Tourism has been perceived as an environmentally friendly industry for a long term, but the negative impact of tourist irresponsible behavior on the environment cannot be ignored. Tourists’ pro-environmental behavior is crucial to the destination’s sustainable development. Taking stimulus-organism-response theory as a framework, this study explores the factors and mechanisms influencing tourists’ pro-environmental behaviors by integrating cognitive-affective-conative (CAC) theory and motivation-opportunity-ability (MOA) theory. Through the PLS-SEM analysis with 548 questionnaires, the driving mechanism of tourists’ pro-environmental behavior was deeply explored. The results indicate that tourists’ environmental knowledge, participation motivation, and opportunity have significant positive effects on pro-environmental behavior, but the role of participation ability on tourists’ pro-environmental behavior is not confirmed. Tourists have significant environmental self-identity and environmental commitment, and these affections actively promote tourists’ pro-environmental behavior. Except for the participation ability, the other three factors, respectively, influence tourists’ environmental behavior through the mediating effects of environmental self-identity and environmental commitment. Given the limited literature integrating CAC and MOA theories within research on tourists’ pro-environmental behavior, these findings provide new perspectives for future research. This research enriches the literature on the factors influencing tourist pro-environmental behavior and also provides practical guidance for promoting tourists’ pro-environmental behavior. Although tourism has been considered as a green industry, the burden of tourism development on the destinations’ ecological environment cannot be ignored . In addiPro-environmental behaviors are those behaviors that tourists perform during tourism activities to reduce the negative impact on the destination environment and promote the sustainability of resource use . CurrentWith further research, scholars have argued that cognitive and affective factors play an important driving role in tourists’ pro-environmental behavior . Among tThe tourists’ inappropriate behaviors not only bring negative effects to tourist attractions but also hinder the sustainable development of tourist destinations. Currently, it is an important issue to guide and enhance tourists’ pro-environmental behavior in academia. Assessing the factors influencing tourists’ pro-environmental behavior is significant to improve tourism sustainability . TherefoThe Stimulus-Organism-Response (SOR) theory proposed by environmental psychologists The SOR paradigm has well explained the emotional changes and behavioral characteristics of individuals and has been widely validated in many research domains. The framework has been extended to many other research fields, like online behavior , consumeThe theoretical framework in the present study mainly derived from the trilogy of mind, namely Cognitive-Affective-Conative (CAC) theory . The corThe CAC theory plays a significant role in explaining an individual’s behavioral process. Therefore, the current study will take it as a theoretical basis to further explore the formative mechanisms of tourists’ pro-environmental behavior. Specifically, tourists’ environmental-related knowledge will influence their initial judgments when carrying out activities in tourist destinations, from which they will generate certain emotions and feelings. This research will focus on exploring the environmental self-identity and environmental commitment of tourists. The environmental self-identity and environmental commitment generated by tourists will drive their actual pro-environmental behavior. In other words, according to the CAC theory, tourist environmental knowledge (cognitive stage) will stimulate the environmental self-identity and environmental commitment (affective stage), which in turn influence the pro-environmental behavior (conative stage).Environmental knowledge is defined as general knowledge of facts, concepts, and solutions to environmental issues, and reflects the degree of understanding and concern for the relationship between humans and nature . SpecifiThe person-environment relationship structure provides a fundament for understanding tourists’ pro-environmental behavior . The preSelf-identity is a label that an individual uses to describe oneself . In otheCommitment is a widely researched topic in business operations and organizational management . It playRealizing that tourism activities have many negative impacts on the ecological environment, a growing number of studies have focused on tourists’ behavior. As important stakeholders, tourists’ pro-environmental behavior can have a significant impact on destinations’ environmentally sustainable development. Pro-environmental behavior is the action of an individual to consciously reduce the negative impact on the environment , such asIn tourism research, tourists’ pro-environmental behavior refers to the behavior of individuals or groups that promote the sustainable use of natural resources . AttribuThe Motivation-Opportunity-Ability (MOA) theory was proposed by In recent studies, scholars have extended the MOA theory to examine environmental behaviors. For example, Based on the above discussion of relevant literature, the theoretical framework of the present study extended the SOR theory by incorporating the MOA theory and the CAC theory. According to the framework, three key hypotheses have been made: (1) tourists are stimulated by multiple factors to produce psychological changes in the tourism environment, which in turn cause them to perform pro-environmental behavior; (2) tourists’ environmental knowledge (cognition) will inspire environment-related self-identity and commitment (affection), which in turn drives tourists’ pro-environmental behavior (conation); (3) participation motivation and participation ability factors are intrinsic factors that drive individual behavior, and participation opportunity is an external environmental factor that influences individual behavioral intention. To sum up, participation motivation, participation opportunity, participation ability, and environmental knowledge are considered as Stimulus (S) in the research framework, environmental self-identity and environmental commitment represent organism (O), and tourists’ pro-environmental behavior is considered as the response (R).Some studies have confirmed the impact of certain knowledge on identity and commitment, such as the positive impact of food safety knowledge on food safety commitment . HoweverH1: Tourists' environmental knowledge positively influences environmental self-identity.H2: Tourists' environmental knowledge positively influences environmental commitment.Self-identity is thought to be highly related to environmental behavior. It has been suggested that environmental self-identity predicts behavioral consistency. For example, green self-identity is related to organic food consumption behavior , sustainIndividuals who make environmental commitments are more likely to practice their sense of environmental responsibility through actual actions. With high levels of environmental commitment, people are more likely to increase their pro-environmental behavior. Considering the role of environmental commitment in shaping behavior, it has also received considerable attention in tourism research . For exaH3: Environmental self-identity positively influences tourists' pro-environmental behavior.H4: Environmental commitment positively influences tourists' pro-environmental behavior.Environmental knowledge not only represents an individual’s level of awareness on relevant environmental issues, but also characterizes the degree that he or she cares about ecology. Environmental knowledge can increase people’s awareness of environmental issues, which in turn increases pro-environmental behavior. It has been shown in several empirical studies that environmental knowledge plays an important role in predicting individual behavioral intentions . In the H5: Environmental knowledge positively influences tourists' pro-environmental behavior.In the original MOA theory, H6: Participation motivation positively influences environmental self-identity.H7: Participation motivation positively influences environmental commitment.Motivation is an important antecedent variable in explaining individual behavioral characteristics. The role of motivation in predicting individual attitudes and behavioral intentions has been widely confirmed. Regarding the influence of motivation on pro-environmental behavior, H8: Participation motivation positively influences tourists' pro-environmental behavior.According to H9: Participation opportunity has a positive impact on tourists' environmental self-identity.H10: Participation opportunity has a positive influence on tourists' environmental commitment.H11: Participation opportunity has a positive impact on tourists' pro-environmental behavior.According to the original concept of the theory, ability refers to consumers’ ability to understand advertising messages , which gH12: Participation ability positively influences tourists' environmental self-identity.H13: Participation ability positively influences tourists' environmental commitment.H14: Participation ability positively influences tourists' pro-environmental behavior.In addition, the research also considered the possible mediating role of environmental self-identity and environmental commitment separately. We proposed the following hypotheses:H15: Environmental self-identity (a)/environmental commitment (b) mediates the effect of environmental knowledge on tourists' pro-environmental behavior.H16: Environmental self-identity (a)/environmental commitment (b) mediates the effect of participation motivation on tourists' pro-environmental behavior.H17: Environmental self-identity (a)/environmental commitment (b) mediates the effect of participation opportunity on tourists' pro-environmental behavior.H18: Environmental self-identity (a)/environmental commitment (b) mediates the effect of participation ability on tourists' pro-environmental behavior.Based on the above discussion of the relationship between variables, this research exploratively proposed a research model for the influence mechanism of tourists’ pro-environmental behavior .The measurement of variables involved in this research is derived from established scales in domestic and international studies with appropriate modifications, which ensures the validity and reliability of the research data. To avoid the semantic differences or cultural background differences affecting the quality of the questionnaire, the questionnaire was first translated independently by the research team members and formed into a primary Chinese version scale. Then, it is reviewed by three tourism experts and two PhDs in the relevant fields. Until the language was accurate and unambiguous, the research scale was determined. The four construct items of environmental knowledge are referenced from Due to the impact of COVID-19, this research collected questionnaire data from an online platform WenjuanxingThis study analyzed the influence mechanism of tourists’ pro-environmental behavior using partial least squares structural equation modeling (PLS-SEM). The empirical analysis was based on a two-step process. First, the measurement model was assessed to test the consistency reliability, convergent validity, and discriminant validity. Next, we assessed the structural model and tested the hypotheses.Generally, consistency reliability, convergent validity, and discriminant validity are used to assess the measurement model. As shown in Next, we assessed the discriminant validity with the use of the Fornell-Larcker criterion . Table 3We tested the structural model and the research hypotheses with the help of the PLS Algorithm, Blindfolding, and Bootstrapping method . First, we evaluated the model fit by the standardized root mean square residual (SRMR) value. In this research, we obtained an SRMR value of 0.071 (less than 0.08), which indicated that the model specification was within the eligible threshold .β), determination coefficient (R2), and the cross-validated redundancy measure (Q2) provide an important reference for assessing the structural model . The model explained 64.4% of environmental self-identity, 60.2% of environmental commitment variance and 68.8% of tourists’ pro-environmental behavior.The values of the path coefficient , environmental commitment , and tourists’ pro-environmental behavior , which supported H1, H2, H4 and H5. The effects of environmental self-identity and environmental commitment on tourists’ pro-environmental behavior  were positively significant. Hence, hypotheses H3 and H4 were supported. As well, participation motivation had a significant effect on environmental self-identity , environmental commitment , and tourists’ pro-environmental behavior , and hypotheses H6, H7 and H8 were supported. In addition, the effect of participation opportunity on environmental self-identity , environmental commitment  and tourists’ pro-environmental behavior  were positively significant, which supported hypotheses H9, H10 and H11. Lastly, participation ability significantly and positively affected environmental self-identity  and environmental commitment , and supported hypotheses H12 and H13. The effect of participation ability on tourists’ pro-environmental behavior  was not significant, hence hypothesis H14 was rejected.Specifically, β = 0.093, p < 0.001; β = 0.075, p < 0.001). Environmental self-identity and environmental commitment mediated the effect of participation motivation on tourists’ pro-environmental behavior . Meanwhile, the mediation effects of environmental self-identity and environmental commitment between participation opportunity and tourists’ pro-environmental behavior were confirmed . None of the bias-corrected lower limit (2.5%) and upper limit (97.5%) confidence intervals included zero. Hence, the mediation hypotheses for H15a, H16a, H17a, H15b, H16b, and H17b are well supported. While the mediating effects of environmental self-identity and environmental commitment between participation ability and tourists’ pro-environmental behavior were not confirmed in this research. Thus, H18a and H18b are not empirically supported.Finally, we assessed the mediating role of stimulus factors  between organism factors  and tourists’ pro-environmental behavior. As shown in Based on an empirical test, this study has broadened our understanding of tourists’ pro-environmental behavior by integrating the CAC theory and the MOA theory grounded in the SOR theory. Overall, this research assessed the role of stimulus and organismic factors in an attempt to reveal the driving mechanisms of pro-environmental behavior in tourists. Our research supports the effects of environmental self-identity and environmental commitment in the relationship between stimulus factors  and tourists’ pro-environmental behavior. Specifically, the following main findings are found in the research.H14. What is interesting to note is that in examining tourists’ social media engagement behavior, First, by investigating the direct effect of environmental knowledge on tourists’ pro-environmental behavior, the findings confirm that environmental knowledge positively affects tourists’ pro-environmental behavior, which supports previous research findings . This reSecondly, the findings that environmental self-identity and environmental commitment positively influence tourists’ pro-environmental behavior are consistent with previous research findings . In the Finally, we also find support for the research hypotheses that the relationship between stimulus factors  and tourists’ pro-environmental behavior are mediated by environmental self-identity and environmental commitment. These results are similar to the findings of This study makes a theoretical contribution to advancing the research on tourists’ pro-environmental behavior. Based on empirical tests, this research provides new ideas for exploring tourists’ pro-environmental behavior. On one hand, in the SOR theoretical framework, this research introduces the CAC theory and MOA theory to explore the antecedents of tourists’ pro-environmental behavior. Our findings show that environmental knowledge, participation motivation, and opportunity have a positive impact on tourists’ pro-environmental behavior. This research is the first time to integrate the CAC and MOA theories to consider individual pro-environmental behavior in tourism context, which extends the application of related theories in the tourism domain.On the other hand, to deeply analyze the driving mechanisms of tourists’ pro-environmental behavior, the study incorporates the person-environment relationship into the model framework. Previous studies have mainly explored the possible driving factors of tourist pro-environmental behavior from the perspective of the person-place relationship , while tFrom a practical perspective, the findings from this work provide some guidance on enhancing tourists’ pro-environmental behavior. The results show that the level of environmental knowledge significantly affects the pro-environmental behavior of tourists. Therefore, the government should strengthen the popularization of environmental knowledge and continuously enhance social awareness of environmental protection. It is necessary to actively seek ways to translate environmental knowledge into real pro-environmental action. It should strengthen the breadth and depth of environmental knowledge popularization through social media platform publicity and organizing expert knowledge lectures. Tourist destination managers can organize plentiful environmental education activities in scenic spots to disseminate scientific knowledge and general knowledge about the environment and protection skills to tourists. As tourists’ environmental knowledge grows, their self-identity and commitment to environmental protection will strengthen, which eventually transform into actual pro-environmental behavior. In the meantime, tourism participation motivation also positively stimulates tourists’ pro-environmental behavior. Destination managers can encourage tourist pro-environmental behavior by setting external incentives. It can also enhance tourism participation motivation in the form of internal incentives by promoting tourists’ environmental concern and environmental responsibility, which will promote tourists’ pro-environmental behavior.In addition, given the facilitating role of participation opportunity, destinations should provide a convenient environment and material support for tourists to carry out pro-environmental behavior, by continuously improving ecological infrastructure construction and enhancing the supply of pro-environmental tourism products. Government and destination managers should also actively develop management policies to provide specific guidance for regulating tourists’ behavior and enhancing pro-environmental behavior. Finally, in view of the important role of environmental self-identity and environmental commitment, destination managers should actively explore ways to enhance tourists’ environmental self-identity and environmental commitment. Aiming to enhance the awareness and connection of the person-environment relationship, tourism destinations can use multiple social media platforms to widely promote their good ecological environment and continuously create a social atmosphere for eco-environmental protection. These favorable external environments will trigger tourists’ appreciation and approval of the destination, inspire them to develop an environmental identity, make positive environmental commitments, and in turn exhibit specific pro-environmental behavior.The limitations of this work provide insights for further research. First, this study investigated tourists’ subjective environmental knowledge through self-reporting, and the tourists’ environmental knowledge level may be overestimated. Therefore, future researchers need to adopt a more comprehensive approach to examine the tourists’ objective environmental knowledge to more accurately assess the level of environmental knowledge of tourists. Second, many researchers have classified tourism motivation in terms of push-pull motivation, altruistic motivation, and egoistic motivation. Although this research measured tourists’ motivation in terms of both egoistic and altruistic motivation when collecting data, the empirical analysis did not explore whether the two motivations differ in their effects on tourists’ pro-environmental behavior. Hence, we suggest that future research could further explore the mechanism differences in the effects of egoistic and altruistic motivations on tourists’ pro-environmental behavior. Third, future research can further analyze other antecedents, such as social norms, values, risk perceptions, and destination image, in order to fully reveal the driving mechanisms of tourists’ pro-environmental behavior. In some research, scholars have considered the boundary condition that affects tourists’ pro-environmental behavior, such as destination reputation and environmental level . In futuThe raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.Ethical review and approval was not required for the study on human participants in accordance with the local legislation and institutional requirements. The patients/participants provided their written informed consent to participate in this study.HT: data collection, literature review, data compiling, data analysis, and text formulation. YM: development of research concept, research framework development and guidance, expert editorial guidance, and funding acquisition. JR: research framework development and guidance, expert editorial guidance and text edition and modification. All authors contributed to the article and approved the submitted version.This research was funded by the Project of National Social Science Fund of China (19BJL036).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


MicroRNAs have been projected as promising tools for diagnostic and prognostic purposes in cancer. More recently, they have been highlighted as RNA therapeutic targets for cancer therapy. Though miRs perform a generic function of post-transcriptional gene regulation, their utility in RNA therapeutics mostly relies on their biochemical nature and their assembly with other macromolecules. Release of extracellular miRs is broadly categorized into two different compositions, namely exosomal (extracellular vesicles) and non-exosomal. This nature of miRs not only affects the uptake into target cells but also poses a challenge and opportunity for RNA therapeutics in cancer. By virtue of their ability to act as mediators of intercellular communication in the tumor microenvironment, extracellular miRs perform both, depending upon the target cell and target landscape, pro- and anti-tumor functions. Tumor-derived miRs mostly perform pro-tumor functions, whereas host cell- or stroma-derived miRs are involved in anti-tumor activities. This review deals with the recent understanding of exosomal and non-exosomal miRs in the tumor microenvironment, as a tool for pro- and anti-tumor activity and prospective exploit options for cancer therapy. Every crisis comes with an opportunity, which is also true for COVID-19. Treatment options for SARS-CoV-2 infections led to the early adoption of RNA therapeutics . mRNA-baThe genesis of unicellular and multicellular organisms shares a common trait that is crucial for their success of life, which is the propagation of signals to the environment, including cellular environments. The uncanny perfection with which cells communicate with their environment using chemicals, including nucleic acids, is responsible for the evolution of complex life forms, and dysregulation therein plays an important role in their diseases. An established tumor acts as a multicellular parasitic organism within its host and relies on this communication. The tumor microenvironment consists of a plethora of soluble factors, either derived from the tumor cell or stroma, which take part in intercellular communication. These factors play a pivotal role in tumor growth and dissemination. Amongst these factors, small nucleic acids such as microRNAs (miR) play a prominent role in the quest of regulatory factors for cancer therapy since virtually all biological pathways are under the control of miRs as most mRNAs contain at least one functional miR target site ,6. FurthClassically, exosomal and non-exosomal miRs were differentiated using RNase to digest non-protected miRs. However, this differentiation has a serious limitation as it cannot discriminate AGO-protected miRs from exosomal miRs . FurtherClassification of extracellular vesicles (EVs), such as the exosome, is a complex process and is based on size and composition. Studies have identified at least two distinct subpopulations of exosomes, e.g., Exo-L  and Exo-S , which show distinctive biophysical and molecular properties as well as non-membranous “exomeres” (~35 nm) . These eRelease and uptake of EVs are one of the most important modes of intercellular communication, including in the tumor microenvironment . HealthyExosomal miR can be exchanged between the tumor and stroma and repress gene expression ,33. Howe+CD206+ macrophages then decrease CD8+ T cell activity and increase tumor growth ..+CD206+ Inflammatory conditions favor tumor growth and migration. Tumor-derived factors also alter the tumor microenvironment in a way that low-grade inflammation persists. Cancer cells also use miRs to achieve these objectives. Casadei et al. reported that liposarcoma cell-derived exosomes are rich in miR-24 and miR-92a. These miRs stimulate the production and release of proinflammatory IL-6 from TAMs via toll-like receptor (TLR)7/8 activation and NF-κB signaling . The relras and K-ras mRNAs, and Rab proteins . When these exosomes are exposed to adipose stem cells, it enhances prostate tumor formation in vivo by inducing genetic instability, mesenchymal-to-epithelial transition, and oncogenic transformation . In. In63]. These studies underscore the importance of exosomes in the tumor microenvironment. Though exosomes from tumor cells and stroma are involved in pro- and anti-tumoral effects, their retrovirus-like nature ,75 is anThere are many iterations of non-exosomal miRs, but they all share the common principle that extracellular miRs should be stable and protected from nuclease digestion. The predominantly studied non-exosomal miRs are AGO-bound or RNA-induced silencing complex (RISC)-containing AGO ,77,78. HContrary to the tumor microenvironment, almost all miRs in plasma of healthy donors could be immunoprecipitated by AGO2 antibodies, suggesting that the majority of plasma-borne miRs are associated with non-exosomal fraction and are free to interact with anti-AGO2 antibodies . HoweverNot all non-exosomal miRs in the tumor microenvironment are necessarily protected or show the comparable miRome profile to exosomal miRs or miRome. It has been shown that AGO-bound non-exosomal miRs are subjected to degradation based on the exposed 3′ end of miRs, which may be due to the mutation in the AGO protein . These mTumor-derived non-exosomal miRs are scarcely reported in the literature, partly due to difficulty in isolating them or due to the major focus on EV-derived miRs. However, in biological fluids, there is a variety of miRs that are not encapsulated in EVs. One of the most remarkable examples is miR-375. miR-375 has been shown to be highly expressed in breast cancer cells and released upon undergoing apoptosis. This tumor-derived miR was shown to be associated with LDL (low-density lipoprotein). In the tumor microenvironment, LDL was taken up by macrophages via CD36 . The nonThe extracellular miR landscape is quite heterogeneous and the binary nature of exosomal and non-exosomal miR is context-dependent . SeveralWhen the first oncomiRomes were analyzed from healthy tissue vs. cancer samples, it was surprising that miRomes were better at predicting the cancer type and stage than transcriptome profiles ,90. RNA Cancer cell-specific targeting of RNA therapeutic molecules would severely reduce side effects and enhance the therapeutic efficacy. There are approaches which have been exploited for targeted delivery of miR mimetics. It has been shown that hyaluronic acid (HA)-chitosan nanoparticles delivered tumor suppressor miR-34a and doxorubicin (DOX) to triple-negative breast cancer cells in vitro and in vivo by targeting the HA receptor, which is overexpressed in breast cancer a 99]. T. T99]. TAnother way to exploit tumor-derived exosomes for targeted miR memetic delivery was recently reported in the literature. The authors developed anti-exosomes antibody-oligonucleotide complexes (ExomiR-Tracker) that exploit tumor exosomes. ExomiR-Tracker binds to the surface of an exosome and hijacks it to carry the therapeutic miR payload and gain entry into target cells. 9-mer of D-arginine enhances the endosomal escape of the anti-miR oligonucleotides, thereby executing the therapeutic miR memetic action . It is nTargeting a single miR for therapeutic purposes has its limitations as a single miR might be insufficient for clinical purposes due to a variety of related oncomiRs and their multiple targets. This degeneracy of miR action along with their stability poses the biggest challenge in RNA therapeutics of cancer ,113. SimStudies using exosomes as a source of miRs should be interpreted in the context of their origin, localization, target cells, and pathologies that are involved. Exosomes not only contain regulatory nucleic acids such as miRs but are also enriched with immunomodulatory proteins such as CSF-1, CCL2, FTL, TGFβ, etc. . These iThe notion that miRs are reliable diagnostic and prognostic markers is based on the premise that a substantial number of miRs can be detected in the plasma of cancer patients. However, the concentration of miRs in plasma is in the range of about 5 pM , which iSome anti-cancer compounds alter the expression of the oncomiRome  and miRsIt has been shown that the administration of mouse- or human-derived exosomes, in low doses, does not evoke strong immune responses ,145. HenTargeting small RNAs such as miRs should also take into consideration that the expression profile of lncRNA is not affected. It has been shown that lncRNA regulates miR expression . Further


Parkinson’s disease(PD) is a progressive, neurodegenerative nervous system disease.Psychiatric symptoms are common in PD,it is important to consider the suicide risk of these patients.The aim of this study is to investigate the risk of suicide in PD with a case control study and to determine the factors that may be associated.126 Parkinson’s patients and 117 age,gender matched healthy controls were included.Montreal Cognitive Assessment Scale(MoCA),Suicide Probability Scale(SPS),Beck Hopelessness Scale(BHS),Apathy Rating Scale(AES) and Multidimensional Scale of Perceived Social Support(MSPSS) were completed by participants.The PD group completed Parkinson’s Disease Quality of Life Questionnaire(PDQ-39) and data on disease duration,Movement Disorders Society-Unified Parkinson’s Disease Rating Scale(MDS-UPDRS),levodopa-equivalent daily doses(LEDD) were obtained.Hamilton Anxiety Rating Scale(HAM-A) and Hamilton Depression Rating Scale(HAM-D) were administered to all participants.SPS total scores were significantly lower and total scores of HAM-A,HAM-D and MSPSS were found to be significantly higher in the PD group.There was no significant difference between the two groups in terms of BHS total scores.In PD group a linear relationship was found between SPS total scores and BHS,PDQ39, HAM-A and HAM-D and an inverse relationship with AES.In the regression analysis,it was concluded that a one-unit increase in BHS total scores increased the probability of suicide by 17.1%.It is seen that SPS scores in patients are lower than controls and observed that SPS is correlated with hopelessness,depression,anxiety and quality of life.Although the possibility of suicide is found to be low in PD,this risk increases in patients with untreated depression and anxiety.Therefore,psychiatric evaluation may be recommended in these patients.No significant relationships.


CA-125 is a clinical biomarker with predictive effect on the prognosis of different cancers. Numerous clinical trials have been conducted to investigate the possibility of using the pretreatment level of CA-125 to predict the prognosis of epithelial ovarian cancer (EOC). However, its value in predicting prognosis remains controversial. The purpose of this meta-analysis was to assess the predictive value of pretreatment CA-125 levels for prognosis in EOC patients.We searched the EMBASE, Cochrane library, PubMed and Web of Science databases for studies published up to 3 December 2021, according to specific inclusion and exclusion criteria. The clinical studies that were included investigated the relationship between pretreatment CA-125 levels and ovarian cancer prognosis. Combined hazard ratios (HR) of overall survival (OS) and progression-free survival (PFS) reported in the studies were compared and analyzed using fixed-effects/random-effects models. Sensitivity analysis was used to assess study stability, while Egger’s and Begg’s tests were used to assess publication bias.This meta-analysis included 23 studies published in 2004 - 2021 with a total of 10,594 EOC patients. Comprehensive analysis demonstrated that the serum level of CA-125 before treatment was significantly correlated with overall survival  and progression-free survival . After comparing data from different FIGO stages and treatments, we discovered that a high pre-treatment serum CA-125 level was associated with a low survival rate.According to the results of this study, a higher pre-treatment serum CA-125 level is associated with poor survival outcomes, which can be utilized to predict the prognosis of EOC patients. Pre-treatment serum CA-125 level might provide reliable basis for predicting the risk of EOC disease progression. This study is registered with the International Prospective Register of Systematic Reviews (CRD42022300545).https://www.crd.york.ac.uk/prospero/display_record.php?RecordID=300545, identifier [CRD42022300545]. Ovarian cancer (OC) is one of the most common malignant tumors in the world, with the highest fatality rate among gynecological malignant tumors with a high degree of malignancy. The 5-year survival rate is less than 45% , 2. TherCA-125 is a transmembrane macromolecule glycoprotein secreted by coelomic epithelial cells. It has become a consensus that CA-125 plays an important role in the prognosis of OC –12, and This meta-analysis was conducted in accordance with the preferred reporting items for systematic reviews and meta-analyses (PRISMA) . We thorInclusion and exclusion criteria: The inclusion criteria were as follows: (1) EOC was diagnosed according to pathological examination or current clinical guidelines; (2) serum CA-125 level was collected before chemotherapy, surgery and neoadjuvant chemotherapy with an explicit cut-off value; (3) articles investigating the relationship between CA-125 and EOC prognosis [including hazard ratio (HR) and corresponding 95% confidence interval (CI) for overall survival (OS) and/or disease-free progression (PFS)], or providing sufficient data to calculate these indicators, including the clinical pathological parameters; (4) published as full-text papers; and (5) the articles were written in English. The exclusion criteria were as follows: (1) reviews, conference abstracts, letters, editorials, expert opinions, case reports, abstracts, and reviews; (2) basic research or animal studies; (3) duplicate publications; and (4) studies on the tumors originating in the gastrointestinal tract and having metastasis to the ovary.Two investigators independently collected corresponding data from eligible studies, and any disagreements were resolved through discussions with a third investigator. The following information was extracted: (1) study characteristics including the first author, country of origin, year of publication, number of patients, duration of follow-up, and method of survival analysis; (2) patient characteristics including geographical area, age, metastasis, FIGO stage, treatment and cut-off value; and (3) survival measures including HRs of OS, PFS, DFS and their 95% CIs. The HRs were extracted from multivariate or univariate analyses. If HR values were not provided in the original text, Kaplan-Meier survival curves were used to determine HR values and corresponding CIs (Engauge Digitizer version 10.8). In this meta-analysis, the primary and secondary outcomes were OS and PFS. E-mails were also sent to the corresponding author for the requested data.Newcastle-Ottawa Scale (NOS)  was used2 statistic. Data were combined using a random-effects model (DerSimonia-Laird method) if I2>50% or p<0.10 (indicating significant heterogeneity among studies); otherwise a fixed-effects model (Mantel-Haenszel method) was used. To identify sources of heterogeneity between studies, subgroup analyses were conducted based on geographic region, treatment method, and FIGO stage. Furthermore, sensitivity analysis was conducted to assess the impact of individual study data on HRs for OS and PFS. All statistical calculations were performed using STATA MP16 . In all the analyses, p<0.05 (two-sided) indicated a statistically significant difference.Pooled HRs and 95% CIs were calculated to assess the relationship between pretreatment CA-125 level and prognosis. The heterogeneity of the included studies was assessed using Cochran’s q-test and Higgins IA total of 3083 articles were retrieved from the system’s database, with 445 of them being eliminated owing to dataset duplication. Another 2422 articles were eliminated after analyzing their titles and abstracts. Afterward, 216 articles were reviewed using full-text reading, with 192 of them being eliminated for the following reasons: Six studies lacked required data; five studies were not accessible in full text, and 181 studies did not provide the desired result. There are a total of 24 eligible articles, one which was published in 1995 and was excluded because it was too old . Finally2 = 86.40%, p <0.001). The results presented that increased serum CA-125 level before treatment was associated with poorer OS . Therefore, a fixed-effects model was used. No subgroup analysis was done due to the small number of studies reporting PFS.Data from 13 studies with a total of 7145 patients were retrieved and utilized in the PFS analysis , 36–38. Sensitivity analysis was carried out after merging the included literatures. After removing the included literatures one by one, the effect size in the remaining literatures were combined again. The results showed that there was no significant change in the combined HR and 95% CI, indicating that the results of this meta-analysis are relatively stable and reliable . The pre-treatment serum CA-125 level had predictive significance for the prognosis regardless of whether the tumor was at an early or advanced stage or if NACT was employed. Elevated CA-125 may be a strong prognostic marker for disease progression, tumor recurrence, and long-term survival outcomes including OS and PFS.In recent years, the tumor microenvironment has received increasing attention in the field of tumor immunology. The mechanism of CA-125 affecting prognosis may be related to the promotion of metastasis of cancer cells to other sites. CA-125 is a repeated peptide epitope of the mucin MUC16 , which bSeveral previous meta-analyses and retrospective analyses have investigated at the prognostic role of CA-125 in solid tumors. Overexpression of MUC16 is associated with poor prognosis in various malignancies –51. CA-1Due to the biological heterogeneity of EOC, which is not a single disease entity but a group of heterogeneous tumors with different clinicopathological features, different pre-treatment CA-125 levels have been found in patients with different EOC subtypes, such as low levels in mucinous and clear cell carcinomas. Although the included studies did not use a consistent cut-off value for pre-treatment serum CA-125 concentration, most studies found that higher levels of CA-125 in the normal range were associated with a higher risk ratio for recurrence or death .The society of gynecologic oncology and the American society of clinical oncology consider that women with high perioperative risk or a low probability of tumor cytoreduction to a residual lesion <1cm  should receive NACH. Women who are eligible for PDS and have potentially resectable disease may receive either NACH or PDS. However, PDS is preferred if the likelihood of achieving cytoreduction to <1cm  at an acceptable incidence is high . Many stThe study by Merlo et al. indicated that higher pre-treatment CA-125 levels were less likely to achieve optimal cytoreduction. At the cut-off value of 500IU/ml, 58% of patients with advanced EOC achieved complete or at least optimal cytoreductive surgery, and at the cut-off value of 250IU/ml, the probability increased to 74%. Merlo concluded that a cut-off value of 500IU/ml was an ideal threshold for predicting PDS success , which wBased on our findings and those of other relevant studies, pre-treatment CA-125 may contribute to the prognosis of cancer and the development of clinical treatment strategies. For example, EOC patients with low pre-treatment CA-125 may benefit more from PDS, NACT, or other cancer-related treatments than EOC patients with high CA-125. The prognostic relevance of pre-treatment CA-125 has attracted clinical attention, and in order to benefit EOC patients, ongoing and systematic studies are still necessary to fully understand and utilize CA-125.There are several limitations to this study. First, the number of studies we included was relatively small. Second, we were unable to perform further subgroup analyses due to the diversity of cut-off values defined in each study. Third, most of the enrolled studies were retrospective and extracted several HRs from univariate analyses, which may lead to overestimated effect sizes. Therefore, this study may have information bias, selection bias, and misclassification bias, and the optimal cut-off value cannot be determined. Larger-scale prospective studies are needed to confirm the significance of pretreatment CA-125 in predicting the prognosis of EOC and to provide evidence for its clinical application. In summary, according to the current study, the pre-treatment CA-125 level has some certain significance in predicting the prognosis of EOC. The results of this study may help to determine whether patients may benefit from surgery, chemotherapy, and/or neoadjuvant chemotherapy.This meta-analysis has shown that higher pre-treatment CA-125 levels are significantly associated with poorer OS and PFS in patients with EOC. We suggest that CA-125 should be monitored to guide prognosis and provide reliable information on the risk of disease progression in EOC. Nevertheless, due to some limitations of this study, large multi-center prospective trials are needed to examine the prognostic role of pretreatment CA-125 in EOC.The original contributions presented in the study are included in the article/QW designed this study. QW and SZ contributed to literature retrieval, review and data extraction. SZ and XL conducted the statistical analyses. QW, and XL contributed to the manuscript drafting. QW, and XL contributed to the manuscript revision. XF provided fund help. All authors contributed to the article and approved the submitted version.Project supported by the National Natural Science Foundation of China (Grant NO.81973894).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


SLOfit Lifelong is a public health initiative which was created to upgrade a well-established, national physical fitness surveillance system for Slovenian schoolchildren that has been collecting annual fitness and health data for over three decades. The ultimate objective of creating SLOfit Lifelong was to build a modern societal infrastructure with the capacity and ability to detect future causal associations between childhood physical fitness trends and future health outcomes based on the lifelong surveillance of one's own fitness status. By instilling citizens with an ambition to test, understand, and follow-up their own physical fitness and health status , this initiative provides the technical support and expert feedback needed to engender greater individual control over understanding (and thus modulating), one's own physical fitness status as they progress into older adulthood. This perspective paper details the extensive approach taken to devise appropriate fitness test batteries for adults and older adults which can also relate to the student version of the original SLOfit test database, including establishing criterion health risk zones and a public approach to establish this national, citizen-driven health feedback framework. Through its sophisticated online web applications, social media, print media, and outreach workshops, SLOfit Lifelong provides the expert support for public health engagement by fostering positive lifelong physical literacy experiences an individual can enjoy across their aging journey. Cardiovascular diseases (CVD) have become the leading cause of global mortality in the 21st century primarily due to lifestyle changes, including increased physical inactivity . Althougvia lifestyle interventions.With aging, negative physiological and physical changes can occur, including sarcopenia , progresThe purpose of this paper is to communicate how SLOfit Lifelong was developed, the health data it conveys, and how this initiative collaborates with individuals to provide quality testing and expert feedback for adults of all ages who wish to better understand their body and its functional capacity across their lifetime.Any data collected within the context of SLOfit Lifelong was approved by the Slovenian National Medical Ethics Committee (ID: 0120-468/2021/3), following the Declaration of Helsinki. Participants provided written, informed consent prior to data collection.N = 13,000 graduates per year who are potential SLOfit Lifelong cohort members. SLOfit Lifelong is designed to be capable of housing their fitness data from childhood to adulthood, thus “slotting” in existing SLOfit members as they age, in parallel to recruiting new participants.The original SLOfit infrastructure program (herein differentiated as: SLOfit Students) monitors child fitness through their entire schooling period . This inwww.slofit.org/ucitelj/administracija). Testing is administered by PE teachers trained from the Faculty of Sport, University of Ljubljana, which is the only institution in Slovenia educating PE teachers in a 5-year specialist program. All schools follow identical testing protocols and use standardized equipment. Results are entered into the “My SLOfit app” where data is first checked with smart algorithms, analyzed, and made available to all registered users, i.e., parents/guardians control access .In Slovenia, the physical fitness of children and adolescents is monitored by a program originally entitled “Sports Educational Card”, implemented in 1982 on a sample of Slovenian schools. After 5 years of refinement and pilot testing, the program was introduced to all Slovenian primary and secondary school curricula in all Slovenian schools . “SLOfitBeginning in 2016, SLOfit researchers developed and launched a free-for-use, online application system called “My SLOfit” . AmongstWe developed a medical screening protocol to mitigate health-risks associated with fitness testing for adults with specific/underlying clinical concerns. SLOfit Lifelong test batteries include preliminary medical screenings performed at rest, before exercise, for:  arterialParticipants completed a warm-up to a prepared video based on age and fitness level of the individual. After warm-up, participants begin the motor tasks. Although testing order is not strictly specified in SLOfit Students or SLOfit Seniors (except that the cardiorespiratory endurance test should be performed last), for SLOfit Adults tests do follow a specific order, namely: vertical jump (leg power), Figure-8 run (agility), handgrip (arm strength), arm plate tapping (coordination), sit and reach (flexibility), partial curl-up (core strength), and a 6-min walk (cardio-respiratory endurance).N = 14–16 senior and junior researchers) at Faculty of Sport, Ljubljana to identify, pilot test and define appropriate fitness tests for adults. Discussions were then held with experts from target end-users, e.g., fitness industry, sports clubs, national public health institute, patients, academia, and educators familiar with SLOfit. The entire test battery development occurred over ~36 months, divided into 5 steps. First, the work group reached consensus on which fitness components needed monitoring. Seven components were prioritized: body composition/adiposity, cardiorespiratory endurance, muscular strength, power, coordination, agility, and flexibility. Second, it was decided to split adult test batteries to encompass advancing age and variable fitness levels. Three distinct batteries were decided for: (a) Young Adults (19–34 y), Adults (35–64 y), and Seniors (65+ y). The group then cross-referenced whether any existing published test batteries fit the specific needs of SLOfit Lifelong and found that none matched the existing SLOfit Student version well enough in terms of breadth or scope of fitness tests included. Thus, step 4 was finding the range of appropriate tests and conducting pilot measurements to assess testing feasibility . In the final step 5, three fitness testing batteries were defined: (a) SLOfit Young Adults (19–34 y) (SLOfit Adults (35–64 y) and (c) SLOfit Senior (65+ y) (A working group was assembled (19–34 y) ; (b) SLO (65+ y) . When idSLOfit Lifelong testing was piloted on each adult group to confirm whether the selected tests were appropriate and determine the best test order for successfully completing all sections with positive vigor. All in-person testing took place at the Faculty of Sport or Faculty of Education, University of Ljubljana, Slovenia, following national guidelines for COVID-19 safety protection.N = 471) and Adults (N = 527) were sampled between May 2021 and June 2022, including ~20 measurement days at the University. Seniors (N = 152) were pilot tested last, from 31.03.2022 to 21.06.2022. On 21.04.2022, measurements were performed at the Coronary Club of Ljubljana with participants having diagnosed cardiovascular disease (s). After this first stage, some tests were modified or improved slightly. Due to problems with feasibility , with the Sorenson test and T-test prompted replacement with “partial curl-up” and “Figure 8 run”, respectively (The test batteries were first piloted on university students (26.04.2021–12.05.2021). Next, Young Adults (ectively . StatistWe performed literature searches for studies published from 1.1.1980) that investigated the ability of a specific test to predict mortality or morbidity  to identify cut-off values related to health risk for each test included. The search was built around two areas: .1.1980 t health-rSince literature reviews failed to identify criterion-referenced standards for most tests, step 2 searched for dose-response relationships between a given fitness component and premature mortality across the life course. Two analyses on male youth found that 20% of people with the lowest cardiorespiratory fitness  and 10% These procedures were repeated for the elderly, based on fitness components included in the SLOfit Senior fitness test battery. The search identified three studies which defined criterion-based standards for Senior fitness tests. We opted to primarily use a large, European-based study . For misAll pilot data collected from adults 19–64 years old were used to construct centile norms for each test item included in the SLOfit Adult test battery. Centile curves were obtained using the Lambda Mu Sigma (LMS) method separately for each gender following methods described elsewhere , 40. To After each measurement, participants enter their results into the application and receive immediate feedback on their physical fitness levels and health risks. Feedback can be interpreted as a relative change from different time-points, or by centile value compared to peers. BMI and fitness data in the My SLOfit application are presented according to health risk based on existing scientific evidence , available to all web users offering program information for different user groups . The My SLOfit website (URL: moj.slofit.org) is a secure web application running a DNN (formerly DotNetNuke) platform. All personal data are longitudinally saved in SQL database. Access to the data is initially granted only to the administrator of SLOfit testing providers . If the participants of SLOfit fitness testing are registered users of MySLOfit, they can also access (their own) fitness data, complete questionnaires, and create reports, e.g., report of fitness testing . In this way, SLOfit is building an online and real-world community. The free online My SLOfit app is customizable for use on different screen sizes . The SLOfit website with the application is educational platform to increase physical and health literacy of children, adolescents, adults, and seniors. My SLOfit app allows individuals of all ages to  manage twww.slofit.org) and free web application My SLOfit (moj.slofit.org), which provides free diagnostic tools, SLOfit reports expert advice in their native language (www.slofit.org/slofit-nasvet), and social media  where citizens can engage directly with fitness, education and health professionals.SLOfit Lifelong encourages individuals to use data for their own exercise planning and to optimize the quality and quantity of their habitual movement behavior. Given the limited possibilities of communication between schools, physicians, municipalities, and parents it was increasingly important for us to establish holistic communication channels for the individual, their households, schools, and community environment. Since 2017/18, SLOfit registered users are already able to access information support through the SLOfit website . Each outlet provides unique tools and tips on how to increase one's health, fitness, and physical literacy. Health literacy is an important determinant of overall health, encouraging individuals to meet the demands of a modern society  informing SLOfit participants about possible weaknesses in their fitness routine, and the importance of regular exercise (b) bond SLOfit participants to competent physical fitness activity providers (c) provide 2-way communication between patients and health care providers (d) enable health care professionals to monitor patient fitness to design appropriate, personalized physical activity programs.SLOfit Lifelong provides a scientific backbone for creating/maintaining population-based policies related to improving physical fitness and activity. In future, causal relationships between children and adolescents' physical performance and future health outcomes can be inferred. Understanding to what extent physical fitness in childhood is a predictor of future disease will improve preventive interventions to increase physical fitness across the lifespan. With these program upgrades, SLOfit Lifelong will allow for detailed secular trends analyses in Slovenian subpopulations at the municipal, regional, and country level.From a research perspective, SLOfit Lifelong will develop a longitudinal cohort infrastructure using the Community-Based Participatory Research (CBPR) approach, one which involves direct participant engagement in which they can be affected by the research questions being addressed. Already, the SLOfit database includes about 8 million sets of measurements of over 1 million children. By enlarging SLOfit to include adulthood fitness surveillance, it is building a unique research platform for future studies to assess the predictive validity of health-related fitness.SLOfit Lifelong will continue as a participatory research platform by adding modules for sharing real world data such as smartwatches, fitness trackers, mobile phones, etc. It will be conceivable to link data from family members, enabling custom analyses, and providing evidence for at-risk, family-based interventions. The SLOfit team is closely following artificial intelligence technology that may allow the collection and analysis of large amounts of data on individual's movement behaviors , which could further improve the predictive models and enhance people's lifelong physical fitness.SLOfit Lifelong has developed unique physical fitness test batteries for adults which are physiologically comparable to those conducted in all Slovenian schools annually. By encasing the SLOfit Lifelong initiative within the existing SLOfit framework, web platform and infrastructure, researchers now have the capacity to store lifelong health and fitness data, including calculating health risk assessments, based on fitness data supplied by the citizen. Everyday people can follow their own fitness and health status, with expert feedback, to increase their physical literacy and fortify a sense of personal responsibility toward their own health outcomes.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by Slovenian National Medical Ethics Committee (ID: 0120-468/2021/3). The patients/participants provided their written informed consent to participate in this study.GJ: project administration, conceptualization, investigation, resources, writing- original draft, and writing- review and editing, writing- approving final submission. SM and VSe: writing- original draft, writing- review and editing, and writing- approving final submission. MS: conceptualization, writing- original draft, writing- review and editing, and writing- approving final submission. BL: conceptualization, investigation, resource, writing- original draft, writing- review and editing, and writing- approving final submission. MK: writing- review and editing and writing- approving final submission. TO: writing- original draft, investigation, resource, writing- review and editing, and writing- approving final submission. JK, KM, PG, VSt, and VH: investigation, resource, and writing- approving final submission. RB: investigation, resource, writing- review and editing, and writing- approving final submission. NM: investigation and writing- approving final submission. GS: conceptualization, investigation, resource, writing- review and editing, and writing- approving final submission. All authors contributed to the article and approved the submitted version.The SLOfit Lifelong initiative was co-funded by the Slovenian National Research Agency (J5-1797) and limited, non-specific funding from the research program group P5-0142 .The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Patent ductus arteriosus (PDA) is one of the most common congenital heart diseases. Once the PDA is diagnosed, it needs to be dealt with in time. At present, main methods include pharmacological treatment, surgical closure, and interventional closure for treatment of PDA. However, the effect of different interventions in PDA management is still controversial. Thus, our study aims to assess the effectiveness of different interventions together and estimate the sequence of these therapies for PDA children. Meanwhile, it is necessary to conduct a Bayesian network meta-analysis to compare the safety of different interventions comprehensively.To the best of our knowledge, this is the first Bayesian network meta-analysis comparing the efficacy and safety of different interventions for the treatment of PDA. PubMed, Embase, Cochrane Library, Web of Science, gray literature, and trial registry databases were searched from inception to December 2022. We will extract and report data according to methodological guidelines for Bayesian network meta-analysis by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocols (PRISMA-P). Primary PDA closure, overall PDA closure, technical success, surgical success rate, mortality during hospital stay, operation time, intensive care unit stay, intraoperative radiation dose, radiation exposure time, total postoperative complication rate, and postoperative major complication rate will be defined as the outcomes. The quality of all random studies will be assessed using ROB, and quality of evidence for all outcomes will be judged by using the Grading of Recommendations Assessment, Development and Evaluation (GRADE).The results will be disseminated through peer-reviewed publication. Since no private and confidential patient data will be contained in the reporting, there are no ethical considerations associated with this protocol.INPLASY2020110067.The online version contains supplementary material available at 10.1186/s13643-023-02195-4. This paper will be the first study to compare the efficacy and safety of surgical procedures for PDA patients using Bayesian network meta-analysis.We will use the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach to evaluate the quality of evidence.The results of this study will help surgeons and patients to select appropriate treatments.Our results will be limited by both the quantity and quality of the trials available for review.Congenital heart disease (CHD) is the most prevalent human birth malformation with an incidence of one approximately in 100 newborns. CHD is also a major cause of perinatal and infant mortality, with about 220,000 deaths worldwide each year . If it iCurrently, three pharmacological agents—ibuprofen, acetaminophen, and indomethacin—are used for PDA closure in preterm infants . IndometAt present, some pairwise meta-analyses have compared the efficacy and safety of some procedures. Network meta-analysis (NMA) can estimate the relative effectiveness of all interventions and the sequence of interventions, even in the absence of a head-to-head comparison of all interested interventions . Thus, oBayesian network meta-analysis will be carried out. This protocol is reported according to the Preferred Reporting Items for Systematic Review and Meta-Analysis Protocols (PRISMA-P) , and theWe will perform electronic search until December 2022 in the following databases: PubMed, Embase, Cochrane Library, and Web of Science. We will also search ClinicalTrials.gov, WHO International Clinical Trials Registry Platform (ICTRP), and International Standard Randomized Controlled Trial Number Registry for all registered clinical trials and randomized controlled trials. Furthermore, we will search the gray literature in Google Scholar, OpenGrey, and ProQuest Dissertation. The reference lists of articles and relevant systematic reviews will be tracked to identify other relevant studies. No restrictions on language will be set. If we find more relevant key words during our primary search, we will modify the search syntax. All searches will be updated before submitting the review.#1 “Ductus Arteriosus, Patent” [MeSH Terms]#2 “patent ductus arteriosus” [Title/Abstract]#3 #1 OR #2#4 "Thoracic Surgery, Video-Assisted "[MeSH Terms] OR "video-assisted thoracoscopicThe PubMed search strategies are as follows:surgery"[Title/Abstract] OR "Minimally Invasive Surgical Procedures"[MeSH Terms] OR "minimally invasive"[Title/Abstract] OR "Minimal Access Surgical Procedures"[Title/Abstract] OR "mini-invasive"[Title/Abstract] OR "minimal surgical procedure*"[Title/Abstract] OR "Surgical ligation"[Title/Abstract] OR "surgical closure"[Title/Abstract] OR "Transthoracic"[Title/Abstract] OR "transcatheter"[Title/Abstract] OR "percutaneous closure"[Title/Abstract] OR "pharmacological"[Title/Abstract] OR "Cyclooxygenase Inhibitors"[MeSH Terms] OR "acetaminophen"[MeSH Terms] OR "COXI"[Title/Abstract] OR "Indomethacin"[Title/Abstract] OR "indometacin"[Title/Abstract] OR "indocid"[Title/Abstract] OR "ibuprofen"[Title/Abstract] OR "brufen"[Title/Abstract] OR "motrin"[Title/Abstract] OR "paracetamol"[Title/Abstract]. #5 #3 AND #41) Types of study: We will include randomized controlled trials. The number of cases included in each group must be greater than ten. Retrospective cohort studies, prospective cohort studies, case–control studies, and case series will not be included.2) Types of participants: PDA children were confirmed clinically and by transthoracic echocardiography and scheduled for pharmacological treatment, surgical closure of PDA, or interventional closure. There were no restrictions in children weight and age. PDA diameter ranged from 2 to 5.5 mm.3) Types of interventions: Pharmacological treatment , mini-thoracotomy, VATS, fluoroscopy-guided transfemoral venin and artery of percutaneous closure, fluoroscopy combined with TTE-guided transfemoral venin and artery of percutaneous closure, TTE-guided transfemoral artery of percutaneous closure, and TTE-guided transfemoral vein of percutaneous closure.4) Types of comparison groups: Surgical ligation for surgical interventions and placebo for pharmacological treatment. If there are head-to-head studies, these studies which compare surgical ligation with other mini-invasive treatments, treatment between different medicines, or pharmacological with surgical treatments will be included.5) Types of outcomes.The primary outcomes include the following: primary PDA closure (defined as echocardiography-confirmed closure of PDA after the first course of the treatment), overall PDA closure (defined as echocardiography-confirmed closure of PDA after one or more courses of the treatment), and technical success  with a device or coil in the PDA. Cases were also considered successful in the event of coil/device embolization that was retrieved during the same procedure, and the PDA was closed with a different size of device/coil.)The secondary outcomes include the following: mean days/hours needed for closure of PDA, surgical success rate, mortality during hospital stay, operation time, intraoperative radiation dose, radiation exposure time, total postoperative complication rate, postoperative major complication rate, intensive care unit stay, total hospital stays, postoperative hospital stays, and total cost.Children were diagnosed with other congenital heart diseases .Those studies which assessed the change of cardiac function and cardiac physiology changes after PDA treatmentNo treatment (conservative management)Literature search records will be imported into EndNote X8.1 (Thomson Reuters (Scientific) LLC Philadelphia, PA, USA) software. First, the EndNote X8.1 will be used to exclude duplicate articles, and then, two reviewers will independently screen the titles and abstracts and exclude the articles that do not fulfill the inclusion criteria. If it is hard to judge, full texts will be retrieved. After then, two reviewers will cross-check whether the final selected research is consistent. If there are any disagreements between two reviewers during the selection of eligible studies, a third reviewer will involve resolving the disagreements.www.microsoft.com). Then, two independent reviewers will extract the basic characteristics and the data of outcomes. The extracted data will include the following: authors, gender, age, birthweight, year of publication, study design, inclusion criteria, medicine dosing protocol, sample size, primary closure rate, reopening rate, mortality, intraventricular hemorrhage , necrotizing enterocolitis , pulmonary hemorrhage (blood-stained liquid flowing from the trachea of the infant), bronchopulmonary dysplasia, retinopathy of prematurity, renal dysfunction, hepatic dysfunction, gastrointestinal bleeding, treatment methods, device used, median follow-up, PDA diameter, etc. If there is a discrepancy between the two reviewers, a third researcher will be consulted. We will randomly select five to ten studies to check the completeness of the data abstraction form. The data abstraction form will be complemented by the pilot trial. We will contact the corresponding author to query information when the essential data are insufficient or missing in the original study through sending an email. Studies will be excluded if we are unable to get access to the data, and the reasons for exclusion will be reported in detail.A standard data abstraction form will be created using Microsoft Excel 2019  assessment tool . StudiesA network plot will be created to describe and present the geometry of the treatment network of comparisons across trials using STATA . If the trial is not linked by treatments, we will exclude it from network meta-analysis and just describe the findings of the study.In the network plot, nodes represent different interventions, and edges represent a head-to-head comparison between interventions. The size of the nodes and the thickness of the edges are associated with the sample size of the intervention and the number of trials included, respectively.For pharmacological treatment, a network will be created. The pharmacological treatment includes acetaminophen, indomethacin, and ibuprofen. For surgical treatment, the network includes surgical ligation, mini-thoracotomy, VATS, and interventional therapy. Finally, a network of pharmacological and surgical treatment will be created if possible.Q-test and I2 statistics to assess and report heterogeneity. When I2 ≤ 40%, the heterogeneity is considered acceptable, and the fixed-effects model is used for the combined analysis. If I2 > 40%, the heterogeneity is considered to be large, and the source of heterogeneity will be explored. Sensitivity analysis, subgroup analysis, and meta-regression will be used to find the source of heterogeneity. Meanwhile, the clinical and methodological heterogeneity across studies will be reassessed. If no proper reasons will be found for high heterogeneity, the random effects model will be used to incorporate the heterogeneity. P < 0.05 was considered as statistically significant.We will use Review Manager 5.3  statistical analysis software for pairwise meta‐analysis. For dichotomous outcomes, the combined results will be calculated as odds ratios (ORs) and 95% confidence intervals (CIs). For continuous outcomes, we will calculate and meta-analyze mean differences (MD) or standardized mean differences (SMD) and their 95% CIs. We will use the Cochran’s P < 0.05, the inconsistency model will be used for pooled analysis. Meanwhile, the consistency model will be used [http://cran.r-project.org/) with interfacing to OpenBUGS  (http://www.openbugs.net/w/Downloads) [Network meta-analysis will be conducted using a Bayesian Markov chain Monte Carlo (MCMC) framework. We will use the generalized linear modelling framework for network meta-analysis of randomized controlled trials. The convergence will be assessed using the Brooks-Gelman-Rubin (BGR) plots method. Both random and fixed models will be used. And the best model will be chosen based on the deviance information criterion (DIC) values. The model with lowest DIC value is usually selected because it is considered in most cases to be the most reliable. Transitivity is a key assumption of NMA and refers to the belief that an indirect comparison is a valid estimate of the unobserved direct comparison , 31. The be used , 34. The be used . If the  be used . The MCMwnloads) –39.Participant characteristics Study-specific factors are as follows:(a) Sample size(b) Overall study quality(c) Risk of bias According to the instrument to assess the Credibility of Effect Modification Analyses (ICEMAN) tool , subgrouThe funnel plot and Egger test will be conducted to detect publication bias if the number of studies was more than ten.We will assess the quality of the evidence by following the steps to assess the quality of treatment effect estimates for network meta-analysis as per the Grading of Recommendations Assessment, Development, and Evaluation (GRADE)  approachWe will perform a sensitivity analysis to verify the robustness of the study results. This will be achieved by assessing the impact of the sample size, high risk of bias, missing data, and selected models. Following the analyses, if the quality of a study is judged to be low, it will be removed to ensure the robustness of the results.Ethical approvals and patient consent are not required because this is a network meta-analysis based on published trials. The findings of this project will provide a general review and evidence of the efficacy and safety of pharmacological treatment, surgical ligation, mini-thoracotomy, VATS, fluoroscopy-guided transfemoral venin and artery of percutaneous closure, fluoroscopy combined with TTE-guided transfemoral venin and artery of percutaneous closure, TTE-guided transfemoral artery of percutaneous closure, and TTE-guided transfemoral vein of percutaneous closure for PDA in children. The results will be submitted to a peer-reviewed journal for publication. We hope that these findings will help clinicians and patients choose a more appropriate repair method for PDA.Additional file 1: PRISMA-P (Different surgical procedures for the treatment of patent ductus arteriosus in children: a protocol for a network meta-analysis*


Background: Necroptosis is closely related to tumorigenesis and development. Accumulating evidence has revealed that long non-coding RNAs (lncRNAs) are also central players in osteosarcoma (OS). However, the role of necroptosis-related lncRNAs in OS remains unclear. In the present study, we aim to craft a prognostic signature based on necroptosis-related lncRNAs to improve the OS prognosis prediction.Methods: The signature based on necroptosis-related lncRNAs was discovered using univariate Cox, least absolute shrinkage and selection operator (LASSO), and multivariate Cox regression analysis. The prognosis efficiency of the signature was then estimated by employing various bioinformatics methods. Subsequently, immunological analysis and Gene Set Enrichment Analysis (GSEA) were used to explore the association between necroptosis-related lncRNAs with clinical outcomes and immune status. More importantly, several necroptosis-related lncRNAs were validated with RT-qPCR.Results: Consequently, a novel prognosis signature was successfully constructed based on eight necroptosis-related lncRNAs. Meanwhile, the novel necroptosis-related lncRNAs model could distribute OS patients into two risk groups with a stable and accurate predictive ability. Additionally, the GSEA and immune analysis revealed that the necroptosis-related lncRNAs signature affects the development and prognosis of OS by regulating the immune status. The necroptosis-related lncRNA signature was closely correlated with multiple anticancer agent susceptibility. Moreover, the RT-qPCR results indicated several necroptosis-related lncRNAs were significantly differently expressed in osteosarcoma and osteoblast cell lines.Conclusion: In this summary, a novel prognostic signature integrating necroptosis-related lncRNAs was firstly constructed and could accurately predict the prognosis of OS. This study may increase the predicted value and guide the personalized chemotherapy treatment for OS. Osteosarcoma (OS) is the most prevalent primary bone cancer in children and adolescents, caused by mesenchymal stem cells or osteoclasts . OS accoNecroptosis, a new form of cell death resembling, is characterized by three essential proteins: receptor-interacting protein kinase 1/3 (RIP1 and RIP3) and mixed lineage kinase-like protein (MLKL) . As the In this study, we aimed to screen out independent prognostic necroptosis-related lncRNAs based on the gene expression data and clinical information of OS in public databases and constructed a novel necroptosis-related lncRNA prognostic signature for OS, providing insights for evaluating the clinical prognosis of OS. Additionally, we tentatively explored the potential molecular mechanism of necroptosis-related lncRNAs included in the prognostic models through Gene Set Enrichment Analysis (GSEA). Finally, immune infiltration and drug sensitivity analysis were utilized to further recognize the connections between necroptosis and tumor immune infiltration status, improving the understanding of tumor microenvironment and OS treatment. Hence, the results of our study would provide new insights for exploration of the molecular mechanism of OS pathogenesis and development, as well as provide a new therapeutic strategy for OS treatment in the future.https://ocg.cancer.gov/programs/target)  (https://xenabrowser.net/) and normalized by log2 (x+1) (The RNA sequencing (RNA-seq) and the corresponding clinical characteristics of OS come from The Therapeutically Applicable Research to Generate Effective Treatments database  , and theg/home/) . The trag2 (x+1) . In tota2| > 0.3 and p < 0.001 were considered as necroptosis-related lncRNAs . Subsequently, they were incorporated into LASSO and the multivariate COX regression analysis to seek independent prognostic necroptosis-related lncRNAs for constructing a precise prognosis signature. The risk score was calculated as n” represents the number of pyroptosis-related lncRNA, coef (lncRNA k) represents the coefficient of each lncRNA in the signature, and exp (lncRNA k) represents the lncRNA expression level. The risk score was obtained by multiplying and accumulating the two groups. Patients with OS were separated into high- and low-risk groups according to their median risk score. The predictive value was estimated using Kaplan-Meier (K-M) survival analysis. The Receiver operating characteristic (ROC) curve was drawn, and the area under curve (AUC) value was calculated to assess the predictive power of the prognostic signature.To explain the prognostic significance of necroptosis-related lncRNAs systematically and comprehensively in OS, we employed 84 patients with overall survival data to build prognostic signatures. For OS patients, we extract survival data, including survival time and survival status, and merge them with the expression files of necroptosis-related lncRNAs. The necroptosis-related lncRNAs associated with overall survival were screened out through univariate COX regression analysis and drew a forest plot , gender, and metastatic status. To further evaluate the independent predictive ability of the novel model, the K-M survival curve was conducted one by one to compare the prognosis differences of two risk groups in each subgroup.To further clarify the association between the novel signature and the prognosis of OS, we constructed a nomogram and corresponding calibration curves. A nomogram that included different variables, such as age, gender, and metastasis, was established to predict the survival possibility of OS patients after 1-, 2-, and 3-years. The calibration curve was used to validate the nomogram’s prediction performance between different patient groups. The R software package “rms” was utilized to build nomograms and calibration curves.P< 0.05 and false discovery rate (FDR) q < 0.25.GSEA can calculate the enrichment score of a specific gene set based on the expression level of each gene in the gene expression matrix, thereby identifying the pathway enrichment of the different groups . The MolEstimation of Stromal and Immune cells In Malignant Tumor tissues using Expression (ESTIMATE) is a method to infer the fraction of stromal cells and immune cells in tumors based on the expression of characteristic genes in transcriptome data and is widely used to assess tumor purity . The R pThe related survival data is merged with the amount of immune cell infiltration to investigate the association between the novel prognostic signature and immune cell infiltration. Finally, we also compared the expression of immune checkpoints in the high- and low-risk group. The potential immune checkpoint was obtained from the previous report .Currently, the “pRRophetic” R package is commonly implemented for chemotherapeutic response prediction . To expl2 in a humidified incubator.Human osteosarcoma cell lines 143B, U2OS, HOS, and normal osteoblast cell line hFOB1.19 were obtained from the American Type Culture Collection (ATCC). All the cell lines were cultured in Dulbecco’smodified Eagle’s medium  with 10% fetal bovine serum  and 1% penicillin-streptomycin solution  at 37°C with 5% COTotal RNA was extracted with RNA Express Total RNA Kit . Then, the Revert Aid First Strand cDNA Synthesis Kit  was used for cDNA synthesis. Subsequently, RT-qPCR was performed with Hieff qPCR SYBR Green Master Mix (High Rox Plus) . The primers used for the validation are listed in All statistical analyses in this study were conducted using R statistical software (Version 4.0.5). The Wilcox test analyzed differentially expressed necroptosis-related lncRNAs between osteosarcoma and normal tissues. K-M curve was performed for survival analyses, and the log-rank test was applied to analyze differences between subgroups. Univariate and multivariate Cox regression analyses were used to identify independent prognostic factors. The ROC curve was drawn to examine the sensitivity and specificity of the proposed predictive model for OS. The “Rtsne” R package was used to perform principal component analysis (PCA) and T-distributed stochastic neighbor embedding (t-SNE).The flow diagram of our study is presented in The 218 differentially expressed necroptosis-related lncRNAs were used for subsequent prognostic model construction. First, we performed a univariate Cox regression analysis to identify necroptosis-related lncRNAs connected with OS prognosis. The results discovered that 34 differentially expressed necroptosis-related lncRNAs were closely connected with the overall survival of patients with OS . Then, tp < 0.001 ). Similarly, the risk score curve and survival status plots indicated that the OS patients in the low-risk group had a better prognosis, with fewer deaths and longer survival time , gender, and metastasis status and analyzed the survival differences between the high- and low-risk groups in different subgroups. The results indicated that OS patients in the low-risk group had a better clinical prognosis than the high-risk group among different ages, genders, and metastatic status . MoreoveIncorporating the risk score and clinical parameters, we constructed a nomogram for predicting 1-, 2-, and 3-years overall survival. As shown in The GSEA was performed to investigate the potential process and pathway involved in molecular heterogeneity between the two risk groups. p > 0.05). This supported the observation that the degree of immune infiltration in the low-risk group was higher than in the high-risk group.To explore the roles of necroptosis-related lncRNAs signature in the tumor immune microenvironment, we further analyzed the association between the immune status and the risk score in OS. First, we calculated the immune score of OS through ESTIMATE and compared its correlation with the risk score and survival prognosis. The immune, stromal, and estimation scores were all greater in the low-risk group than in the high-risk group, but tumor purity was lower in the low-risk group than in the high-risk group . SimilarIn addition to analyzing the correspondence scores of the immune microenvironment, it is also necessary to understand the proportion of the immune landscape of OS. We performed the CIBERSORT algorithm to explore the proportion of immune cells in OS. As shown in Additionally, we compared the susceptibility of OS to commonly used anticancer drugs between the two distinct risk groups to identify potential OS treatment agents. We discovered that the OS patients in the low-risk group might positively react to Imatinib, Midostaurin, Pazopanib, Bexarotene, Shikonin, Dasatinib, Bortezomib, and Bryostatin, while patients with high-risk scores maybe respond better to Lenalidomide, Camptothecin, Elesclomol, and Metformin . Under tThe expression level of these signature necroptosis-related lncRNAs in cell lines  were further detected through RT-qPCR. As presented in OS is the most common bone malignancy in children and adolescents . AlthougA total of 67 necroptosis-related genes were obtained from previous studies, and 218 differentially expressed necroptosis-related lncRNAs were identified through Pearson correlation analysis and difference analysis. Then, we screened eight necroptosis-related lncRNAs for the subsequent signature establishment based on univariate Cox regression analysis, Lasso regression, and multivariate Cox regression analysis. Among these prognostic necroptosis-related lncRNAs in OS, some of them have been demonstrated to play pivotal roles in carcinogenesis and progression. For instance, the lncRNA VPS9D1-AS1 was significantly upregulated in esophageal squamous cell carcinoma, which can promote the malignant progression of esophageal squamous cell carcinoma through the Wnt/catenin signaling pathway . In coloTo systematically analyze the prognostic accuracy of necroptosis-related lncRNAs in OS, we conducted a novel prognosis signature based on these eight necroptosis-related lncRNAs. The patients were split into two risk groups according to the risk scores, and various bioinformatics analyses were performed. First, our survival analysis results indicated that the survival prognosis of patients in the high-risk group is significantly lower than that in the low-risk group. The survival analysis results between different clinical subgroups are also consistent with that. Meanwhile, the 1-year, 2-years, and 3-years ROC curves confirmed the accuracy of the novel signature in survival prediction. Additionally, univariate and multivariate analyses further verified that the risk model could be an independent prognostic factor for OS. Finally, the necroptosis-related prognostic nomogram and calibration curve demonstrated that the signature could accurately predict OS patient’s prognosis. Consequently, these above results revealed that the novel prognostic risk signature based on these eight necroptosis-related lncRNAs has better predictive performance and universal applicability.via the Hedgehog signaling pathway (After that, to further explore the association between the novel necroptosis-related lncRNAs signature and OS GSEA was utilized, and the results revealed that the high-risk group significantly enriched several cancer-related signaling pathways, including DNA replication, Hedgehog signaling pathway, and cell cycle. Accumulated evidence has demonstrated a vital role for these signaling pathways in the development and progression of OS. To name a few, microRNA-524-5p inhibits the proliferation of OS cells and induces cell cycle arrest by targeting CD6 . URG4, w pathway . On the  pathway . For ins pathway . The nov pathway . Additio pathway . For exa pathway . Togethe+ T cells play a critical role in anti-tumor immunity. High CD8+ T cell infiltration levels are significantly related to the better prognosis of various cancers, including OS (To further explore the relationship between the necroptosis-related lncRNAs signature and the immune status of osteosarcoma, we performed ESTIMATE, ssGSEA, and CIBERSORT algorithms to analyze the immune infiltration status and the tumor microenvironment in O.S. First, the ESTIMATE analysis indicated that the overall immune score of OS was negatively correlated with the risk score and was correlated with the prognosis of patients with OS Similarly, the ssGSEA showed that most of the immune cell infiltration and corresponding immune functions in the low-risk group were better than those in the high-risk group. This was in line with the results of several previous research. Xinxin Bu et al. showed a lower immune infiltration score in the high-risk group of pyroptosis-related lncRNAs signature for OS . Ting Leuding OS . Previouuding OS . Finallyuding OS . Hence, via MMP13 inhibition (Subsequently, we determined the drug sensitivity to candidate anticancer drugs in the two risk groups. Our results revealed that Imatinib, Midostaurin, Pazopanib, Bexarotene, Shikonin, Dasatinib, Bortezomib, and Bryostatin showed better responses in the low-risk score group. In contrast, Lenalidomide, Camptothecin, Elesclomol, and Metformin in the high-risk group showed better response. In the past, it has been shown that the application of imatinib and adriamycin had a synergistic antiproliferative effect in platelet-derived growth factor receptor-expressing OS cells . Shikonihibition . Equallyhibition . Overallvia regulating the immune status. It is reasonable to believe that our study provides new insight into prognosis prediction and individualized chemotherapy therapy for OS.In summary, our study constructed a novel necroptosis-related lncRNAs signature, which could effectively predict the prognosis of OS. The signature could be served as an independent prognostic factor for OS. Furthermore, the GSEA and immune analyses revealed that these necroptosis-related lncRNAs affect the OS prognosis


While antiviral strategies are vulnerable to the effects of viral mutation, the relatively conserved Mpro makes an attractive drug target: Nirmatrelvir, an antiviral targeting its active site, has been authorized for conditional or emergency use in several countries since December 2021, and a number of other inhibitors are under clinical evaluation. We analyzed recent SARS-CoV-2 genomic data, since early detection of potential resistances supports a timely counteraction in drug development and deployment, and discovered accelerated mutational dynamics of Mpro since early December 2021.The current coronavirus pandemic is being combated worldwide by nontherapeutic measures and massive vaccination programs. Nevertheless, therapeutic options such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main-protease  near the active site since December 2021.The analysis showed a significant recent event of mutational dynamics in MThe increased mutational variability in close proximity to an antiviral-drug binding site as described herein may suggest the onset of the development of antiviral resistance. This emerging diversity urgently needs to be further monitored and considered in ongoing drug development and lead optimization. Still, therapeutic options such as the SARS-CoV-2 main-protease (Mxlovid™) , generalAs mutation rates are estimated to be moderate to high in coronaviruses , treatmepro  were neglected in order to ensure the correct mapping of an exchange to its position within the sequence. Mutations not directly adjacent to deletions, insertions, or “X” were neglected if the surrounding region of 10 residues on each side of the potential exchange showed more than 50% mismatches to the wild-type sequence in combination with at least one deletion, insertion, or “X.”Time-resolved analysis of mutation events was performed by assigning the collection dates of genome sequences retrieved from GISAID  to the rA python workflow was used to combine and compile sequence-collection information with mutation lists as we extracted them from the MSA file. Dates of first occurrences of unique amino-acid exchanges were identified by comparing all dates for any particular unique amino-acid exchange at a given position in a given protein. The results were saved to disk for further analysis and as input for visualization.Visualizations in pro sequence.The conservation and variability across aligned protein-sequence sites are computed by two approaches, (1) frequency-based and (2) amino-acid’s physicochemical-property-based scoring methods , 40. Fori,j of each amino acid j at a position i in the sequence, while P(exp) is the expected probability of an amino acid j in a protein sequence. N denotes the number of unique amino acids present at a position.Pairwise alignment has been used as an input for calculating the probability Ppro, the B factors within PDB entry 7SI9  in new unique mutations within a region of the Mpro sequence corresponding to the target site of current antiviral agents such as the protease inhibiting component of Paxlovid™, nirmatrelvir.We investigated the mutational dynamics of the SARS-CoV-2 Mt GISAID  by 30 Jupro, with an average number of 5.9 ± 2.1 exchanges per residue  until June 2022. However, a specific region of eight amino acids (residues R188-G195) revealed an unprecedented average number of 14.8 ± 1.3 exchanges per residue. A time-resolved analysis  over time is illustrated in pro sequence  as well as a measure of disorder along the protein sequence derived from PDB entry 7SI9  of Mpro, namely six residues each at the C-terminus of NSP4, the C- and N-termini of NSP5-NSP15 as well as the N-terminus of NSP16 , the net effect for the virus might still be advantageous.For decades, multidrug-resistant bacteria have been posing a severe threat. Antiviral-drug resistance is equally dangerous, in particular in the examples of resistant human immunodeficiency viruses, influenza, herpes, or hepatitis strains, which pose a serious threat to public health. Consequently, the acquisition of mutations reducing the susceptibility and clinical activity of antiviral drugs has been a perpetual hurdle in developing effective antiviral therapies –66. Whilpro expressed in yeast resulted in low mutational sensitivity of residues 188-191 and 193-195, meaning that the protease allowed several amino-acid exchanges at these positions without significant loss of function, and, in contrast, high sensitivity at position 192  resulted in decreased susceptibility to nirmatrelvir when expressed in Escherichia coli  in order to be aware of the potential development of antiviral-drug resistance at an early stage. For the establishment of resilient drugs with long-term efficacy, efficient binding of the inhibitor should not rely on interaction with residues which are insensitive to mutation. For instance, as the current situation indicates, essential interactions of the inhibitor to side chains of residues R188-G195 should be avoided. Clearly, it is of equal importance to gain structural insights, examine pathways of resistance, and constantly investigate new classes of drugs to target multiple key factors of the viral infection machinery.It is therefore of great importance to monitor a possible global increase in the occurrence of the herein-described new unique mutations near the active site of Mpro’s mutational dynamics, it had not occurred—not even partially. On the contrary: diversity at the positions of interest had already been stagnant over a long period of time, cf. the date range from April 2020 to November 2021 in For coronaviruses it is still uncertain whether drug-resistant variants will persist or evolve into globally prevalent dominant variants. VOCs like Alpha, Delta, Omicron, and others are mainly characterized by new spike-RBD variants. This is most likely due to evolutionary pressure from tropism and host adaptation as well as immune- and vaccine-evasion , which cpro variants in complex with its natural substrate and inhibitors. Evolutionary studies with remdesivir-treated (pro; a more thorough investigation of the other SARS-CoV-2 proteins belongs to the realm of future research.While analyzing this data set, we have come across interesting starting points for future research, which we plan to pursue and report elsewhere. These include a quantitative model of the in-host evolutionary dynamics of coronaviruses under the pressure of a targeted antiviral drug, accompanied by structural-bioinformatic studies of M-treated  and untr-treated , 73 patihttps://www.gisaid.org/. An acknowledgment table including accession numbers and the origin of the processed sequences is accessible at https://doi.org/10.6084/m9.figshare.20303979. The genome sequence herein used as a wild-type reference is available at https://www.ncbi.nlm.nih.gov/nuccore/NC_045512.2. Additional supporting data is provided in the https://github.com/innophore/virus.watch-mpro.Publicly available genome sequences were downloaded from LP prepared and evaluated data, performed sequence analysis, contributed to protein structure modeling and structural analysis, and drafted the manuscript with input from all authors. AK supervised data analysis, created various visualization graphs, and drafted the manuscript with input from all authors. TS assisted in data analysis. AS performed sequence analysis regarding Shannon entropy and surprisal indices. KT and KK supported the analysis of structural aspects, druggability information, and comparison to other viral models. MH and KG contributed to structural models, structural analysis, and helped interpret the structural consequences of the observed mutations. GS contributed to evaluating, preparing, and interpreting the data. CG contributed to evaluating, preparing, and interpreting data, drafting the manuscript and designed, managed, and supervised the project. All authors edited the manuscript to its final form.