alone . Details of ROC analysis were shown in According to the ROC curve analysis, ICA stenosis severity had a higher AUC compared with either CTA stenosis severity or FAIlesion measurement was 0.87 (95% CI:0.81–0.90) for average measures.The ICC of inter-observer FAIlesion or CTA lesion stenosis severity was individually inferior to invasive stenosis severity measurement, the predictive value of the combination model using these two CTA parameters was significantly improved and comparable to the ICA stenosis measurement.In the present study, we found that patients with or without functional ischemia caused by CAD (as determined by invasive FFR) might share similar EAT features and lesion-free PCAT attenuation. In contrast, lesion-specific FAI was independently associated with reduced FFR. Although the diagnostic performance of FAITo assess whether the difference of lesion-specific FAI between ischemic or non-ischemic lesions resulted from the difference of general EAT status, the comparison of EAT volume/attenuation between patients with and without functional ischemia was conducted. Some investigations have shown that EAT volume was associated with myocardial ischemia detected by nuclear imaging , 29, butlesion was significantly higher than FAInormal. Thus, we assumed that vessel-specific FAI measurement could be affected by the ratio of normal to lesion segment in the measuring area, and lesion-specific FAI measurement might be a better indicator with lower variability. Two recent published studies used lesion-specific PCAT attenuation to predict abnormal FFR, but they reached contradictory results (lesion) in comparison to general EAT parameters.Instead of EAT, the PCAT was taken to be a better indicator of the development of coronary atherosclerosis and the FAI had been proven to be associated with not only vessel inflammation but also patient's prognosis , 20. Stu results , 24. Thi results  manuallylesion is higher for lesions with FFR <0.8.Studies have proposed that vascular inflammation was considered to be a contributor to endothelial dysfunction and would inhibit lipid accumulation in adipocytes of PCAT , 30, 31.lesion), as shown in the present results. Although stenosis severity assessed by coronary CTA was highly consistent with ICA, there were interference factors such as inappropriate intraluminal contrast enhancement, spatial resolution, and coronary calcification that would affect its accuracy  could be improved by a focal inflammatory parameter (FAIaccuracy . This co3). Moreover, due to the inconsistent results of the individual plaque characteristic and high-risk plaque features on abnormal FFR  and high-risk plaque features were independently associated with abnormal FFR . Accordirmal FFR , 37, 38,lesion and CTA-assessed coronary stenosis severity had a much higher sensitivity than any other parameter for the diagnosis of functional ischemia lesions. That meant FAIlesion and CTA-assessed stenosis severity, as a non-invasive method, could help us to rule out ischemic coronary stenosis more safely. As patient safety is the primary concern of clinical practice and the measurement of lesion-specified FAI does not require extra protocols or radiation exposure within routine coronary CTA, we believed that it was reasonable for quantifying lesion-specified FAI to facilitate clinical decision-making in patients with intermediate coronary stenosis assessed by CTA.Our study indicated that the combination of FAIlesion. Finally, because fixed tube voltage (120 kV) was applied in the present study and the attenuation of adipose tissue could be affected by the tube potential, the correction of HU value might be in demand for EAT/PCAT analysis if other scanning parameters were applied.This study also had several limitations. First, as the enrolled subjects were clinically selected with intermediate lesions, they might not represent people with a low risk of CAD and severe diffuse coronary atherosclerosis. Second, based on the cross-sectional design, no causal relationship could be confirmed although multivariate regression analysis was performed. Moreover, no outcome data were included in the present study, and further research was needed to test the prognostic value of FAIlesion could be used to facilitate clinical decision-making and formulate therapeutic strategies.Lesion-specific FAI measured by coronary CTA was independently associated with abnormal FFR and could improve the diagnostic performance in combination with morphological stenosis severity assessment. Therefore, the combined diagnostic model with FAIThe original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by the Institutional Review Board of Shengjing Hospital of China Medical University. Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.SM and YH contributed to conception and design of the study. HL, JZ, LX, YW, TL, and KW conducted the data collection. SM, XC, YM, and JY conducted the image analyzing and statistical analysis. SM wrote the first draft of the manuscript. All authors contributed to manuscript revision, read, and approved the submitted version.This study was granted by the National Natural Science Foundation of China ; The Key Research & Development Plan of Liaoning Province (No. 2020JH2/10300037); 345 Talent Project in Shengjing Hospital of China Medical University.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Both social and medical factors can negatively affect health outcomes, especially in vulnerable populations. To address these two types of factors in a hospital post-discharge population, two non-profit organizations collaborated to combine their novel decision support programs to address the question: could combined programs have greater potential for improved health outcomes? HomeMeds (HM), a social health program in which trained social services staff make home visits to vulnerable clients, was combined with MedSafety Scan (MSS), a medical health, clinical decision support tool. Data captured in the home visits were entered into the HM and MSS programs to detect those patients at greatest risk of adverse health outcomes due to medications. Patients received a post-discharge home visit by trained social services staff. The number of drugs reported as being taken was on average 10.4, which was less than prescribed at discharge in 62% of patients. Both programs detected serious risk of medication-induced harm, mostly from different causes such as drug-drug interactions or for use not recommended in older adult. Combined analysis of data from two novel decision support programs yielded complementary findings that together address both medical and social determinants of health. These have the potential to reduce medication-induced harm, costly re-hospitalization and/or emergency room visits and support the further evaluation of this combined approach in other vulnerable populations such as the seriously mentally ill, frail, those confined to home, opioid-dependent or otherwise impaired.


Additive manufacturing (AM) is the automated production of three-dimensional (3D) structures through successive layer-by-layer deposition of materials directed by computer-aided-design (CAD) software. While current clinical procedures that aim to reconstruct hard and soft tissue defects resulting from periodontal disease, congenital or acquired pathology, and maxillofacial trauma often utilize mass-produced biomaterials created for a variety of surgical indications, AM represents a paradigm shift in manufacturing at the individual patient level. Computer-aided systems employ algorithms to design customized, image-based scaffolds with high external shape complexity and spatial patterning of internal architecture guided by topology optimization. 3D bioprinting and surface modification techniques further enhance scaffold functionalization and osteogenic potential through the incorporation of viable cells, bioactive molecules, biomimetic materials and vectors for transgene expression within the layered architecture. These computational design features enable fabrication of tissue engineering constructs with highly tailored mechanical, structural, and biochemical properties for bone. This review examines key properties of scaffold design, bioresorbable bone scaffolds produced by AM processes, and clinical applications of these regenerative technologies. AM is transforming the field of personalized dental medicine and has great potential to improve regenerative outcomes in patient care. Hard tissue deficiencies in the maxillofacial region are the result of numerous diseases, disorder and injuries, and appropriate rehabilitative therapies are necessary to restore quality-of-life for affected individuals. The Global Burden of Diseases, Injuries, and Risk Factors Study 2017 (GBD 2017) revealed that oral disorders had the greatest age-standardized prevalence and incidence in the world . PeriodoSimilar to trends for periodontal disease, incidence rates for cancers of the lip and oral cavity are also increasing . SquamouThe regeneration of periodontal defects in humans is case-sensitive due to the involvement of multiple tissue types and variability in defect morphology . For insThe prognosis of regenerative periodontal therapy is dictated by the defect morphology, which primarily considers the number of remaining bone walls and the defect angle . 3-wall The key elements of periodontal regeneration are cells, scaffolds, growth factors, and blood supply . ImproveDental implant therapy is often the treatment of choice to replace missing teeth, offering patients high satisfaction and improved oral health-related quality-of-life following treatment . The suiAlveolar ridge deficiencies are categorized by their severity and defect type, generally described as horizontal, vertical, or combined . More seThe clinical limitations, implications of invasive reconstructive surgical procedures, and prognostic uncertainty are current challenges in regenerative dental medicine. Clinical scenarios in which predictable treatments have yet to be achieved include ridge defects with severe horizontal or vertical components of alveolar bone loss, class III furcations, papilla deficiencies, and advanced peri-implant defects . Additioin vivo  techniques. Suitable biomaterials must demonstrate process compatibility with the specific AM technique applied, as well as appropriate biochemical and physical characteristics to function successfully in vivo . Althougin vivo . Certainin vivo . Additioin vivo , ultraviin vivo , or orgain vivo  may precAM technology enables scaffold production with a diverse array of materials, including polymers, metals, ceramics, hydrogels, and carbon-based nanomaterials . Thermopin vivo. In contrast, although polyglycolic acid (PGA) and poly lactic-co glycolic acid (PLGA) offer superior mechanical properties, they degrade quickly when used as a bioresorbable scaffold and within 2 weeks, their tensile strengthen is reduced by half  and an organic phase, consisting of extracellular matrix proteins, of which approximately 90% is collagen type 1 . The preBy mimicking the physiologic characteristics of native bone, material property tailoring enhances the regenerative capacity of tissue engineered constructs in the presence of biomechanical stresses . This isThe layered construction process utilized in AM is advantageous for the production of lightweight and porous constructs that can support tissue regeneration in an irregular defect. CAD files can be used to generate scaffold configurations that accurately replicate the overall defect shape and dimensions. Further, customized 3D surface topology can be generated by using standard triangle language (STL) files to topologically subtract defects from a digital scaffold design . This prMechanical cues provided by scaffold materials can regulate the fate of stem and progenitor cells . In 2D cViscoelasticity, a key property of living tissues, is another regulator of MSC behavior. Viscoelastic materials exhibit a combination of storage of elastic energy as a solid, and loss of mechanical energy as a fluid. These materials exhibit stress relaxation and hysteresis in the stress-strain relationship during loading and unloading. When a mechanical load is applied then removed, viscoelastic materials can dissipate energy . Chaudhuex vivo prior to transplantation, and to tune stem cell behavior in vivo following transplantation in order to improve their regenerative capacity , vascular endothelial growth factor (VEGF), osteoprotegerin (OPG), and bone morphogenetic protein (BMP)-2 . ComputeRegenerative medicine is based upon the manipulation of known physiologic processes to create a microenvironment that simulates a desired stage of tissue development, thus inducing tissue formation and renewal. Not only must tissue dynamics be replicated at a macroscopic organ or tissue scale, but on the cellular and molecular levels as well. Scaffolds primarily serve to provide an osteoconductive matrix and benefit from the addition of growth factors that exert osteoinductive influence on cellular activity. Biologics such as recombinant human BMP-2 and BMP-7, growth differential factor-5 (GDF-5), EMD, and rhPDGF-BB, have all been well-studied for their capacities to promote osteogenic differentiation and enhance bone formation in regenerative dental medicine . Growth in vivo  technology. It is often used interchangeably with SFF, which also implies the use of a fixture-less platform without part-specific tooling or human intervention, or rapid prototyping (RP), often used in contexts that describe fast fabrication of scale models or parts. AM processes first emerged in the 1980’s and have rapidly evolved to become a powerful tool for biomedical scaffold design, capturing particular interest from the research community in bone tissue engineering. Research involving bone-related applications account for approximately 20% of the existing publications in searches for articles with the terms “additive manufacturing” and “3D printing.”AM fabrication of personalized biomedical constructs begins with the acquisition of high resolution, 3D datasets, typically with computed tomography (CT) or magnetic resonance imaging (MRI), from an individual patient. The images are then converted to a common medical file format referred to as digital imaging and communication in medicine (DICOM). The DICOM file is then imported into a software package that performs segmentation to produce serial sectional data (slices) and reconstruction of a volumetric model composed of voxels. A voxel is a single unit of volume within a 3D grid as opposed to a 2D pixel. A density threshold is then established and the signal intensity of point data is used to determine which points will be included. Surface polygons are then extracted to reconstruct a tessellated, polyhedral model, also known as a mesh which can be exported as an STL file. Surface refinement is performed with algorithms such as non-uniform rational B-spline (NURBS) functions; this can occur either before the creation of the STL-triangulated surface, known as reverse modeling or after, which is the STL-triangulated model converting approach . With thDentistry is a field that has embraced AM techniques and frequently uses commercially available equipment to fabricate patient-specific constructs in everyday practice. AM is considered a technologic hallmark of the 4th industrial revolution and has resulted in a paradigm shift from design for manufacturing to manufacturing for design . In the Depending on the specific tissue and critical defect size, there are numerous options for AM regenerative scaffolds in the oral and craniofacial arena. The predominant methods for non-metallic bone scaffold production can be categorized broadly into extrusion-based, laser-assisted, or binder jetting type processes . The detStereolithography apparatus (SLA) and selective laser sintering (SLS) are the primary laser-assisted techniques for non-metallic bone scaffold production. First developed in 1983, SLA utilizes photochemical reactions with UV lasers to produce scaffolds out of photosensitive polymers. Because of specific material and post-processing requirements associated with toxicity concerns, SLA is not as commonly used for craniofacial regeneration. The main advantage of SLA is its capability for high accuracy and refined internal resolution relative to other AM techniques . First dAggregatibacter actinomycetemcomitans, a facultative anaerobe commonly implicated in periodontal infections s, polyurethanes, polyacetals, poly(ester amide)s, polyanhydrides, polyphosphazenes, and pseudo poly(amino acids) . Their uin vivo compared to bare scaffold controls  may also be combined with synthetic polymeric scaffolds to provide appropriate molecular cues for osteogenic activity. AM-printed constructs have been coated with dECM obtained from bone cells  or non-bcontrols . Furtherin vitro . Applicain vitro .Bioprinting, generally defined as “the use of computer-aided transfer processes for patterning and assembly of living and non-living materials with a prescribed 2D or 3D organization to produce bioengineered structures” , is a prThe selection of bioinks is one of the most critical steps in the process of bioprinting, mainly relying on two important aspects: biofabrication and biocompatibility. Biofabrication usually refers to the printability of the ink, such as the compatibility with the printer and printing resolution, which is highly related to the rheological properties of the bioink. Viscous and shear-thinning hydrogels, such as gelatin and methylcellulose , are oftDental, oral and craniofacial tissues are organized with complex 3D architectures involving multiple types of cells and tissues. Mimicking their 3D complexity and multicellular interactions represents one of the main barriers in dental and craniofacial regeneration . 3D biopin vitro cell culture. The BMP peptide construct group exhibited the highest calcification as compared to the growth medium, osteogenic medium, and control groups with robust expression of osteogenic genes. In addition to pulp-dentin complex, the feasibility of whole tooth bioprinting has been studied by co-printing the hDPSCs-laden bioinks with PCL. The results not only achieved localized differentiation of hDPSCs in the outer region of the 3D cellular construct but also successfully produced 3D patient-specific cellular constructs for tooth tissue engineering in a predefined pattern  can differentiate into several cell types, including odontoblasts, neural progenitors, osteoblasts, chondrocytes, and adipocytes with high proliferative capability . Therefo pattern .Engineering craniomaxillofacial bone tissue is a unique challenge due to the complex architecture of bone, consisting of organized calcified regions with interpenetrated vasculature . In ordein vitro systems that would otherwise be challenging for extrusion-based processes to realize, including a liver lobule model  contour models; (2) guides; (3) splints; and (4) implants . ContourAdditive manufacturing is now widely employed in dentistry for a variety of purposes, including fabrication of dentures, occlusal splints, temporary crowns and bridges, orthodontic appliances, and surgical guides. More recently, customized, non-resorbable titanium metal cages intended for extraosseous alveolar ridge augmentation have become available to clinicians as well. Despite prolific use of commercially available AM equipment to create custom dental devices in every day clinical practice, attempts to utilize bioresorbable bone scaffolds produced by this technology have only recently begun in academic, clinical settings. In 2015, the first dental use of a personalized, bioresorbable scaffold produced with AM in humans was reported. A PCL scaffold fabricated by selective laser sintering was loaded with rhPDGF-BB solution and implanted into a periosseous defect in the periodontium localized to a mandibular canine site . The desCustomized, bioresorbable bone scaffolds created by AM processes have been tested more extensively in the fields of maxillofacial and craniofacial surgery. In 2016, a clinical case series described 20 patients who received artificial bone constructed with 3D inkjet printing to graft non-weight bearing, maxillofacial bone deformities in 23 sites . These sBone grafting is a routine procedure in clinical dentistry and may occur in approximately half of all dental implant sites . AugmentContemporary biomedical research is steadily approaching a reliable AM strategy to create bioresorbable bone scaffolds for clinical use and the implications for patient care are enormous. Recently, a workflow for AM fabrication of porous, bioresorbable scaffolds consisting of medical grade PCL for the reconstruction of large, posterior mandibular defects was demonstrated . The resAM holds tremendous promise for the advancement of regenerative medicine; however, this impressive technology must overcome several obstacles before it can be extensively introduced to clinical settings for the purpose of fabricating personalized bone tissue scaffolds. First, AM has historically been limited by relatively low production speed. In 2017, the average build time to create a personalized object constructed by AM techniques in the field of craniomaxillofacial surgery is approximately 18.9 h but can be as high as 96 h per object . Practicin vivo.Despite these potential limitations, the technologic infrastructure necessary to produce bioresorbable scaffolds for bone regeneration by AM is available for implementation. Successful translation to clinical use now relies upon the ability to manipulate biomaterials and precisely coordinate their architectural and biochemical features with known physiologic mechanisms of tissue formation. This process is further complicated by the addition of exogenous agents, such as viable cells, growth factors, gene vectors, drugs, and other bioactive components, which must be present in appropriate quantities to simulate a suitable microenvironment for regeneration. Simultaneous incorporation of viable cells during scaffold fabrication remains a preeminent challenge. Ensuring cell survival during the AM process and preserving their phenotype and morphology post-processing will require continued development of cell-deposition techniques and cell-carrier systems. Improved control of cellular responses will benefit from the progressive advancement of scaffold-based gene therapy techniques. Finally, innovation in the use of hydrogel bioinks and 3D bioprinting processes will further refine spatiotemporal regulation of biomolecular signaling and progress efforts to regenerate bone tissue with bioresorbable, biomimetic scaffolds in vivo conditions  printing, in which time is the fourth dimension of the printed construct . 4D prinnditions . Potentinditions  and shapnditions  are settnditions , which me defect , is an ie defect , especiaThe treatment of dental, oral, and craniofacial bone defects is currently restricted by available biomaterials, which have limited capacity to facilitate true regeneration of new tissues that exhibit native physiologic form, function and esthetics. Further research efforts are needed to optimize AM for the production of bioresorbable scaffolds that yield safe, predictable, and efficacious clinical outcomes in the reconstruction of bony defects. More preclinical studies are needed to improve the material properties and clinical performance of polymer-ceramic composite scaffolds for bone reconstruction and to refine understanding of the architectural features that promote formation of an anatomic periodontal ligament compartment. Additionally, tremendous opportunity exists to functionalize scaffolds for therapeutic purposes, especially with regards to gene therapy. As the understanding of multifaceted biomaterial interactions and tissue dynamics improves within the scientific community, AM offers a promising future in which a superior generation of sustainable regenerative biomaterials will become accessible for everyday clinical use. Commercialization of custom scaffold technology will dramatically accelerate the trend toward increased usage of synthetic bone substitutes and expand their existing market share within the multibillion dollar industry for biomaterials. Successful adaptation of AM technology for bone tissue engineering will expose a new realm of regenerative possibilities within dental medicine, thus expanding treatment options for patients and significantly improving their oral health related quality-of-life. Eventually, personalized bone constructs for dental regenerative medicine will evolve from state-of-the-art technology to a new standard in patient care.WG and JL contributed to conception and design. JL led the writing of the manuscript. SM, YY, DW, and MC wrote sections of the manuscript. All authors contributed to manuscript revision. All authors read and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The global community decided in 2015 to improve people’s lives by 2030 by setting 17 global goals for sustainable development. The second goal of this community was to end hunger. Plant seeds are an essential input in agriculture; however, during their developmental stages, seeds can be negatively affected by environmental stresses, which can adversely affect seed vigor, seedling establishment, and crop production. Seeds resistant to high salinity, droughts and climate change can result in higher crop yield. The major findings suggested in this review refer nanopriming as an emerging seed technology towards sustainable food amid growing demand with the increasing world population. This novel growing technology could influence the crop yield and ensure the quality and safety of seeds, in a sustainable way. When nanoprimed seeds are germinated, they undergo a series of synergistic events as a result of enhanced metabolism: modulating biochemical signaling pathways, trigger hormone secretion, reduce reactive oxygen species leading to improved disease resistance. In addition to providing an overview of the challenges and limitations of seed nanopriming technology, this review also describes some of the emerging nano-seed priming methods for sustainable agriculture, and other technological developments using cold plasma technology and machine learning. World food production must increase 50% by 2050 to meet the needs of 9 billion people . The groSeeds are miniature plants which, when fertilized, hatch into ovules containing both a germinating embryo and enough food to grow. A variety of treatments can be applied to a seed before planting in order to enhance its quality and potential yield. The process of priming seeds, in its broadest sense, involves soaking them in a solution where enough water is absorbed so that germination of the seed becomes possible, but not enough to allow the radicle to protrude through the seed coat . Using t2•− and converts it into H2O2, and the enzyme catalase transforms H2O2 into water. A significant increase in antioxidant enzyme amounts could be the result of nanoparticle-mediated reduction of ROS. Due to the relationship between reactive oxygen species and antioxidant enzymes, seeds that are nanoprimed tend to contain a lot of antioxidant enzymes through nanoparticle-mediated ROS mitigation. Furthermore, aquaporin and ROS-mediated interactions have been found to influence seed development [2O2 and ROS through biological membranes as well as water uptake [Sclerospora graminicola, which causes downy mildew. By priming seeds with chitosan nanoparticles, plants display improved immunity, with increased gene expression of alkaline peroxidase, polyphenoloxidase, and phenylalanine ammonia lyase. The nanochitosan seed priming over expression involves PR1 and PR5, which are involved in the salicylic acid pathways. Using zinc oxide nanoparticles to combat environmental stress factors was studied by Chaudhary et al. [An analysis of the studies shows that the benefits of nanomaterial in promoting seed germination include (a) developing nanopores in the seed coat, (b) introducing reactive oxygen species (ROS) to the seed, and (c) using the nanocatalyst to boost enzyme activity at the starch-degrading site  nanoparticles were shown to mitigate heavy metals uptake and toxicity in wheat seedlings. Physiological mechanisms were investigated in wheat seedlings to determine how magnetic nanoparticles (nano-Fe3O4) mitigated the toxic effects of heavy metals  [By seed priming with cerium oxide nanoparticles, An et al. studied the molecular mechanisms behind plant salinity stress tolerance. Poly (acrylic acid)-coated cerium oxide nanoparticles (PNC) show morphological, physiological, biochemical, and transcriptomic effects on cotton seedling priming under salinity stress, according to An et al. An advantage of PNC nanoparticle seed priming is that it can be used to increase a crop tolerance to stress during the early seedling stage in a sustainable, practical, and scalable way. These results showed promising molecular mechanisms that could be synergistically operated to enhance plant salt tolerance . Accordiiculture . The stuiculture . Khan eticulture . The roltributes . A studytributes . In a st and Cu) .P. grisea and X. oryzae, Sathayabama and Muthukumar investigated the antimicrobial activity of chitosan guar nanoparticles (CGNP). The treated rice showed no symptoms of blast disease. Rice plants with CGNP-applied antimicrobial protection displayed an enhanced rate of seed germination and growth [Fusarium species) as shown in As a bioprotectant against rice borne pathogens d growth . T. Xu ePseudomonas species enhancing growth, physiology, and antioxidant metabolism in Melissa officinalis L. According to P. fluorescens and P. putida significantly increased plant biomass indices, the relative water content of leaves, photosynthetic pigment values, essential oil yield, and all major components (except thymol). It has been found that seed priming with nano-silicon particles combined with inoculation with pseudomonas strains boosts the number of both primary and secondary metabolites in lemon balm plants [P. agglomerans improved germination, size, and weight of the roots. Moreover, seed coating with Bacillus californianus increased leaf number and dry weight. In view of this, the technique applied in the current study to prime seeds with nanofiber-immobilized PGPR could be regarded as a promising eco-friendly strategy to enhance soybean production by using microbial inoculants [. Combining seed treatment with colloidal solution of Mo nanoparticles and microbial preparation results in a four-fold increase in nodule formation compared to control plants [Seed germination and seedling growth characteristics of several plant species have been enhanced by nanopriming with a variety of nanomaterials ,44. Sevem plants . Shcherbm plants . De Gregoculants . Accordil plants .Cichorium intybus with selenium nanoparticles. This study provides a better understanding of the potential advantages of cold plasma in improving early growth, protection, and production. The combined potential benefits of working with cold plasma and metal-based nanoparticles seed priming are rarely studied. The reactive oxygen and nitrogen species (ROS and RNS) are important signaling molecules that are involved in seed signaling pathways, cellular physiology, gene expression, differentiation, and growth . Seed trMelissa officinalis has shown to respond differently to zinc oxide (nZnO) and selenium (nSe) nanoparticles when primed with cold plasma. The primed seeds with plasma were cultured in nutrient solution modulated with nSe and nZnO. As a result of plasma priming, the growth-related characteristics  as well as biomass accumulation were improved, as well as the toxicity signs of nSe were attenuated [Capsicum annuum) under in vitro and in pot conditions using a functional scientific device and metal-based nanoparticles. Eliciting peroxidase enzyme activity in both culture media was achieved by treating either with plasma or nZnO on both roots, and it was found that plasma activity and/or nZnO activity of phenylalanine ammonia-lyase were significantly greater. Plant growth was most influenced by soaking seeds with nanomaterials before plasma treatment in the pot experiment [Using non-thermal plasma, A. Babajani et al. studied selenium and zinc oxide nanoparticle reactions of seed priming. Medicinal plant tenuated . The stutenuated . Cold plperiment .Seeds with better seed germination and seedling emergence rates can ensure reliable emergence under a variety of agricultural conditions and are therefore instrumental in ensuring yield potential and uniformity. As of today, germination scoring is typically done by human observation, and therefore, has a limited frequency, scale, and accuracy. In order to handle this bottleneck, many attempts have been made to automate both seed diagnostics and associated phenotypic analysis, with more than one research-based solution such as germinator, phenoSeeder, and the MultiSense tool as a result . As showRecently, advanced computer vision and machine learning methods have been applied to germination assays; among them, the Rice Seed Germination Evaluation System which uses artificial neural networks to gauge the germination status of Thai rice species . NumerouIt is recognized that despite remarkable progress, scientists do not have a complete understanding of how these nanomaterials can affect the macro- and micro-environments of seeds. It is alarming that there is still a lack of basic understanding regarding the possible health and safety effects of engineered nanomaterials on both human and non-human receptors considering the actual and projected levels of exposure . A generNanopriming can be a potential future target for sustainable agriculture with the current global supply versus demand trend. A fortification program consists of adding vitamins and minerals to processed foods as a public health measure to improve the nutritional quality of the food supply. The goal is to enhance the nutrient intake by the population . As a re


As numbers of sandwiched caregivers in the United States grow, it is essential to document the literature on the impact of dual care responsibilities on aspects of psychological well-being and physical health. This scoping review examined the literature on sandwiched caregivers’ psychological well-being and physical health, identified gaps in the literature, and provided suggestions for future studies to advance the literature on sandwiched caregivers in the United States. Guided by the Arksey and O’Malley (2005) framework, this scoping review comprised of 15 peer-reviewed articles between 1980 and 2019, that examined aspects of the psychological well-being  and physical health  of sandwiched caregivers in the United States. Findings showed that there was ambiguity surrounding the conceptualization of sandwiched caregivers, specifically how older and younger care recipients were defined. Also, most studies examined psychological well-being while physical health was understudied. The findings of this review also showed that, compared to non-sandwiched caregivers  and non-caregivers, sandwiched caregivers exhibited greater depressive symptoms and psychological distress as well as poorer health behaviors. Furthermore, sandwiched caregivers who were female and employed were more susceptible to greater depressive symptoms than their employed male counterparts or employed non-caregivers. In considering future directions, more work is needed that examines physical health. Additionally, sandwiched caregivers of minority status merit attention as multigenerational care occurs at greater rates in these populations. Finally, caregiving during the pandemic may have a detrimental impact on sandwiched caregivers’ lives which should be investigated.


Host-microbiota interactions have increasingly been shown to play important roles in supporting host health . In humaLiu et al.). They found that microbial diversities were similar between these two techniques, suggesting SMS as an economical approach for future clinical tests. This study identified dominant species, such as Prevotella copri, Bacteroides dorei, and Bacteroides vulgatus, that might be responsible for the progression of colorectal diseases. These strains may serve as chassis to diagnose, or even engineered to treat colorectal diseases. Next-generation sequencing (NGS) technology was also demonstrated to be a powerful technique in the study by Chen and coauthors to detect fetal copy number variant for non-invasive prenatal testing . With increased read depth and reduced cost, it is foreseeable that NGS will have more contributions in the future.In order to modulate microbiota, we need to understand the composition of microbiota and their interactions with host. In this respect, Liu and coauthors used 16S rRNA gene amplicon and shallow metagenomic sequencing (SMS) to explore the gut microbiota directly from patient samples with colorectal disease . They discovered that abundance of membrane GLUT1 and cytoplasmic G6PD was greatly reduced and the GPR109a-AKT signaling pathway was vital in regulating these changes. More strikingly, they found that butyrate could promote chemotherapeutical efficacy of 5-fluorouracil on cancerous colonocytes, possibly due to exacerbated impairment of DNA synthesis. This study provides valuable information for the molecular mechanism of butyrate on glucose metabolism of colorectal cancer cells. These discoveries could promote the development of beneficial metabolites as therapeutical or adjuvant anti-cancer drugs. It is worth noting that butyrate could improve the antivirus capacity of human . They focused on LuxR, a bacterial quorum sensing regulator from Vibrio fischeri, which can be engineered to sense environmental signals and control overall microbiota behaviors. In this work, they identified a key residue, isoleucine-46, when modified to F or R, can have different binding affinity to the same substrate 3-oxo-hexanoyl-homoserine lactone. They implemented the designed transcription factors into genetic circuits in E. coli to detect target bacterial pathogens such as Yersinia pestis. The overall workflow showcased how synthetic biology, especially protein engineering and circuit design, can facilitate microbiota engineering for bio-therapeutical applications.Rather than relying on existing microbiota and molecular parts, with the advances in synthetic biology, we can design and engineer valuable regulatory parts as needed. Li and coauthors employed molecular dynamics simulation and the Molecular Mechanics Generalized Born Surface Area (MM-GB/SA) method to artificially designed transcription factors with different binding affinities . Chen and coauthors summarized impacts of the host-microbiome interactions on osteomyelitis pathogenesis, covering direct and indirect microbiome-associated osteomyelitis . Finally, Yang and coauthors reviewed the immune responses of severe COVID-19 in the gastrointestinal microenvironment and the gut-lung axis . This review highlighted the potential of using probiotics as an alternative drug to alleviate or even prevent severe COVID-19 outcomes.Several excellent reviews were also assembled for our readers in this topic. Zhang and coauthors provided a comprehensive review of the recent advances in understanding mechanisms of microbiomes as well as synthetic biology technologies that can be utilized to prevent, detect, and treat various diseases (We hope the articles and reviews collected in this research topic can show how fascinating this emerging field in the intersection of synthetic biology and microbiota is, and the great therapeutical potentials this field might offer. We want to take this chance to thank all the authors and reviewers for contributing to this special topic issue. As we are starting Volume II, we encourage and welcome more experts from this field to submit their research work to our topic.


Structural variants (SVs) are a major source of human genetic diversity and have been associated with different diseases and phenotypes. The detection of SVs is difficult, and a diverse range of detection methods and data analysis protocols has been developed. This difficulty and diversity make the detection of SVs for clinical applications challenging and requires a framework to ensure accuracy and reproducibility. Here, we discuss current developments in the diagnosis of SVs and propose a roadmap for the accurate and reproducible detection of SVs that includes case studies provided from the FDA-led SEquencing Quality Control Phase II (SEQC-II) and other consortium efforts.The online version contains supplementary material available at 10.1186/s13059-022-02636-8. Structural variants (SVs) are typically defined as genetic variants of greater than 50 base pairs (bp) in length and include insertions, deletions, duplications, and chromosome rearrangements . AdvanceAlthough great progress is being made in detecting SVs in the human genome and correlating this with phenotypic impact, accurately and precisely identifying SVs in specific samples and/or across samples is challenging . The preSVs are responsible for more nucleotide changes than any other genetic variants in humans and other species. Many technologies have been developed to identify different types of SVs in the past 15 years, from cytogenetic-based detection , array-based technologies , short-read high throughput sequencing , and linked-read sequencing  to long-read sequencing  , 19. AccThe accurate and reproducible detection of SVs underpins reliable clinical implementation and reduces the chance of misdiagnosis. Similar to the quality control steps made for SNVs and small indels , 21, iniThe Sequencing Quality Control Phase II (SEQC-II), led by the U.S. FDA, is the most current initiative to develop actionable best practice for sequencing data analysis. The aim is to define reproducible and accurate genetic variant calling to facilitate the development and regulation of precision medicine in clinical practice –32. In tIn this perspective, we discuss findings from these consortiums and identify gaps where current approaches for SV calling may be suboptimal, as well as propose potential solutions. Our analysis highlights three key components essential for an accurate and reproducible SV detection Fig. : (1) chaReference standards can be used to measure the false-positive and false-negative rate of SV calling . SpecifiWhen reference materials are designed to improve assay development for inherited disease diagnosis, patient-parent trio samples offer more sensitive identification of the de novo SVs and any underlying predisposition mechanism. Examples of such trio-based reference materials  were employed by the National Institute of Standards and Technology (NIST)  and the Unlike these two working groups focusing on somatic mutations, the SEQC-II germline mutation working group focuses on a comprehensive and reproducible assessment of the detection of germline variants based on a trio sample set. For this, sample materials are the HapMap trio from the 1000 Genome project  and HG00It is apparent that this is yet to be solved as many samples are “only” healthy cells derived from limited ethnic backgrounds. Especially in cancer, scientists are often interested in detangling complex SV that are chained, including virus insertions, somatic vs. germline mutations, low-frequency mutations escaping traditional diploid genomic patterns or even circularized DNA are not present in any of the reference materials yet. Several attempts have been made to develop reference material for somatic variants identification and NGS technology calibration in clinical laboratories. Specifically, the genome editing technologies such as CRISPR/Cas9 were adopted to biologically engineer mutation clones in mammalian cells , 43. HowOne approach could be to use tumor-normal or mixed cancer cell lines samples with different tumor purity and heterogenicity that may be closer to actual cancer patients. Cancer genomics aims to identify somatic (tumor-specific) variants with potential diagnostic, prognostic, and therapeutic implications and to detect germline variants with inherent information for both patients and their families. Although substantial clinical tumor testing does not currently involve analysis of a matched germline sample due to cost and time to delivery, concerns have been raised that the detected genetic variants are truly somatic variants . MoreoveSpecifically, the SEQC-II somatic mutation working group developed a tumor-normal reference sample using a normal B-lymphocyte cell line  and a triple-negative breast cancer cell line  from the same donor purchasable through the American Type Culture Collection (ATCC) . The HCChttps://www.fda.gov/medical-devices/in-vitro-diagnostics/list-cleared-or-approved-companion-diagnostic-devices-in-vitro-and-imaging-tools). To enhance the clinical application of panel-based sequencing for cancer diagnosis, we suggest it is indispensable to establish reliable, robust, continuous, and generally available genomics reference samples for assessing and calibrating different NGS assays.While large whole-exome and whole-genome sequencing studies are capable of providing new insights of genetic variants into cancers, clinical adoption of these approaches is still lagged and is not routinely offered by clinical laboratories . In conthttps://www.horizondiscovery.com/reference-standards/type/oncospan). To overcome the shortcoming, the SEQC-II Oncopanel working group synthesized sample A by equal mass pooling gDNA of ten cancer cell lines that were used to make the Universal Human Reference RNA (UHRR)  to ensure the wide coverage of actionable cancer genes under different VAF magnitudes [VAF of somatic mutation  is far less than the VAF in germline cells . It is technically challenging to utilize normal cell lines to establish a reference material covering a wide range of VAF magnitudes. Commonly available commercial reference samples typically consist of the limited number of genes and variants with distinct VAF ranges, hampering its utility for benchmark NGS assays in genetic variants with lower VAF (< 2~5%) and resulting in the overall variant detection performance is inversely correlated to the VAF of the analytes targeted  in inherited diseases and cancers has been widely reported , 55. TriMatrix effect-spiked reference materials. The SV detection limit within formalin-fixed, paraffin-embedded, or liquid biopsy-based samples from patients is much lower than that in normal tissues. Furthermore, some complex effect of fixatives such as in vitro fertilization (IVF) and preimplantation genetic diagnosis (PGD) screening help patients to select embryos free of rare diseases . As the Use genome editing for spiked specific SV types in the reference materials. Genome engineering technology with programmable nucleases  has been well-established, enabling precise and efficient genome-editing spiking of specific SV types into cells, and offering the opportunity to develop synthetic reference materials . For exaCommutability, defined as the ability of reference materials to perform comparably to actual patient samples, is one of the most critical parameters in qualifying reference materials . To estaAdvances in genomics technologies continually improve the resolution of SV detections , 18. FroAn increasing body of literature has demonstrated the benefits of employing multi-platform genomics technologies for more comprehensive SV detection across the human genome . ChaissoThe SEQC-II adopted multi-platform and multi-lab designs for a comprehensive assessment of reproducibility and accuracy of the detection of SVs. The SV working group set out to investigate the reproducibility and variability of SV calls when the sample was sequenced across multiple sequencing instruments or in different laboratories , 32. IntAlthough the promise of multi-platform genomics technologies for complete SV detection is apparent, it may be more appropriate to use a combination of different sequencing technologies; insight into their different strengths and weaknesses is key for effective deployment. The pros and cons of different genomics technology for SV detection were discussed extensively elsewhere , 17, 62.First, a multi-technology approach is not scalable given the costs and DNA, cell, or sample requirements such as quantity and quality for the clinical samples. One technology that may enable more accurate SV detection is known as long reads , 63. DesSecond, given the trend of reducing sequencing costs and improving yield as well as error rate, multiple long-read sequencing projects are underway from the assembly of individual genomes through the study of the genomic architecture of individual human cells . This trThere are multiple short read-based assays currently available. Linked reads constitute a fast and inexpensive method that provided information across a long molecule , leading to improvements in mappability and phasing. In theory, this approach could improve SV calling itself, but the software for achieving this was limited. Other interesting concepts such as HiC and StrandSeq also show promises. StrandSeq enables the accurate detection of inversions but requires laborious preparation, and there is not yet a standardized kit available. However, both technologies are limited by the mapping into repeats given their read length of 100–150 paired-end reads. Nevertheless, a combination of StrandSeq and PCR-free WGS may improve the detection of SVs at a cost and sample requirement that outcompetes long reads.Currently, WGS remains the workhorse of genomics, with million genomes sequenced every year. Therefore, we need to establish robust pipelines, ensure technical reproducibility, and understand the risk of bias. SEQC-II has so far approached this by sequencing samples across multiple centers to improve our understanding of potential variabilities and the impact on SV detection .The evolution of genomics technologies has also resulted in the proliferation of different SV calling algorithms. Over 85 publicly available SV calling algorithms have been developed for different NGS data types . These aThis work started over a decade ago. SV calling is improving but has by no means reached the reliability and quality of SNV calling. While multiple reasons may attribute to this problem, SV alleles are longer and often more complex than SNV which can be contained in a single-short read. Nevertheless, what steps can be taken to improve the field further or spark a new evolution of methodologies and approaches? Here we highlight some potential directions and discuss their promises. These are (1) extending the number and diversity of SV benchmark sets, (2) alignment vs. assembly vs. graph genome approaches, (3) single caller vs. consensus vs. genotyping approaches to improve SV precision, and (4) incorporation of machine learning and AI.Over the past few years, multiple methods have been proposed and implemented to simulate SV  have already been heavily sequenced and studied, but no official benchmark SV set is available. Cancerous but also other human disease genomes will be important as these include challenging regions  that are not trivial to resolve. Additionally, the need to include different ethnicities and the potential for novel sequences  suggests that there is still some way to go. In any case, the current existing benchmark sets are already showing an improvement in SV calling methods and methodologies and are also providing standards for wet lab technology development.There is often a debate about the best approach to identify SVs, either at scale (multiple hundred to thousand samples) or comprehensively (complex structures within a sample). Mapping  compared to de novo assembly  often has different advantages and disadvantages . In geneAlternatively, for assemblies, there are still risks of over-merging or splitting regions due to high  or low  heterogeneity. This is mainly due to the central paradigm of an assembly of when are two genomic regions the same and some sequencing artifact altered them slightly or if these are indeed two different regions. Prominent assembler for short reads  or evenAnother option that might be able to combine these advantages over time is a graph genome approach. Here multiple genomes  are first assembled into high-quality genomes and then utilized to improve mapping and thus variant detection. As one can imagine, if this graph genome carries a novel sequence or complex SV, it enables the detection of these in the analysis of other samples. However, there are apparent limitations such that there needs to be a balance between comprehensiveness  and complexity . While these problems are being addressed, graph genomes have already shown significant promises for SV calling: Paragraph  and VGToThe SV detection limit differs across different types , sizes , and genomic regions . SV calling algorithms were developed based on different heuristics, hypotheses, and thresholds, resulting in a considerable divergence. This is illustrated by comprehensive benchmark papers across SV callers , 26. BotThe use of consensus calls generated from multiple SV callers can achieve better precision and recall rate , 26. NevAnother less-studied area is the use of a SV genotyper after initial SV calling. Here, a SV genotyper uses a VCF with predefined SVs and summarizes the evidence for these SVs across a given sample or bam file. There are currently multiple SV genotyping methods available such as STIX, Paragraph, SVTyper, GraphTyper, and others. Their performance varies similar to SV callers over sizes and types of SV . Still, Machine learning algorithms especially neural networks have impacted almost every facet of our daily life and have revolutionized genomics –99. For The fundamental difference between conventional SV calling algorithms and deep learning approaches lies in retrieving the SV information from the BAM file. Traditional methods aim to detect the divergence between the sample sequence and reference at a per read level first. In contrast, deep learning approaches such as convolutional neural networks (CNNs) transform variant detection as a classification problem by considering the BAM file or its genomic regions as an image .A few initial attempts to apply deep learning algorithms for SV detection have been conducted, and some encouraging results were obtained –103. OneBesides SV detection, machine learning and AI may also be a good option for SV pathogenicity prediction to facilitate clinical application. Although NGS has tremendously improved the resolution of SV detection, it also poses a significant challenge to prioritize and pinpoint a small subset of SV that is clinically relevant. Therefore, accurately discerning the pathogenicity of the SVs identified through NGS testing is profound for its clinical adoption . The guiSome efforts have been made for pathogenicity prediction of SV, such as strategies by aggregation of SNP pathogenicity within the SV intervals  by integrating over 30 informative features correlating with SNV pathogenicity, to quantitively predict the pathogenicity of CNVs across various ethnic groups across approximately 93% of the human genome [Machine learning-based approaches by integrating different genome features and ethnic information across the population could effectively improve the prediction performance of SV pathogenicity in the whole genome scale. As part of the FDA-led SEQC II effort, we developed a novel machine learning-based framework X-CNV . Crowdsourcing efforts led by government agencies such as NIST and FDA will be tremendously helpful to prioritize fundamental and translational AI research consistent with regulatory priorities for robust, safe, secure, and privacy-preserving machine learning in different real-world applications. One example is PrecisionFDA, led by the U.S. FDA, which is a secure, collaborative, high-performance computing platform to advance precision medicine, inform regulatory science, and enable improvements in health outcomes. Several PrecisionFDA challenges have been launched, such as calling variants from short and long reads in difficult-to-map regions to standardize NGS testing in the precision medicine practice  [Although machine learning and AI shed light on SV detection and interpretation, more comprehensive evaluation and further investigation on the context of use are highly recommended towards a robust, secured, privacy-preserving, and explainable machine learning and AI solution for clinical application. The U.S. Food and Drug Administration (FDA) recently issued the “Artificial Intelligence/Machine Learning (AI/ML)-Based Software as a Medical Device (SaMD) Action Plan” to enable the FDA and manufacturers to evaluate and monitor a software product in a lifecycle-based regulatory framework for machine learning and AI technologies and allow for modifications to be made from real-world learning and adaptation (llenges) . The SEQAlthough NGS has tremendously improved the resolution of SV detection and facilitated our understanding of disease etiology and pathogenesis, substantial challenges remain for its full clinical implementation . The infThe strategies of promoting NGS towards its clinical implementation and adoption have been intensively discussed elsewhere , 121. OnThe potential high false-positive rate of NGS-based SV detection is one of the major concerns for its clinical applications compared to currently clinically accredited laboratory genetic tests. Since the NGS pipeline for SV detection involves multiple steps, a standard framework for managing and standardizing the NGS-based SV detection is urgently needed for clinical application. To fill the gap, a ClinSV was recently proposed to provide a “one-stop” solution for WGS based SV integration, annotation, prioritization, and visualization . The proThe accurate and reproducible detection of SVs is required for use in clinical applications. As a direct benefit of the rapid evolution of genomic technologies coupled with the work of different stakeholders, an increased appreciation of the contribution of SVs in genetic diversity and disease etiology continues to emerge , 124–126The field of SV detection continues to expand with advances in genome technologies, the establishment of and more reference standards and the release of high-quality calling sets. These resources allow us to further investigate some of the critical questions that remain and facilitate novel genomics technology development towards reliable and comprehensive SV identification (see Outstanding Questions). Notably, regulatory attention is on standardizing structural variant detection and on classification for convenient clinical adoption . We suggThe ongoing development of many genomics technologies sees a period of excitement, huge investment and perhaps disappointment that, in turn, may trigger a new wave of innovation. We foresee that more accurate and reproducible SV detection approaches will emerge soon, generating a more complete picture of the landscape of the human genome. Next steps will require different stakeholders to work towards a common goal of further resolving the difficulty in SV detection and accelerating clinical application.Additional file 1. Review history.