Early information on treatment response of HCC to local ablative therapy is crucial. Elastography as a non-invasive method has recently been shown to play a potential role in distinguishing between benign and malignant liver lesions. Elastography of hepatocellular carcinoma (HCC) in early response to local ablative therapy has not been studied to date.We prospectively included a cohort of 14 patients with diagnosis of HCC who were treated with local ablative therapy . We used 2D shear-wave elastography (RT 2D-SWE) to examine stiffness of HCC lesion before and 3, 30 and 90 days after local ablative therapy. Contrast-enhanced imaging after 90 days was performed to evaluate treatment response. Primary endpoint was stiffness of HCC in response to local ablative therapy. Secondary end point was tumor recurrence.Stiffness of HCC nodules and liver showed no significant difference prior to local ablative therapy. As early as three days after treatment, stiffness of responding HCC was significantly higher compared to non-responding. Higher stiffness before treatment was significantly associated with tumor recurrence.Nodule stiffness in general and RT 2D-SWE in particular could provide a useful tool for early prediction of HCC response to local ablative therapy. In recent years, liver stiffness assessed by different elastography techniques  has become a standard tool for non-invasive staging of liver fibrosis and detection of clinically significant portal hypertension in patients with chronic liver disease –3. PatieThis study prospectively included a total of 14 patients with HCC diagnosed by typical contrast-enhanced imaging (CT and/or MRI) and were referred to local ablative therapy by our institution’s interdisciplinary tumor board. HCC that were previously treated were excluded. In total, three patients were excluded . A flow chart of patient selection is shown in We included only lesions that were well visible in B-mode sonography in the right liver lobe for real-time two-dimensional shear wave elastography (RT 2D-SWE) . The region of interest was set according to the size of the evaluated lesion. The assessment was performed according to manufacturer’s recommendations. RT 2D-SWE examinations were performed in a fasting condition, supine posture and intercostal probe position. At least three viable measurements were taken and mean values in kPa were recorded. In addition, RT 2D-SWE of the non-HCC liver parenchyma was assessed according to manufacturer’s recommendations and with the region of interest set at a diameter of at least 15 mm, at least 2 cm away from the HCC nodule and at least 1.5 cm deep from the liver capsule.We examined RT 2D-SWE of HCC before local ablative therapy as well as three, 30 and 90 days after local ablative treatment. The lesion of interest was selected based on the planned treatment. Therefore, only lesions that were treated with local ablation were measured and included.The diagnosis of HCC was done with contrast-enhanced imaging (multidetector row CT or magnetic resonance imaging (MRI)). Commercially available clinical 3.0 Tesla MR imaging system  or a 1.5 Tesla MR imaging system  or CT imaging systems  were used. HCC was diagnosed only if typical imaging features were detected . Follow-Descriptive statistics were computed for all variables. Non-parametric testing was used to compare different groups when suitable. Paired non-parametric testing was used to compare data during follow-up of the same patients. Continuous variables are presented as mean ± standard error of the mean (SEM) or median (range). Categorical variables are presented as absolute cases or percentage. Distinguishing cut-off values was performed using receiver operating characteristics analysis. Time-to-Event analysis was performed using Kaplan-Meier curve with log-rank test. All data were analyzed using SPSS  and plotted using Prism .A total of 14 patients with HCC were included . Eight pAfter completion of the 3-months follow-up, the subsequent contrast-enhanced imaging revealed that 11 patients had complete local response to local ablative treatment , while 3Of the 14 patients, 8 developed tumor recurrence (Non-TL) during follow up at a median time of 220 days (82–371). Stratifying the patients for development of Non-TL showed no significant difference in general characteristics. Of note, the stiffness of the target HCC nodule before treatment was significantly higher in the patients without Non-TL compared to those who develop Non-TL .Two patients died during follow up. One due to gall bladder perforation 5 months after treatment and one due to progressive tumor disease 10 months after treatment.Median stiffness of the non-infiltrated liver before local ablative therapy was 28.2 kPa, which is above healthy liver stiffness and in line with diagnosis of liver cirrhosis. There was no significant difference between responding and non-responding patients . As expeComparing the patients with local response to those without, the stiffness of HCC nodules was not significantly different. However, the stiffness of the nodules of the non-responding patients tended to being higher  . InteresDue to this result, we aimed to identify a possible cut-off value to distinguish the patients who develop Non-TL with elastography of the HCC nodule. We therefore performed ROC analysis and found the optimal cut-off value at 17.5 kPa  . We thenRECIST criteria are still commonly used in tumor imaging and therefore we also rated the patients according to RECIST. As shown in the general characteristics in As early as three days after treatment, nodule stiffness of the RECIST responders remained stable, while HCC stiffness of RECIST non-responders increased, rendering the difference between response and non-response statistically significant . SimilarProbably due to the small sample size, none of the assessed parameters were significantly and independently associated with non-response, including, in particular, treatment modality, evolution of elastography, and severity of disease.This study provides first data of non-invasively measured HCC stiffness in response and outcome to local ablative therapy. We not only demonstrate that stiffness before treatment might distinguish patients at risk of developing Non-TL but also show that an increase in stiffness as early as three days after therapy could be useful in predicting outcome of local ablative therapy according to RECIST criteria.Local recurrence and residual tumor mass in local ablative therapy is more frequent compared to surgical resection . TherefoElastography has been shown to be useful in distinguishing between benign and malignant focal liver lesions ,8,18,19.It has been shown that some HCC have microvascular invasion and/or micrometastases ,23. DepeOne might argue that after RFA mostly the treated HCC nodules are completely ablated and the lesion after treatment therefore only consists of necrosis . NeverthImportantly, since—according to the MELD score—non-responders had an even better liver function than responders, it is unlikely that HCC stiffness was due to an overall increased liver stiffness in these patients (which was similar). Rather, this was due to the mechanical and biological properties of the nodules.This work has several limitations, mainly the small sample size and therefore serves more as a proof-of concept study. Another limitation of the study might be a variability in measurements depending on a different of factors like volume status of the patients. Of note, in our study, the HCC-to-liver stiffness ratio did not provide significant predictive value, which is in line with previous studies  and undeNon-invasive evaluation of stiffness of HCC nodules before and after local ablative treatment is feasible and an easy and promising tool to predict response long before the usual contrast enhanced imaging.


To present the rates of referral of participants in the National Eye Health Survey (NEHS) for further eye care.A national sample of 3098 non-Indigenous Australians aged 50–98 and 1738 Indigenous Australians aged 40–92 years living in 30 randomly selected sites was recruited using a door-to-door approach. Participants completed a general questionnaire and a series of eye tests, including vision and anterior segment assessment, intra-ocular pressure measurement, visual field testing and fundus photography. A predefined protocol was used to guide the referral of participants for follow up eye care. An ophthalmologist was on-call to assist with the triaging of participants.Of the total sample, 32.1% (994/3098) of non-Indigenous participants and 43.6% (757/1738) of Indigenous participants were referred for further eye care (p<0.001). A significant difference in referral rates for Indigenous Australians was observed between regions of differing geographic remoteness . After adjusting for covariates, males , older age  and longer time since previous eye examination  were associated with higher rates of eye care referral in the non-Indigenous population. In the Indigenous population, older age , self-reported diabetes , greater geographical remoteness  and longer time since previous eye examination  were associated with a higher rate of referral after multivariate adjustments. A total of 25 participants (1.4%) were referred for urgent follow-up of potentially sight threatening conditions.Our data has identified several high risk groups that required ophthalmic referral including older Australians, non-Indigenous men, Indigenous Australians with self-reported diabetes and those residing in very remote populations who may benefit from improvements in the provision and/or uptake of eye health services. Future longitudinal research is warranted to evaluate the feasibility and efficacy of implementing a referral protocol within a population-based research setting. In 2012 the World Health Organisation (WHO) reported that more than 300 million people worldwide were vision impaired or blind [Most population-based eye health studies either fail to include referral guidelines in their study protocols, or do not report referral rates or the reasons for referral –8. Only The implementation of a referral protocol in a research study requires adequate resources including personnel, clinical expertise and governance systems. Whilst a population health study should not be misconstrued as the provision of health screening or health care, the detection of disease or individuals at risk of disease in the course of a study should, in our view, warrant timely referral. This is particularly relevant in ophthalmic research given that the large majority of eye diseases are asymptomatic in their early stages and vision loss is often treatable or avoidable through appropriate interventions . This paThe protocol for this study was approved by the Human Research Ethics Committee of the Royal Victorian Eye and Ear Hospital (RVEEH) (HREC-14/1199H), the Aboriginal Health and Medical Research Council of New South Wales (HREC-1079/15), the Aboriginal Health Council of Western Australia (HREC-622), the Menzies School of Health Research (HREC-2015-2360) and the Aboriginal Health Council of South Australia (HREC-04-15-604). This research was conducted in accordance with the Declaration of Helsinki.The selection of recruitment sites was based on the Australian Statistical Geography Standard (ASGS), developed by the Australian Bureau of Statistics (ABS) to report 2011 Census data. Thirty sites, across five Remoteness Areas , were selected using a multi-stage, random cluster sampling methodology. To obtain a nationally representative sample of the population, 100 non-Indigenous Australians aged 50 years and older and 50 Indigenous Australians aged 40 years and older were to be recruited at each site. Recruiters went door-to-door to determine the eligibility of the residents. All eligible residents were invited to participate. Engagement of local Aboriginal health workers and community elders played a central role in the recruitment of Indigenous participants.Clinical examinations took place in a venue that was within 6km of the target recruitment area. The testing protocol took approximately 30 minutes per participant.All participants provided informed, written consent prior to examination. Each participant underwent an interviewer-administered questionnaire to collect information on socio-demographic factors, history of ocular problems, stroke and diabetes. Participants then underwent a series of eye tests, administered by trained eye examiners.Presenting distance visual acuity (VA) was measured using a logMAR chart . If presenting VA was worse than 6/12  in either eye, vision was retested with a pinhole occluder. Auto-refraction was performed using a hand-held auto-refractor/keratometer  to objectively measure refractive error of participants whose VA improved with pinhole to ≥6/12. Presenting near vision was assessed using the CERA E near vision card at the participant’s preferred reading distance [Anterior segment assessment of both eyes was conducted using a hand-held slit lamp  at 10x magnification for the presence of pterygium and lid abnormalities. Trachoma grading was conducted using the WHO Trachoma Simplified Grading System  in IndigThe N-30-5 screening protocol of the Frequency Doubling Technology (FDT) perimeter  was used to assess visual field loss. If the sensitivity was reduced in any of the field locations, the test was repeated to determine the reproducibility of the defect and the best result was graded. Intraocular pressure (IOP) was measured using a tonometer .Two-field, forty-five degree colour fundus photography was performed using a non-mydriatic fundus camera  to assess the retina and the optic disc. If retinal photographs of reduced quality were due to small pupil size (<3.00mm), the pupil was dilated with tropicamide 0.5% and photographs were retaken. Anterior chamber angle depth estimation was performed with a hand-held slit lamp using the Van Herick grading system, prior to pupil dilation to exclude participants at risk of anterior chamber angle closure .At the completion of the examination, each participant was provided with verbal feedback on the health of their eyes. This included a discussion of VA, IOP, visual fields as well as anterior and posterior ocular health. A referral protocol was developed by study investigators in conjunction with ophthalmologists . ParticiReferrals were made by optometrists, orthoptists or medical doctors within the research team. A consultant ophthalmologist was available at all times via telephone to provide advice on clinical findings and to triage referrals when necessary. Records of referrals for Indigenous participants were provided to the corresponding local Aboriginal health services with the consent of the participants.Statistical analyses were performed with Statistical Package for the Social Sciences (SPSS) . A p-value of 0.05 was used for significance testing. Descriptive statistics were calculated and Kolmogorov-Smirnov or Shapiro-Wilk significance statistic was used to assess the normality of the distribution of scores with a non-significant result (p = ≥0.05) indicating normality. Chi-square tests were utilised to compare proportions and an independent-samples t-test was used to compare continuous variables of interest between; 1) referred and non-referred participants, 2) referred Indigenous and referred non-Indigenous participants. Chi-square tests were utilised to compare referral rates between the 5 Remoteness Areas for Indigenous and non-Indigenous populations. Multivariable logistic regression analysis was used to examine the association between referral status and all covariates that were significant in univariate analysis.A total of 4836 individuals were recruited and examined in the NEHS, including 3098 (64.1%) and 1738 (35.9%) non-Indigenous and Indigenous Australians, respectively. Almost one-third  of non-Indigenous participants  in the study were referred for further eye examination and management. In contrast 43.6% (757/1738) of Indigenous participants  were referred (p<0.001) . A signiIn the non-Indigenous population, adjusted analysis revealed that males , older age  and longer time since previous eye examination  were associated with higher rates of eye care referral. For Indigenous Australians, older age , self-reported diabetes , greater geographical remoteness  and longer time since previous eye examination  were associated with higher rates of eye care referral after multivariate adjustments .Over three quarters of referrals for non-Indigenous  and Indigenous  participants were recommended for one month , while aThe reasons for referral were grouped into 5 categories: repeatable visual field loss, high IOP (>21mmHg), anterior segment abnormality, posterior segment abnormality and non-adherence to recommended guidelines for general or diabetes ocular examinations . The thrOf the 25 participants referred for urgent follow-up, 12 (48%) presented with an IOP of >30mmHg, 10 (40%) presented with symptoms consistent with possible retinal tears (persistent photopsias of recent onset), 2 (8%) showed signs of uveitis, and 1 (4%) participant had presumed periorbital cellulitis.Of those participants who required semi-urgent (1 week) referrals (n = 153), the five primary reasons for referral were: presenting vision less than the Australian driving standard (6/12) in both eyes ; a recent onset of unilateral vision loss (VA<6/60) ; retinal hemorrhage , and elevated IOP (≥21mmHg) with an enlarged vertical optic cup to optic disc ratio (≥0.8) .The NEHS is a population-based study conducted between March 2015 and April 2016 that aimed to determine the prevalence of vision impairment and blindness in Australia. Herein we have described the rate of referral of NEHS participants for further eye care and identified risk factors for referral.The overall referral rate for the study was high, with more than 1 in 3 participants being directed to a local eye health or primary care service for follow-up assessment. The significance of this finding must not be overstated as the vast majority of referrals were due to non-urgent reasons, such as poor compliance to recommended guidelines for general or diabetic ocular examinations. Of note however, approximately 10% (178/1738) of the referrals made by NEHS examiners recommended urgent (same day/next day) or semi-urgent (within one week) follow-up, accounting for 3.9% of the total sample. Urgent and semi-urgent referrals represent undetected and potentially sight-threatening ocular conditions that were opportunistically detected. It can be speculated that had these individuals not participated in the NEHS their ocular disease may have gone undetected, particularly for those who had never had an eye examination. It is not easy to put these referral rates into context given the paucity of similar data from other population-based eye studies. The urgent referral rate in the current study (1.4%) was lower than that reported amongst homeless adults in the Canadian city of Toronto of 8% [With the exception of non-adherence to recommended guidelines for general or diabetic ocular examinations, the main reason for referral of participants was the finding of suspicious retinal or optic nerve pathology. Population studies have demonstrated that the prevalence of asymptomatic retinal abnormalities increases with age . While tIndigenous Australian participants 757/1738, 43.6%) in the NEHS were more likely than non-Indigenous Australian participants  to be referred for follow-up and this was the case for all Remoteness Areas studied. It appears that this finding can be largely explained by a higher prevalence of self-reported diabetes (Indigenous = 37.1% vs. non-Indigenous = 13.9%) coupled with poorer adherence to recommended guidelines for diabetes ocular examinations in Indigenous Australians. A lack of awareness of the more stringent recommendation for annual diabetic retinal screening for Indigenous Australians, as opposed to biennial screening for most non-Indigenous Australians, may account for the sub-optimal compliance to these guidelines 38, 43.6%–22. IndiReferral rates differed significantly for Indigenous study participants by Remoteness Areas (range = 39.1%–60.4%) but not for non-Indigenous participants (range = 30.2%–35.7%). Notably, referral rates for Indigenous participants in very remote sites were approximately 1.5 to 2 times higher than those of Major City and Inner Regional sites. While referral rates may not directly correlate with the burden of eye disease within communities, these findings may be a reflection of the well-established barriers that Indigenous Australians face in accessing specialist services in remote areas, including communication, distance, and cultural inappropriateness ,26. To cA strength of the NEHS is its representativeness, achieved through the use of population-based sampling stratified by remoteness. The use of a predefined clinical referral protocol by qualified eye health professionals has allowed the study to provide useful insights into the prevalence of referable eye disorders and vision impairment. Nevertheless, the results of the current study must be considered in light of two key limitations. Firstly, the clinical referral protocol implemented in this study has not been validated and due to time and logistical constraints it was beyond the scope of the study to obtain access to the outcomes of follow-up for referred participants. This precludes appraisal of the appropriateness of the referral criteria, including false-positive referral rates. The social costs  and indiOur data has identified several high risk groups requiring ophthalmic referral who may benefit from improvements in the provision and/or uptake of eye health services, including; older Australians, non-Indigenous males, Indigenous Australians with self-reported diabetes and those residing in very remote communities. Whilst we are not in a position to assess the impact of the referrals that were made on the eye health of participants, we hope that participants were assisted by this approach. Future longitudinal research is warranted to further evaluate the feasibility and efficacy of implementing a referral protocol within a population-based research setting.


Ku-Shen (Radix Sophorae Flavescentis). Here, the network target of matrine was firstly predicted by drugCIPHER, a genome-wide target prediction method. Based on the network target of matrine, we performed a functional gene set enrichment analysis to computationally identify the potential pharmacological functions of matrine, most of which are supported by the literature evidence, including neurotoxicity and neuropharmacological activities of matrine. Furthermore, computational results demonstrated that matrine has the potential for the induction of macropinocytosis and the regulation of ATP metabolism. Our experimental data revealed that the large vesicles induced by matrine are consistent with the typical characteristics of macropinosome. Our verification results also suggested that matrine could decrease cellular ATP level. These findings demonstrated the availability and effectiveness of the network target strategy for identifying the comprehensive pharmacological functions of natural products.Comprehensively understanding pharmacological functions of natural products is a key issue to be addressed for the discovery of new drugs. Unlike some single-target drugs, natural products always exert diverse therapeutic effects through acting on a “network” that consists of multiple targets, making it necessary to develop a systematic approach, e.g., network pharmacology, to reveal pharmacological functions of natural products and infer their mechanisms of action. In this work, to identify the “network target” of a natural product, we perform a functional analysis of matrine, a marketed drug in China extracted from a medical herb  Structurally diverse compounds from natural products have historically been used as therapeutic agents for disease control and prevention and as a fertile source of lead compounds for the development of novel drugs . Howeverin silico approach, have provided big biological data of natural products toward network pharmacology, yielding important insights into pharmacological actions  into:The potential targets were predicted by the drugCIPHER-CS method . The drudj is the known drug j binding to the given protein p. β′p and α′pdj are the model coefficients. The likelihood between matrine d and protein p is defined as concordance score:where According to the concordance score of each target protein for matrine, drugCIPHER-CS prioritizes the proteins in the PPI network, and the candidate proteins with high concordance score are hypothesized to be predicted targets of matrine with high confidence.As the top 100 predicted targets of drug prediction can achieve the high prediction accuracy (77.3%), the network target of matrine is composed of the top 100 predicted targets of matrine in the protein–protein interaction network . To gainTM Green Dextran and Alexa Fluor®488 Phalloidin were purchased from Invitrogen . EIPA was purchased from Tocris. HepG2, PLC, HCT-8, HCT-116, HT29 and DLD-1 were obtained from ATCC .Matrine and amiloride hydrochloride was purchased from Sigma . Lucifer yellow (LY), pHrodoHuman colon cancer DLD-1 cells were exposed to 1.25, 2.5 and 5 mM matrine for 12 h and then fixed, dehydrated, and infiltrated for Transmission Electron Microscopy (TEM) as described previously . Ultrath2 at 37°C. The chamber was placed on the stage of a Nikon Eclipse Ti inverted microscope, equipped with a digital camera and Slide-book software . The software was set to automatically acquire phase-contrast images every 1 s for the indicated period of time.DLD-1 cells  were plated in a 35-mm glass-bottom microwell culture dish. One day after plating, cells were treated with 5 mM matrine in RPMI-1640 medium with 10% FBS. The dish was immediately placed in a humidified Live Cell chamber  equilibrated with 5% COTM Green dextran (50 μg/ml in the same medium) for 5 h and 10 h in a 37°C, 5% CO2 incubator, respectively. The tracer was removed and the cells were washed once or twice with the medium. Phase-contrast and fluorescent images of the living cells were acquired on a Nikon Eclipse Ti fluorescence microscope with a digital camera and NIS-Elements AR software .Labeling of endocytic compartments with these fluid-phase tracers was performed as previously described . BrieflyFor colocalization experiments, DLD-1 cells that had been treated with matrine were prepared for immunofluorescence microscopy as described previously . PhalloiTo inhibit macropinocytosis, DLD-1 cells were washed twice with PBS and then pretreated for 30 min with RPMI-1640 + 0.5% BSA in the presence or absence of 50 μM EIPA or 4 mM Amiloride. Following pretreatment, matrine was added to the dishes at a final concentration of 5 mM and phase-contrast images were acquired 5 h later.t-test was performed to determine the statistical significance. P < 0.05 was considered significant.At least three independent experiments were conducted in each study. The values are expressed as the means ± SEM. A two-tailed Ku-Shen that has not only been experimentally shown to cause a neurotoxic response, but has been shown to exert neuropharmacological activities, e.g., antinociceptive effects, hypothermic actions, neuroprotective role, antiepileptic effects, sedative effects, memory enhancement and sleep improvement. In addition, matrine may serve other pleiotropic functions, such as immunological regulation, anti-inflammatory activity, cardiac effects and anti-tumor effect .Actually, matrine is one of several alkaloids in Figure 1B).It is true that bioactive natural products can essentially regulate a molecular network by binding multiple targets, which may imply possible multiple effects. Therefore, to obtain a comprehensive functional characterization of matrine, we firstly validated the reliability of the predicted molecular network regulated by matrine. Here, we searched for reported molecular mechanisms related to matrine from Pubmed and CNKI database by the way of literature mining. The predicted targets of matrine can be connected to its reported biomolecules by direct or indirect targeting  . The resKu-Shen is reported to cause toxicity and side effects, including salivation, tachycardia and abnormal gait  as well as the transmission of nerve impulses (P = 2.92E-15). The results of literature validation suggested that many enriched biological processes have been reported in the literature (Table 1). Further, to evaluate this enrichment results, we manually collected 101 literatures related to biological processes of matrine. The results showed that 90.1% of reported biological processes were covered by enriched biological processes of the network target of matrine . The representative biological functions of matrine are summarized in Table 1 and Figure 2B. Anti-tumor effect, anti-viral effect, lipid metabolism, cardiovascular and blood pressure-lowing effect in the enriched biological functions of predicted targets of matrine are completely supported by literature evidence. Other important enriched biological functions of matrine can also be partly validated. These results suggested that featured as high-throughput, computational and capable of rapid analyses, our network pharmacology approach would have a strong potential to be applicable to other natural products for precise and comprehensive functional characterization.To explore comprehensively functional annotations of the matrine, all of enriched terms, regardless of Figure 3A). The structure of this functional network shown that known and novel GO BP functions share the same molecular mechanisms. Despite being a drug with well-characterized pharmacological actions, matrine has never been previously linked to ATP metabolism and membrane organization/endocytosis. To verify the effect of matrine on the ATP metabolism, we evaluated the ATP levels in the DLD-1 cells treated with matrine by a well-established assay for the levels of ATP. Following treatment with matrine, we observed the decrease of cellular ATP level in a concentration-dependent manner as shown in Figure 3B.Multiple and complex actions of natural products can be described in terms of molecular networks capturing the intricate web of connections among their targets . To furtFigure 3C). Then, we examined whether induction of cytoplasmic vacuolization by matrine is cell-specific. Therefore, we performed the same treatment experiments in various cancer cell lines. Strikingly, we observed more significant morphological changes with increasing concentrations of matrine in the DLD-1 than other cancer cell lines .Based on the molecular network regulated by matrine, we tried to infer the response of different cancer cell lines to endocytosis induced by matrine. We extracted the mRNA expression data of hepatocellular and colorectal cancer cell lines from the Cancer Cell Line Encyclopedia . The resIn fact, previous studies have demonstrated that matrine increases the cell volume and induces the formation of abundant cytoplasmic vacuoles in the SGC-7901 human gastric cancer cells and human hepatoma G2 cells , but howFigure 4A. The results suggest that high expression of these predicted targets of matrine in DLD-1 cells is associated with biological response to matrine. Therefore, observation of DLD-1 cells with standard phase contrast optics and a transmission electron microscope revealed that the biological processes initiated by the addition of matrine at increasing concentrations (1.25–10 mM) to DLD-1 cells within 24 h involve massive membrane ruffling and blebbing as well as cell rounding, followed by the formation of vacuoles that increase in number and size in a concentration-dependent manner . 5 mM matrine was selected for further study because this concentration yielded the most significant number of vacuoles. Time-lapse phase-contrast microscopy was also used to capture the dynamic changes of cytoplasmic vacuolization over time within 6 h and found that nascent vesicles could fuse with each other to form progressively larger vacuoles within the cytoplasm . The results confirm the induction of endocytic-like activity by matrine. Since macropinocytosis is defined as the formation of large endocytic vesicles of irregular sizes and shapes by cells that avidly incorporate extracellular fluid . To further confirm that the matrine-induced vacuoles observed by phase-contrast microscopy were indeed derived from macropinosomes, DLD-1 cells were incubated with the tracer LY during the first 5 h after the addition of matrine. As shown in Figure 5B, LY was also incorporated into most of the phase-lucent vacuoles. After fixing and staining with rhodamine-phalloidin, phalloidin counterstaining on DLD-1 cells shows that matrine disturbed the actin filament cytoskeleton of DLD-1 cells . Macropinocytosis can be distinguished from some types of endocytosis by its susceptibility to EIPA and amiloride (inhibitors for Na+/H+ exchangers). We investigated the effects of EIPA and amiloride on macropinocytosis in DLD-1 cells. Figures 5D,E show that the cytoplasmic vacuolization during the 5 mM matrine treatment was markedly inhibited by 50 μM EIPA and 4 mM amiloride, respectively. We concluded that the origin of a substantial portion of the cytoplasmic vacuoles was macropinocytosis. These results demonstrated that macropinocytosis was involved in matrine-induced vacuolization and verified the efficiency of our computational method.Macropinocytosis is known to be a form of actin-dependent endocytosis that leads to the internalization of fluid and membranes into large vesicles as macropinosomes. Next, we tested whether the large vesicles induced by matrine are consistent with the typical characteristics of macropinosome. As we know, rapid incorporation of extracellular-phase fluid tracers is a hallmark of macropinosomes . TherefoMacropinocytosis plays an important role in multifunctional biological processes, such as nutrient supply and non-apoptotic death of tumor cells .Experimental methods for target identification of natural products have had substantial success, yet many limitations still remain, including a high false-positive rate and bioactivity changes in labeled ligands . In receKu-Shen, have been approved as drugs, including matrine, oxymatrine, and sophoridine. In fact, the biological activities of matrine are more relevant to other applications of Ku-Shen other than anti-tumor activities. It is reported to possess a variety of pharmacological activities, including antipyretic , and provided experimental evidence to demonstrate the availability of a network-target-based approach. The network target regulated by matrine provides evidence for the diverse functions of matrine in Ku-Shen and shows that matrine can induce macropinocytosis in cancer cells and can decrease cellular ATP level. Our general network-based approach to fully elucidate comprehensive functions of matrine can be applied to many other natural products for the identification of their pharmacological functions.In summary, we developed a network target strategy for comprehensive functions of matrine in a Chinese medical herb, Sophorae Flavescentis Radix (SL and BZ designed and supervised the study. BZ, S-YW, MW, and YL performed the experiments, obtained and analyzed the data, and did the statistical analysis. XW and SL performed the computational analysis. All authors discussed the results and wrote the manuscript.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. The reviewer TL and handling Editor declared their shared affiliation.


Transposable elements comprise a huge portion of most animal genomes. Unlike many pathogens, these elements leave a mark of their impact via their insertion into host genomes. With proper teasing, these sequences can relay information about the evolutionary history of transposons and their hosts. In a new publication, Larson and colleagues describe a previously unappreciated density of long interspersed element-1 (LINE-1) sequences that have been spliced (LINE-1 and other reverse transcribing elements are necessarily intronless). They provide data to suggest that the retention of these potentially deleterious splice sites in LINE-1 results from the sites’ overlap with an important transcription factor binding site. These spliced LINE-1s  lose their ability to replicate, suggesting they are evolutionary dead ends. However, the lethality of this splicing could be an efficient means of blocking continued replication of LINE-1. In this way, the record of inactive LINE-1 sequences in the human genome revealed a new, though infrequent, event in the LINE-1 replication cycle and motivates future studies to test whether splicing might be another weapon in the anti-LINE-1 arsenal of host genomes. A piddling portion of most animal genomes encodes proteins. Instead, much of the DNA real estate in the genome of humans and other animals was built by the replication of selfish genetic elements. Selfish elements are stretches of DNA that encode the information necessary to replicate their own sequence in the context of a host cell , 2. By dIn humans, a single group of copy-and-paste transposons called the long interspersed elements-1 (LINE-1s) is responsible for most transposition events  do not manifest in nature. One could certainly imagine that LINE-1 sequences are spliced at a low enough rate that splicing presents no real detriment to the elements. Alternatively, as proposed by Larson et al., a deleterious sequence could be retained because in another context that same sequence provides an outweighing benefit. Here, the authors show that the splice donor site in one of the SpIRE forms overlaps precisely with the binding site of a transcription factor, RUNX3, which drives transcription of LINE-1—an apparently beneficial function . In thisLarson and colleagues proceed beyond a description of SpIREs. They use molecular biology to better understand the consequences of splicing to LINE-1s, in essence trying to mechanistically understand the historical events recorded by the human genome. First, the researchers describe a previously unrecognized SpIRE that is 10 times more abundant in the human genome than previously described SpIREs. They proceed to show that the transcripts from full-length LINE-1 sequences are spliced at very low levels; the vast majority of transcript remains unspliced, and some SpIREs that can be found in the human genome are not produced in measurable quantities using in vitro transcription assays. The clear next question is whether these spliced forms of the LINE-1 sequence are capable of retrotransposition or whether these sequences may be evolutionary dead ends for LINE-1.The authors venture to measure the inherent transcription levels of various SpIREs using constructs in which the 5′-UTRs of spliced and unspliced LINE-1s were cloned upstream of a reporter gene; normally, the LINE-1 5′-UTR encodes promoter activity sufficient to drive transcription. While the full-length, unspliced UTR efficiently drives transcription, the spliced sequence drives about 30-fold less transcription than the unspliced version. Last, the authors show that SpIREs are severely impaired in their ability to retrotranspose. One SpIRE that transcribes at a low level compared to unspliced LINE-1s can still retrotranspose, but only at a similarly low level. When driven with a strong promoter, however, this SpIRE replicates at about half the level of the unspliced sequence, suggesting the primary block to this element’s activity derives from low expression. These data demonstrate that even though spliced LINE-1 transcripts can complete retrotransposition, the resulting SpIREs are likely evolutionary dead ends, incapable of completing the next round of retrotransposition with any appreciable frequency.As a testament to the power of studying the LINE-1 fossil record, Larson and coauthors provide a compelling case for the series of events that led to the birth of a new SpIRE. The authors utilize constructs generated by Jacobs and colleagues that contain or lack a 129-bp stretch of DNA shown to bind ZNF93, a restriction factor that initiates silencing of transcription  18]. El. El18]. In a very cool final experiment, the authors show that a construct that still contains the 129-bp ZNF93 binding sequence produces a previously undescribed SpIRE and that 9 copies of this new SpIRE can be found in the human genome. The authors reason that adaptation to host repression (the 129-bp deletion) should have altered the spliced forms of LINE-1. With these data, they demonstrate that these altered spliced transcripts are produced in vitro, and they (amazingly) show that indeed the in vitro analysis reflects what happened in nature because these variant splice forms can be found in the human genome.In nature, successful reproduction requires propagation of the information necessary for replicating. The act of copying is alone insufficient : the new copies must be able to copy themselves. For selfish genetic elements, hosts encode specific mechanisms to block the replicative ability of descendant copies. These include proteins like APOBEC3s that deaminate intermediates in the replication of pathogens like LINE-1 and infectious viruses . The newThe selection for deletion-containing LINE-1 sequences that evade ZNF93 seems to have altered the patterns and perhaps the frequency of splicing of these pathogens’ RNA. One could imagine that certain deletions would make LINE-1 more susceptible to repression by splicing. Alternatively, splicing could generate dramatic variation in the 5′-UTRs of these elements, some of which could have new functions; for example, some LINE-1 splice variants could delete sequences that the host targets to repress these elements, thereby creating a new escape variant of LINE-1. As always, this benefit would be weighed against the detriment of losing the beneficial aspect of this sequence, for example, transcription factor binding sites that increase expression. These studies of dead LINE-1 sequences in the human genome reveal the push and pull of sequence changes that provide both benefit and detriment. Future studies will advance the limits of our attempts to decode the mass of information written in genome sequences with the goal of better understanding the long-standing coevolution of LINE-1s and their hosts.


The use of anti-tubercular therapy (ATT) along with anti-retroviral therapy (ART) in human immunodeficiency virus-tuberculosis (HIV-TB) co-infected individuals could at times lead to paradoxical worsening due to an increase in the inflammatory activity due to immune reconstitution inflammatory syndrome (IRIS) in the eye. This is characterized by anterior and posterior segment inflammatory reactions which may occur in the form of serous retinal detachment. We describe a case where the use of intravitreal anti-vascular endothelial growth factor (anti-VEGF) agent led to resolution of the serous retinal detachment, which had failed to respond to other common modalities of treatment.An HIV-TB co-infected 18-year-old, male patient, who was started on ART and ATT developed IRIS in the form of worsening of serous retinal detachment around a pre-existent asymptomatic tuberculoma. The patient was initially treated with oral and topical steroids without a satisfactory response. Intravitreal bevacizumab was then tried for this patient. Serial fundus photos and optical coherence tomography (OCT) taken before and after treatment showed complete resolution of the serous retinal detachment with two doses of intravitreal bevacizumab.Intravitreal anti-VEGF agents may have a role in the reversal of serous retinal detachment, which occurs as a part of IRIS in HIV-tuberculosis co-infected individuals who have been started on anti-tubercular and anti-retroviral therapies. TB is one of the most common opportunistic infections in HIV patients; however, ocular TB in HIV patients is relatively rare. It can occur in the form of anterior or posterior uveitis, most commonly in the form of choroidal granulomas , 2. We iWe describe a case of serous retinal detachment that developed around an asymptomatic, pre-existent, tuberculoma as a complication of paradoxical worsening of IRIS, who was successfully treated with intravitreal bevacizumab.An HIV infected 18-year-old, male patient, with newly diagnosed sputum smear negative pulmonary TB and CD4 count of 22 cells/μl presented to us for routine ophthalmic examination, without any ocular complaints. The patient was not on ATT or ART at the time of presentation. On examination, his visual acuity was 20/20 in both eyes, and a quiescent anterior chamber. The fundus examination of the right eye showed a well-defined choroidal tuberculoma of 1.5 disc diameter in size in the posterior pole close to the inferior arcade, not involving the fovea  is a humanized monoclonal antibody against VEGF. We used intravitreal bevacizumab in two doses at 1 month apart for our patient, and there was a marked improvement in both the functional and anatomical outcomes without any deterioration in the next 12 months of follow-up period. In our case, reversal of the vascular permeability by intravitreal bevacizumab played a role in the reversal of the serous retinal detachment. Phase II trials of bevacizumab in HIV patients state that in well-controlled HIV patients, bevacizumab can be given safely, but data on immune-deficient individuals is still awaited and so its use in low CD4 counts remain questionable .Primary treatment for ocular TB remains steroids with ATT in non-HIV patients. But in HIV patients, ATT alone is given without oral steroids and ART is started at 2- to 3-week interval. With the advent of ART, there has been increased incidence of IRIS during the phase of immune recovery. ATT has also solely shown to cause IRIS, even in immune-competent patients. Treatment guidelines for paradoxical IRIS syndrome in HIV are empirical. Our patient developed IRIS in the form of increased serous fluid after being initiated on ATT and ART, and we document its resolution with intravitreal bevacizumab.


The NH2-terminal fragment of DSPP is highly glycosylated but not phosphorylated, whereas the COOH-terminal fragment (named “dentin phosphoprotein” or “DPP”) is highly phosphorylated but not glycosylated. These two fragments are believed to perform distinct roles in dentin formation. To analyze the functions of DPP in dentinogenesis, we created “Dspp-/-;DPP Tg mice”, which expressed transgenic DPP driven by a Type I collagen promoter but lacked the endogenous Dspp gene. We characterized the dentin of the Dspp-/-;DPP Tg mice using X-ray radiography, histology, scanning electron microscopy, double fluorochrome labeling, immunohistochemistry and in situ hybridization. Micro-computed tomography analyses revealed that at postnatal 6 months, the transgenic expression of DPP increased the dentin thickness of the Dspp-null mice by 97.1% and restored the dentin material density by 29.5%. Histological analyses showed that the Dspp-null mice manifested an abnormal widening of the predentin while the predentin in Dspp-/-;DPP Tg mice was narrower than in the Dspp-null mice. Scanning electron microscopy analyses showed that the dentinal tubules in the Dspp-/-;DPP Tg mice were better organized than in the Dspp-null mice. The double fluorochrome labeling analyses demonstrated that the dentin mineral deposition rate in the Dspp-/-;DPP Tg mice was significantly improved compared to that in the Dspp-null mice. These findings indicate that the transgenic expression of DPP partially rescued the dentin defects of the DSPP-null mice, suggesting that DPP may promote dentin formation and that the coordinated actions between DPP and the NH2-terminal fragment of DSPP may be necessary for dentinogenesis.Mutations in the dentin sialophosphoprotein (DSPP) gene cause dentinogenesis imperfecta. After synthesis, DSPP is proteolytically processed into NH Dentin is composed of an organic matrix that is primarily made of Type I collagen and a mineral phase that consists of hydroxyapatite crystals. During the formation of dentin, odontoblasts secrete a collagen-rich matrix termed “predentin”, which is not mineralized. As the precursor of dentin, predentin lies between the mineralization front and the cells; it is converted to dentin when hydroxyapatite crystals are laid down within and around collagen fibrils. During normal dentinogenesis, a rather uniform layer of predentin is maintained, suggesting that the rate of predentin formation must equal the rate of mineralization. The uniformity of these rates indicates that balanced mechanisms must be involved to control the site and rate of hydroxyapatite formation and growth. Likewise, in pathological conditions, the widening of the predentin layer must involve the incapacitation of a controlling mechanism. Dentinogenesis is a dynamic process involving multiple changes that occur within the window of time when predentin is formed and converted to dentin. Changes that are well documented include increases in collagen fibril diameters, secretion and removal of proteoglycans and deposition of acidic phosphoproteins at the mineralization front .Small Integrin-Binding LIgand, N-linked Glycoprotein) family [Dspp gene with dentinogenesis imperfecta in humans [Dspp gene loss with dental defects in mice [Dspp gene knockout mouse is hypomineralized (widening of predentin), giving rise to a phenotype similar to the manifestations of dentinogenesis imperfecta Type III in humans.During dentinogenesis, Type I collagen, secreted by odontoblasts, forms the undergirding that is mineralized in a highly controlled manner. In addition to Type I collagen, the extracellular matrix (ECM) of dentin contains a number of non-collagenous proteins (NCPs) and proteoglycans. Dentin sialophosphoprotein (DSPP) belongs to the SIBLING  family . Excludi) family . The impn humans , 5 and o in mice . The pre2-terminal fragment [n and (Asp-Pse)n [In vitro mineralization studies indicate that DPP is an important initiator and modulator for the formation and growth of hydroxyapatite crystals [in vitro experiments [in vivo investigations are needed to further elucidate the biological functions of this unique phosphoprotein and test these beliefs.DSPP is a large protein that is proteolytically processed to form the COOH-terminal fragment and NHfragment , 7. The fragment . The mossp-Pse)n , 9. Compsp-Pse)n . These scrystals –13. Otheeriments , 15, in 2-terminal fragment of DSPP encoded by the 5’ portion of the DSPP transcript exists in two forms: the core protein form known as “dentin sialoprotein” (DSP) [In vitro studies showed that DSP had little or no effect on the formation and growth of hydroxyapatite crystals [The NHn” (DSP)  and the n” (DSP) . DSP andn” (DSP) , in clean” (DSP) . In vitrcrystals  while th2-terminal fragment (including DSP and DSP-PG) of DSPP suggest that these molecular variants may perform different functions in biomineralization although they are derived from the same mRNA. The abundance of DSPP fragments, along with the scarcity of full-length DSPP in the dentin, suggests that the processed fragments of DSPP may be the functional forms directly involved in dentin formation. Previously, we showed that the blocking of DSPP processing by substituting Asp452 with Ala452 at the cleavage site of mouse DSPP leads to the functional loss of this protein in vivo, indicating that the posttranslational cleavage to convert DSPP into fragments is an activation event, transforming an inactive precursor to active fragments that are essential to the formation and mineralization of dentin [2-terminal fragment failed to rescue the dentin defects in Dspp-null mice [Dspp-null background. We observed that the transgenic expression of DPP partially rescued the dentin defects of Dspp-null mice.The remarkable chemical differences between the COOH-terminal fragment (DPP) and NHf dentin . More reull mice . In the 5’-TGGTGGTGCAAATCAAAGAA-3’, located on the DPP coding sequence, and reverse primer, 5’-CTGTCACTGTCACCATCACCATTAC-3’, located on the SV40 polyadenylation signal. While we obtained a total of fifteen Col1a1-HA-DPP founder mice (15 lines) from multiple pronuclear injections [Dspp-/- mice  to introduce the Col1a1-HA-DPP transgene into the Dspp-null background (lacking the endogenous Dspp gene). The mice expressing the Col1a1-HA-DPP transgene without the endogenous Dspp gene were referred to as Dspp-/-;DPP Tg mice. The primers used to identify the endogenous Dspp alleles and null alleles (containing the LacZ gene) were described in our previous report [Dspp heterozygous (Dspp+/-) mice were used as normal control mice since there was no significant phenotypic difference between the Dspp+/- and wild-type (Dspp+/+) mice as we and others have previously reported [Dspp-/-;DPP Tg mice, in comparison with those from the Dspp+/- and Dspp-/- mice to determine the effects of transgenic HA-DPP on the formation and mineralization of the dentin in the Dspp-null molars.The generation of transgenic mice expressing DPP under the control of a 3.6 kb Type I collagen (Col1a1) promoter has been described in our previous publication . The trajections , only ons report . The agereported , 20. We All the animals used in the present study were housed in a temperature-controlled vivarium on a 12–12 light cycle with free access to food and water. All animal procedures were performed in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals and approved by the Institutional Animal Care and Use Committee of Texas A&M University College of Dentistry .Dspp+/-, Dspp-/-, and Dspp-/-;DPP Tg mice at 3 and 6 months of age were examined by both plain X-ray radiography  and microcomputed tomography (μCT) system . For the μCT analyses, a low-resolution scan (20 μm slice increment) was performed for an overall morphological assessment of the molar regions in the mandibles. A high-resolution scan in 3.5 μm slice increments was performed on 6-month-old samples to examine the mandibular first molars. A morphometric parameter analysis was performed using the built-in μCT software system. The dentin thickness, dentin volume (DV), total tissue volume (TV), DV/TV, and dentin material density of the mandibular first molars were obtained at a threshold of 375. The data acquired from the high-resolution scans of four samples per group were used for the quantitative analysis.The mandibles dissected from the in situ hybridization, or picro-sirius red staining.For histologic analysis, the mandibles were fixed in freshly prepared 4% paraformaldehyde in phosphate-buffered saline (pH 7.4) at 4°C overnight and then decalcified in 15% ethylenediaminetetraacetate (EDTA) solution (pH 7.4) at 4°C for 5~14 days depending on the ages of the animals. The samples were embedded in paraffin using standard histological procedures. Serial sections were cut at a thickness of 5 μm and used for Hematoxylin and Eosin (H&E) staining, immunohistochemistry (IHC), 2-terminal fragment of DSPP) [For IHC analyses, the experiments were carried out using an ABC kit, a M.O.M. kit and a DAB kit  according to the manufacturer’s instructions. We employed polyclonal antibodies against DSP (the NHof DSPP)  at a dilIn situ hybridization was carried out to assess the mRNA level of Type I collagen (Col 1a1) in the mandibular molars of 3-week-old mice as described previously [eviously . The RNAFor picro-sirius red staining, the mandible sections were immersed in haematoxylin solution for 8 minutes to stain the nuclei and washed for 10 minutes in water. Then, the sections were stained in picro-sirius red for one hour, washed in two changes of acidified water, dehydrated in three changes of 100% ethanol, cleared in xylene and mounted. The structure and organization of collagen fibers in the dentin were examined under a polarized light microscope.Microphotographs were taking under an Olympus BX51 microscope equipped with an Olympus DP72 microscope digital camera interfaced with Olympus cellSens Standard 1.6 software .Double fluorescence labeling was performed to analyze the mineral deposition in the dentin in the mouse incisors, as we described previously , 23. BriFor the SEM analyses, the mandibles from 3-month-old mice were dissected and fixed with 4% paraformaldehyde in 0.1 M phosphate buffer solution (pH 7.4) at 4°C for 48 hours. MMA-embedded specimens were prepared as described above. After adjusting a suitable comparable position of the samples, sand paper was used to grind the acrylic block with an increasing order of grit fineness. These samples were then polished using micro cloth with Metadi Supreme Polycrystalline diamond suspensions of sizes 0.1 micron, 0.25 micron and 0.05 micron (Buehler). These samples were then washed using the ultrasonic washer and placed in the vacuum system for 2 days. The dehydrated specimens were sputter-coated with carbon and scanned with a backscattered electron detector in a FEI/Philips XL30 Field emission environmental SEM system . To assess the dentinal tubule structures, the carbon-coated samples were repolished, as described above. The surfaces were acid-etched with 12% phosphoric acid for 7 seconds, washed with 5% sodium hypochlorite for 35 minutes, coated with gold and palladium, and then examined by SEM.The data analysis was performed with a one-way analysis of variance ANOVA for multiple-group comparison. If significant differences were found with the one-way ANOVA, the Tukey-Kramer post-hoc test was used to determine which groups were significantly different from others. The data were expressed in terms of mean ± SD. (standard deviation). P<0.05 was considered as the statistical significant difference.2-terminal fragment of DSPP) in the Dspp+/-  mice and its absence in the Dspp-/- and Dspp-/-;DPP Tg mice .X-ray radiography showed that the mandibular teeth from 3-week-old Dspp-/- mice exhibited thinner dentin and enlarged pulp chambers compared with the Dspp+/- mice, and the transgenic expression of HA-DPP markedly improved the dental defects of the Dspp-/- mice  mice at 6-months of age. However, the transgenic DPP expression only slightly improved the dentin material density  of the Dspp-/- mice.We first performed plain X-ray radiography analyses to assess the tooth structure and morphology. Plain X-ray radiography revealed that the -/- mice . The mic-/- mice . The qua-/- mice  showed tDspp-/- mice. At the age of 3 months, the mandibular molars of the Dspp-/- mice displayed enlarged pulp chambers, reduced dentin thickness, and a widened predentin zone compared with those of the Dspp+/- mice; the structure of the dentin-pulp complex in the Dspp-/-;DPP Tg mice appeared to be similar to that of the Dspp+/- mice. Biglycan is an odontoblast terminal differentiation marker and is primarily localized in the predentin zone of the teeth [Dspp-/- mice had a widened zone of biglycan staining signals, whereas the Dspp-/-;DPP Tg mice displayed a biglycan distribution pattern similar to the Dspp+/- mice, suggesting that the widened predentin zone in the Dspp-/-;DPP Tg mice was restored to a level similar to that in the Dspp+/- mice  compared with the Dspp+/- mice  analyses were carried out to analyze the effects of the HA-DPP transgene on the dentin mineralization defects and the dentinal tubular structural abnormalities in the +/- mice . The tra Tg mice . The acirganized . The HA--;DPP Tg .In situ hybridization analyses showed that there was no apparent difference in the expression level of the Col1a1 gene in the odontoblasts among these three groups of mice  promotes the formation and growth of the hydroxyapatite crystals [in vivo data to elucidate the physiological functions of DPP in dentinogenesis. The findings that the introduction of transgenic DPP significantly improved the dentin formation in the Dspp-null mice confirmed the mineralization-promoting effects of DPP, consistent with the data in previous in vitro analyses.While the fact that the inactivation or loss of DSPP causes dentin defects has clearly illustrated its critical role in dentinogenesis, the exact mechanisms of how this most prominent dentin NCP controls the formation and mineralization of dentin are unclear. DSPP is cleaved into NHunctions . While icrystals –13, therDspp-null mice. Previously, we demonstrated that the transgenic expression of the full-length normal DSPP driven by the same Type I collagen promoter completely rescued the dentin defects in the Dspp-null mice. The partial rescue of Dspp-deficient dentin by transgenic DPP, rather than the full restoration of Dspp-deficient dentin abnormalities, could be attributed to the following two reasons: 1) The proper function of DPP may require the assistance of the DSPP NH2-terminal fragment; the latter may be necessary for proper packaging/folding of the former in the endoplasmic reticulum and Golgi apparatus and for the subsequent delivery of DPP to the correct site. DSPP is cleaved into the NH2- and COOH-terminal fragments by bone morphogenetic protein 1/tolloid-like metalloproteinases [2-terminal fragment of DSPP and other molecules serving as chaperons may assist in the proper folding of DPP. In the trans-Golgi network, DSPP is cleaved and activated into its functional forms by bone morphogenetic protein 1/tolloid-like metalloproteinases. 2) The second reason for the partial rescue may be due to the fact that the expression level of the transgenic DPP was not high enough. As the transgenic founders expressing high levels of DPP could not be passed to subsequent generations, we could not perform rescuing experiments with these mouse lines expressing higher levels of the transgene [The current study showed that the transgenic expression of DPP partially rescued the dentin defects in the teinases , 26. Theteinases . We enviransgene .The transmission electron microscopy studies showed that DPP binds to Type I collagen predominantly at the "e" band in the gap region . The binDspp-null mice. These data provide in vivo evidence that DPP may promote the deposition of hydroxyapatite crystals during the formation and mineralization of dentin, which is in agreement with the in vitro findings described in earlier reports. The partial restoration of Dspp-deficient dentin by transgenic DPP, rather than a full rescue, suggests that the balanced actions between the COOH- and the NH2-terminal fragments of DSPP may be required for dentin development.In summary, this study demonstrated that the transgenic expression of DPP significantly improved the dentin defects in


The 2013 ACC/AHA guideline recommended either no statin therapy or moderate-intensity statin therapy (MST) for intermediate risk patients—those with 5–7.5% 10-year risk and without cardiovascular disease (CVD), hypercholesterolemia or diabetes. The guideline further suggested that the therapy choice be based on patient-clinician discussions of risks and benefits. Since low-density lipoprotein particle (LDL-P) levels were reported to be associated with CVD independently of traditional risk factors in intermediate and low risk patients, we investigated the cost-effectiveness of using LDL-P levels to identify intermediate risk patients likely to benefit from initiating or intensifying statin therapy.We evaluated 5 care strategies for intermediate risk patients. These included the strategies suggested by the guideline: no-statin therapy and MST. We compared each of these strategies to a related strategy that incorporated LDL-P testing. No-statin therapy was compared with the strategy of MST for those with high LDL-P levels and no statin therapy for all other patients (test-and-MST). MST was compared with the strategy of high-intensity statin therapy (HST) for those with high LDL-P levels and MST for all other patients (test-and-HST). We also evaluated the strategy of HST for all. Costs (payer perspective) and utilities were assessed over a 5-year time horizon in a Markov model of 100,000 hypothetical intermediate risk patients.HST dominated all other strategies, costing less and—despite causing 739 more cases of diabetes than did MST—resulting in more quality adjusted life-years (QALYs). For patient-clinician discussions that would otherwise lead to the MST strategy, we found the test-and-HST strategy reduced costs by $4.67 MM and resulted in 134 fewer CVD events and 115 additional QALYs. For patient-clinician discussions that would otherwise lead to no statin therapy, we found that the test-and-MST strategy reduced costs by $3.25 MM, resulted in 97 fewer CVD events and 44 additional QALYs.The HST strategy was cost saving and improved outcomes in intermediate risk patients. For patient and clinicians concerned about the adverse events associated with HST, using LDL-P levels to target intensified statin therapy could improve outcomes and reduce costs.The online version of this article (doi:10.1186/s12872-016-0429-6) contains supplementary material, which is available to authorized users. Cardiovascular disease (CVD) continues to be a major cause of death in the United States. Despite an impressive ~30% decline in CVD-attributable deaths between the years 2000 and 2010, CVD still accounted for roughly 1 in 3 of all deaths in the United States in 2010  or percutaneous intervention [PCI]), statin-related adverse events , and the number of quality-adjusted life years (QALYs).We modeled a population that, based on traditional risk factors, had a 10-year CVD risk estimate with a flat distribution and a range of 5–7.5% (average risk of 6.25%). Elevated LDL-P levels were found to be associated with increased risk of CVD (HR = 1.40 per standard deviation) in a population with intermediate or low risk after adjustment for established risk factors, including standard lipids . We assuThe annual event rates—transition probabilities between states—are based on published literature Table . When neCosts from published sources Table  were infTreeAge Pro software 2015  was used for modeling. Costs and utilities for the base-case strategies were calculated using a half-year correction. The effect of varying input parameters was explored by deterministic sensitivity analyses in which input parameters were individually changed to the upper and lower values of their range. The ranges of input parameters were based on published literature where available, or were set to ±20% of base values for parameters without published ranges . When comparing these 2 strategies the test-and-MST strategy dominated the do-not-treat strategy Tables  and 4. TFor those patients and clinicians whose discussions lead to an MST decision, the alternative strategy evaluated was to treat those in the top decile of LDL-P levels with HST and treat the rest with MST (test-and-HST). The test-and-HST strategy reduced costs by 4.67 million dollars and resulted in 134 fewer CVD and 81 fewer revascularization events when compared to the MST strategy. Despite the occurrence of 74 additional cases of statin-induced diabetes diagnosis, the test-and-HST strategy improved overall outcomes by adding 115 QALYs compared with the MST strategy.We investigated the robustness of the model outcomes in a deterministic sensitivity analysis that changed each input parameter to its highest and its lowest possible values Tables  and 2 whWe also investigated the sensitivity of the model outcomes to the input parameters by varying the parameters in a Monte-Carlo simulation. That is, we simultaneously varied all input parameters by sampling their values from probability distributions chosen to reflect the uncertainty in the parameter estimates Table  and 2. TWe assessed the cost-effectiveness of 5 care strategies for patients at intermediate risk of CVD and found that treating all such patients with HST resulted in lower costs and fewer CVD or revascularization events than did any of the other strategies. Therefore, although the ACC/AHA guideline  suggesteSince the ACC/AHA guideline suggested that information from additional risk assessment could help inform statin initiation decisions for intermediate risk patients, we considered strategies that identified patients with elevated (top decile) LDL-P levels. Although patient-physician discussions may involve a rational comparison of costs and benefits, the actual weights that a particular patient assigns to the benefits, risks and costs are very much a matter of personal preference and, as such, cannot be determined analytically. However, when these discussions would benefit from additional information about a patient’s risk, an analytical approach can be used to assess whether providing that information is likely to be cost effective, which has been the motivation for our analysis. We found that for patients and clinicians, whose discussions would otherwise lead to either a do-not-treat or an MST decision, adopting a strategy that includes a more intensive statin therapy for patients in the top LDL-P decile could be less costly, result in more QALYs and lead to fewer CVD or revascularization events over a 5-year horizon.For intermediate risk patients and their physicians who would otherwise choose a do-not-treat or MST strategy, a decision to choose a more aggressive strategy would probably require compelling additional information about the patient’s risk level. Therefore, in the test and treat strategies, only those in the top decile of LDL-P would be treated. For this LDL-P level, even those with the lowest risk in the intermediate risk group (5% 10-year risk) would have a 10-year risk >7.5%—a 10-year risk that would put them above the risk threshold at which the ACC/AHA guideline suggested considering high-intensity statin therapy. After incorporating the LDL-P level into the risk prediction model, while some of the patients have greater than 7.5% risk based on the new information, some have less than 5% risk; our model incorporated this lower risk estimate into the costs and utilities calculations. However, the model did not suggest a change in the care strategy for these lower risk patients.Several studies have suggested that statins can be cost-effective at lower risk thresholds lower than those in the ACC/AHA 2013 guideline (see Deano et al.  for a reThis current study differs from the Pandya et al. study  in both To account for uncertainty in the estimates of base-case parameters, we investigated the effects of changing parameter values using probabilistic sensitivity analyses. One interesting result of these analyses was that although the test-and-MST strategy dominated the do-not-treat strategy most of the time, the test-and-MST strategy resulted with fewer QALYs than the do-not-test strategy in about 20% of the simulations. A deterministic sensitivity analysis indicated that for these 20% of the simulations, the utility value of being disease free while taking a statin pill daily was probably drawn from the low end of the probability distribution. Although the magnitude of the disutility for taking a statin was modest, the effect of this small disutility was amplified because ~10% of the patients in the test-and-MST strategy were taking a statin pill daily whereas almost no statins were used in the do-not-treat strategy other than patients who had a CVD or a revascularization event. The disutility of taking statins had no noticeable effect when we compared the MST strategy with the test-and-HST strategy because all patients used statins in both strategies. The effect of the disutility of a daily statin pill on reducing the cost-effectiveness of statin treatment in the primary prevention of CVD is consistent with other studies of the cost-effectiveness of statin in the primary prevention of CVD , 30–32. Because pill taking disutility can differ substantially from person to person, clinicians and patients should consider individual preferences selecting a care strategy. Pill taking disutility is based on questionnaires that assess personal preferences, and an accurate assessment of a small disutility may be influenced by the numeracy of the surveyed individuals . AlthougOne limitation of this study is that the model included several simplifying assumptions. For example, the outcome of a third CVD event was assumed to be fatal, all patients with newly diagnosed diabetes were assumed be treated with statins, and all statin-related adverse events were assumed to occur within the first year of taking statins. In general, these simplifications eliminated rare states that, because of their rarity, would have a small effect on the model outcome and have little published support for parameter estimates. We validated the internal consistency of the model by exploring the outcomes of extreme parameter values. However, we were unable to perform external validation by comparing the model outcomes to a real-life study because no outcome study of statin use in patients with estimated 5–7.5% 10-year risk of CVD has been published. Moreover, this study is limited to a primary prevention population with intermediate risk of CVD. Thus, we did not consider patient treatment strategies that could involve angiography-based plaque imaging risk assessment . AnotherIn conclusion, this study indicates that HST is the preferred care strategy in intermediate risk patients. For those patients and physicians who decide to avoid HST, LDL-P testing can assist in making statin treatment decisions and is likely to reduce CVD events and to be cost saving.


High-resolution electronic interface circuits for transducers with nonlinear capacitive impedance need an operational amplifier, which is stable for a wide range of load capacitance. Such operational amplifier in a conventional design requires a large area for compensation capacitors, increasing costs and limiting applications. In order to address this problem, we present a gain-boosted two-stage operational amplifier, whose frequency response compensation capacitor size is insensitive to the load capacitance and also orders of magnitude smaller compared to the conventional Miller-compensation capacitor that often dominates chip area. By exploiting pole-zero cancellation between a gain-boosting stage and the main amplifier stage, the compensation capacitor of the proposed operational amplifier becomes less dependent of load capacitance, so that it can also operate with a wide range of load capacitance. A prototype operational amplifier designed in 0.13- The internet of things (IoT) is a new paradigm, which connects any physical objects embedded with ambient computational intelligence to each other such that these objects can recognize others and exchange collected data ,2,3. WitCapacitive micromachined ultrasonic transducers (CMUTs) ,12,13, pA sufficient direct-current (DC) voltage gain can be generated by multi-stage amplifiers ,39,40,41In this paper, we present a gain-boosted two-stage operational amplifier, whose compensation capacitance size is less sensitive of load capacitance compared to the previously reported operational amplifiers. Compared to the PSS amplifier, rather than decreasing the first-stage gain , the proThe remainder of this paper is organized as follows. Miller compensation, which has been extensively used in integrated operational amplifiers, deals with stability issues in frequency response by introducing capacitor ensation ,44. The To avoid the unity gain frequency higher than An improved compensation technique was introduced by Ahuja. It utilizes the current transformer providing virtual ground to eliminate feed-forward path in Miller compensation , as showthan (4) . The uniCompared to the conventional Miller compensation, a smaller compensating capacitance is required to drive a same load capacitance for a given phase margin. It copes better with heavy capacitive load ,45. HoweThe feedforward compensation technique is introduced to obtain high-frequency performance by implementing pole-zero cancellation ,47. Its mplifier . For the with Cf .The single-stage amplifier only have one high-impedance node at its output, that is why it can drive a large load capacitance without any stability issues. For the pseudo single-stage amplifier, an introduced intermediate resistance ant pole . This twIn summary, there are mainly two problems existing in these previous frequency compensation topologies. First, the size of compensation capacitor is tightly limited by the load capacitance. Specifically, for a 1-nF load capacitance, the compensation capacitance needed to address stability issues is conventionally larger than 10 pF, which occupies a large proportion of the chip area. In addition, these techniques are required to be optimized based on the single specific load capacitance. As a result, the wide range of load capacitance cannot be driven. In next section, a novel two-stage operational amplifier is introduced to solve these two problems by combing Miller compensation and pole-zero cancellation.The proposed operational amplifier architecture is illustrated in In the main amplifier, the first stage transconductance Note that Equation . HoweverThe stability of the operational amplifier is dominated by its phase margin, which is the difference between the phase and 180mplifier  and the While the two-stage amplifier implementing conventional Miller compensation has a second pole determined by the load capacitance, as shown in Equation (4), the dominant and second pole of this work. which are shown in Equations (19) and (21), are both independent of the load capacitance. As a result, this work can maintain a sufficient phase margin with various load capacitance, and this frequency compensation topology is less sensitive to the load capacitance compared to the conventional Miller compensation.Design parameters of the proposed amplifier are shown in  and M13 . The Rm hip area . The biaIn order to drive a wide range of load capacitance, The effect of Miller compensation capacitance variation on the precise pole-zero cancellation is illustrated in The common-mode rejection ratio (CMRR) of the differential amplifier reflects its ability of rejecting identical signal components on both inputs. The CMRR is defined asFor the proposed amplifier, its common-mode rejection ratio at low frequency is typically determined by the first stage of the gain booster. Assuming that the intrinsic gain of the transistor Then the CMRR of the proposed work at DC isThe proposed amplifier is designed using a 130-nm CMOS technology with a total area of 0.00090 The stability simulation with 0.05–17 nF load capacitance over Miller compensation capacitances variations are shown in The simulated frequency response with a 2.5-nF load capacitance is shown in The simulated common-mode rejection ratio (CMRR) is given in Under a transient simulation setup illustrated in The input common-mode range (ICMR) and output swing of the proposed amplifier are shown in Monte Carlo simulations with 50 samples for DC gain, unity gain bandwidth and phase margin with 1-nF load capacitance are shown in Transducers with nonlinear capacitive input impedance need an operational amplifier, which is stable for a wide range of load capacitance. Such an operational amplifier in a conventional design requires a large area for compensation capacitors, increasing costs and limiting applications. In order to address this problem, we present a gain-boosted two-stage operational amplifier, whose frequency response compensation is less sensitive to the load capacitance compared to the conventional Miller frequency compensation. The proposed amplifier cancels the output node pole of the main amplifier stage by the zero of the gain booster, so that a sufficient phase margin can be achieved without using a large compensation capacitor. A prototype CMOS amplifier designed with the proposed architecture uses two-to-four orders of magnitude smaller compensation capacitor compared to the load capacitance. This advantage in the compensation capacitor area requirement extends the applications of the proposed operational amplifier beyond the transducer interface circuits and may benefit general analog integrated circuit applications. The prototype CMOS operational amplifier demonstrates the highest performance trade-off to date when considering unity-gain bandwidth, load capacitance, power consumption, and chip area, which can enable an a compact low-cost transducer interface with unprecedentedly high resolution for emerging applications. It should be also noted that the proposed operational amplifier design technique can be used not only with monolithically integrated transducer interface circuits but also with transducer interface circuits assembled by discrete off-the-shelf components.


Globally mud areas on continental shelves are conduits for the dispersal of fluvial-sourced sediment. We address fundamental issues in sediment dynamics focusing on how mud is retained on the seabed on shallow inner shelves and what are the sources of mud. Through a process-based comprehensive study that integrates dynamics, provenance, and sedimentology, here we show that the key mechanism to keep mud on the seabed is the water-column stratification that forms a dynamic barrier in the vertical that restricts the upward mixing of suspended sediment. We studied the 1000 km-long mud belt that extends from the mouth of the Changjiang (Yangtze) River along the coast of Zhejiang and Fujian Provinces of China and ends on the west coast of Taiwan. This mud belt system is dynamically attached to the fluvial sources, of which the Changjiang River is the primary source. Winter is the constructive phase when active deposition takes place of fine-grained sediment carried mainly by the Changjiang plume driven by Zhe-Min Coastal Currents southwestward along the coast. They are active modern systems formed by complex interactions among single or multiple fluvial sources, physical processes, and the physiographic setting on multiple time and space scales. Most of these systems receive sediment from rivers and are considered as river-dominated sediment dispersal systems3. The shelf morphology and physical processes such as river plume, waves, and ocean currents that transport the fluvial sourced sediment determine the mud distribution pattern and the location of mud depocenters on continental shelves5. Since mud is very mobile, mud systems are easily subject to oceanographic processes, especially resuspension and transport by waves and currents. Consequently, on energetic shelves, constant mud deposition mostly occurs on the mid-shelf in deeper waters8. Mud-rich areas are also located in sheltered depressions1. In rare cases, when there is large supply of fine-grained sediment in low energy environments, mud deposits can be found on the inner shelf5.Mud deposition systems are common features on continental shelves in the world1. Furthermore, these systems are actually conduits linking the source (at river mouths), processes , and sinks (shelf depocenters on margins). Based on this view, 5 types of fine-grained river-sediment dispersal systems have also been proposed: proximal-accumulation-dominated (PAD), marine-dispersal-dominated (MDD), estuarine-accumulation-dominated (EAD), subaqueous-delta-clinoform (SDC), and canyon-captured (CC)2. Although some of the types in the two classification schemes overlap, they nevertheless point out the diversity in these systems.There are eight types of mud deposition systems in the world based on the location of the depocenter on the shelf and their three-dimensional architecture. They include prodelta, subaqueous delta, mud patch, mud blanket, mud belt, shallow-water contourite drift, mud entrapment, and mud wedge9, they preserve signals of river catchments and marine environments and thus the mud deposition systems also act as archives of paleoclimatic and paleo- and modern-environmental changes and coastal evolution during deglaciation15. Because of their archiving ability, they are also sinks for terrestrial substances and pollutants18, especially at depocenters.Since fine-grained sediments are carriers of geochemical signals and environmental proxiesThese mud deposition systems reflect not only how the earth system functions at the land-sea boundary as influenced by the global change, they also reveal the nature of anthropogenic impact on the continental margin. Therefore, it is important to have a holistic understanding of the process and response that lead to the formation and maintenance of these systems.23. Although the northern terminus is clearly defined, the southern end has not been well defined because most studies have been done on the Mainland China side. In this study, we combined the schematic maps of the mud areas based on data from Mainland China24 and Taiwan26 to make a schematic composite map to delineate the entirety of the mud belt system  that extends from the mouth of the Changjiang (Yangtze) River along the coast of Zhejiang and Fujian (Zhe-Min in brief in Chinese) Provinces into the Taiwan Strait (TS) and eventually terminates on the west coast of Taiwan Fig. . The lentem Fig. .Figure 129. Provenance studies of sediment along this mud belt based on clay minerals28, magnetic properties30, and radioactive nuclides16 indicate this mud belt is a conduit for both mainland and Taiwan-derived sediments, and thus, a source-to-sink system that links Chinese Mainland and Taiwan. Sub-bottom profile studies show that the thickness of clinoform and mud wedge decreases from the Changjiang submerged delta along the Zhe-Min Coast31.The mud region along the Zhe-Min Coast has been well studied to show that this is an archive and sink for Changjiang-derived sediment and organic material32. The energetic tidal system in the ECS also affects the dynamics. Many of these processes have been suggested to disperse Changjiang-derived fresh water and sediments and transport southward along the mud belt, such as the Changjiang Diluted Water (CDW) that forms a narrow band attaching to the Zhe-Min Coast33 by the ZMCC34. Additionally, the buoyant CDW and the more saline KBC water create haloclines35 on the ECS shelf, forming stratification that affects the vertical structure of suspended sediment, which in turn, affects sediment transport along the mud belt34. Therefore, this is quite a complicated sediment dispersal system.Dynamically this system is subject to the strong monsoonal seasonality of the river discharge and related plume dynamics, coastal currents in the ECS such as the Zhe-Min Coastal Current (ZMCC) and the intrusion of Kuroshio Branch Current (KBC), South China Sea Warm Current (SCSWC), and wind and wave fieldsDespite the complexity in the dynamics of the system, there have not been studies that directly link the dynamics of sediment to the dispersal of the fluvial source, sediment settling and transport, and eventual deposition along this conduit. Fundamental questions regarding the dynamic maintenance of this system have yet to be answered with certainty. Specifically, how is the mobile mud retained on this energetic shallow inner shelf? What prevents the mud to get dispersed to deeper waters as it does in other systems? How does the river-derived sediment enter the system? What is the evidence in the settling particles in the water column to link the fluvial sources to the seafloor sediment? In order to the address these questions, we designed a comprehensive process-based study. We used an instrumented mooring with a sediment trap to investigate the coupling between the temporal changes in the flow field to water temperature, and salinity and suspended sediment concentration over two spring-neap cycles. We used using shipboard observations to investigate the coupling between structures in the water column  and suspended sediment distribution, to establish cause-and-effective relationship over a diurnal tidal cycle. We compared the properties of settling particles captured in the sediment trap to hypothetical fluvial sources and establish direct provenance of settling particles. Additionally we examined the evidence from the seafloor surficial sediment and sediment cores along the mud belt to establish source-to-sink characteristics of the mud belt. Our goal is to integrate all the measurements of the water column and the corresponding suspended and seafloor sediments to establish a process-based model of this dispersal system that links the mouth of Changjiang River and the west coast of Taiwan.In our field campaign R/V Ocean Researcher 5 (OR-5) deployed a sediment trap mooring off the mouth of Oujiang River on Feb. 17, 2014; took seafloor sediment samples, and box and gravity cores along the entire mud belt Fig. . A chart33 water was evident as shown by the close simultaneous associations of low salinity and temperature  profile in the lower part of the water column Fig. . Higher 40 to the study site as seen in the warmer temperature at −20 mab than that at −17.3 mab measured on the mooring, resulting in sharp temperature reversals  showed pronounced changes over a diurnal tidal cycle on March 17–18 at Sta. M1 Fig. . During ure Fig. , which wues Fig. , whose s*) induced by waves, currents, and wave-current interaction over a diurnal tidal cycle were estimated using the method described before44 were compared against entrainment thresholds of non-cohesive sediment of the grain sizes of 10, 63, and 153 μm, respectively  was the most important mechanism to resuspend seabed sediments of all three non-cohesive size classes. Furthermore, U*cw barely reached the entrainment threshold for the cohesive sediment of 10 μm size during the flooding tide beginning at 06:00, March 18. Therefore, fine-grained cohesive sediment could be resuspended as well. The volume concentrations of grain-size classes of 63–153, 10–63, and <10 μm only appeared in the lower water column, having two cores of high values corresponding to low tides . The results show that a layer of high E values at mid-depth could be simulated along a shore-normal transect  above the mud belt. These particles were fine-grained, predominantly in clay and silt fractions , illite, kaolinite, chlorite, quartz, k-feldspar, plagioclase, and calcite  is constantly supplied in winter.Our findings suggest that the mud area shown in Fig. d46 Fig. . This cosit Fig. , althoug48 that could act as lateral barriers to prevent suspended mud to escape seaward, further confining the mud within the mud belt. In winter the expression of the vertical isopycnal interface at the sea surface can be seen in satellite-derived sea surface temperature gradients, which form a frontal zone that corresponds well with the seaward boundary of the mud belt along Zhe-Min Coast  as the key mechanism under winter conditions. This barrier restricts suspend sediment to the lower water column where the suspended concentration is high. Yet the weaken flow in the lower water column limits effective sediment transport. This barrier in the vertical is likely to exist in the confines of the Changjiang buoyant plume or CDW transported by ZMCC. Furthermore, the three-D density structures, due to the Changjiang buoyant plume and KBC, create shore-parallel vertical isopycnal surfaces21. A sediment dynamics study off the mouth of Minjiang River in summer shows under contrasting wind conditions of SW summer monsoon and NE typhoon winds, a stratified/isopycnal layer existed at mid-depth44. Under both conditions a benthic nepheloid layer below the dynamic barrier formed by the isopycnal layer was also present. Thus, the ‘entrapment mechanism’ to prevent upward mixing of suspended sediment near the sea floor still exists in summer. Since ZMCC is weakened or replaced by northward flowing SCSWC, Changjiang River would not be a major source for fluvial sediment and the predominant sediment transport direction is northward21. Therefore, the Z-M-TSMD is likely to have two phases in the course of a year. During the winter monsoon season, the system receives ample fluvial sediment from the Changjiang River via southward sediment transport, and the deposition on the sea floor is active and the vertical and lateral dynamic barriers are well in place. This is the constructive phase. When the summer monsoon dominates, the system receives sediment delivered from small rivers on the Zhe-Min Coast, but little from Taiwan49. The buoyant water and colder and more saline water brought to the coast by upwelling forms stratification that could create a similar barrier in the vertical44. The amount of suspended sediment in the lower water column is less than that in winter34. Episodic typhoon events are likely to interrupt sediment transport and cause non-deposition21. They also deliver a large amount of fluvial sediment to the system. The impact of the typhoon on the system is yet to be evaluated. Nevertheless, the system is likely in a maintenance or non-constructive phase. The two-phase scenario fits the description of oceanic fronts and hydrodynamic and suspended sediment transport in this region50. Overall, the Z-M-TSMD is directly attached to the fluvial sources and is proximal-accumulation-dominated with a decreasing degree from the major fluvial source. Furthermore, the system’s fluvial sediment is driven by coastal currents over a distance of 1000 km, and thus, is also marine-dispersal-dominated. Therefore, it is a major hybrid dispersal system of distal-to-proximal accumulated and marine-dispersal dominated. Comparing to the characteristics of mud deposition zones and river dispersal systems in the world2, the Z-M-TSMD’s hybrid nature makes it unique in the world.However, our data did not cover the summer season, we therefore speculate based on known facts. In summer the dispersal of CDW is in the NW direction, it does not overlap with Z-M-TSMS. Yet, summer is the flood season of all rivers in the region, the effluent from other rivers along the mud belt replaces the role of the Changjiang buoyant plume and the vertical stratification in the water column still exists°E, 27.8°N) , and a CTD with a turbidity sensor (XR-420). Afterwards, OR-5 took 3 gravity and 7 box cores and sediment samples from the seabed along the mud belt  Fig.  in water°N) Fig. . It was elt Fig.  from Feb44. Seabed sediment samples were analyzed for grain-size distribution using the methods described before44.On March 13 R/V Ocean Researcher 1 (OR-1) arrived at Sta. M1 to conduct 24-hr hydrographic profiling, water and seafloor sediment sampling before retrieving the sediment trap mooring. The profiling included hourly salinity, temperature, chlorophyll-a, and volume concentration of suspended particles of thirty-two grain sizes (using LISST-100X). Water samples were taken at the surface, mid-depth, and near seabed for the analysis of suspended sediment concentration (SSC) at 2-hr intervals using the method described before53. One liter of the residual water in the Catnet container was later filtered through the GF/F (0.7 μm) filter paper. The filtration separated suspended particles into four different grain-size classes . All filter papers were first stored in a refrigerator at −4°C, than dried in the oven at 55 °C in the laboratory for 24 hours and weighed to calculate SSC43.On board RV OR-1 water samples were taken at 3-m below the surface (surface), the location of the thermocline (mid-depth) at the time of sampling, and near bed (bottom-instantaneous depth minus 10%) using 10-l X-Niskin bottles mounted on the CTD rosette. Water samples were filtered onboard immediately using a nested filtration system called Catnet, which contained three funnel-shaped Nitex nets stacked together inside a polycarbonate container having the volume of 60 l. The mash sizes of the nets were 10, 63 and 153 μm, respectively. Particles captured on the Nitex nets were carefully washed into a 100 ml PE bottle by hand using deionized distilled water (DDW), which was filtered using GF/F (0.7 μm) filter paper. Great care was taken in the washing process to avoid breakage of floc particles56:v stands for volume backscattering strength (dB), α is the attenuation coefficient (dB/m), Kc is a scale factor (dB/count), E is echo strength (in count), Er is received noise (in counts), TT denotes the transducer temperature (°C), R is the distance along the central beam axis (m), L is transmit pulse length (m), PT is the transmit power (W), and C is a constant (dB). After combining other constants  with C,  measured from collected water samples at corresponding depths.The active sonar equation has been widely used to convert SSC from backscattering strength for decades3) and is the water density at a reference depth.E is expressed as:58.The analysis was conducted at the Institute of Oceanography, Chinese Academy of Sciences using X-ray Diffraction (XRD) with a D8 ADVANCE diffractometer with CuKα radiation (40 kV and 40 mA) following established procedures7Be and 210Pbex were carried out in the Institute of Earth Sciences, Academia Sinica following the established procedures26.The analyses of 59.Grain-size distribution of samples from the sediment trap and seabed were analyzed in the Department of Oceanography, National Sun Yat-sen University following established proceduresSupplementary Information