AlphaFold2 assists in providing novel mechanistic insights into the interactions among the LUBAC subunits.

The linear ubiquitin chain assembly complex (LUBAC) is the only known E3 ligase complex in which the ubiquitin-like (UBL) domains of SHARPIN and HOIL-1L interact with HOIP to determine the structural stability of LUBAC. The interactions between subunits within LUBAC have been a topic of extensive research. However, the impact of the LTM motif on the interaction between the UBL domains of SHARPIN and HOIL-1L with HOIP remains unclear. Here, we discover that the absence of the LTM motif in the AlphaFold2-predicted LUBAC structure alters the HOIP-UBA structure. We employ GeoPPI to calculate the changes in binding free energy (ΔG) caused by single-point mutations between subunits, simulating their protein-protein interactions. The results reveal that the presence of the LTM motif decreases the interaction between the UBL domains of SHARPIN and HOIL-1L with HOIP, leading to a decrease in the structural stability of LUBAC. Furthermore, using the AlphaFold2-predicted results, we find that HOIP (629‒695) and HOIP-UBA bind to both sides of HOIL-1L-UBL, respectively. The experiments of Gromacs molecular dynamics simulations, SPR and ITC demonstrate that the elongated domain formed by HOIP (629‒695) and HOIP-UBA, hereafter referred to as the HOIP (466‒695) structure, interacts with HOIL-1L-UBL to form a structurally stable complex. These findings illustrate the collaborative interaction between HOIP-UBA and HOIP (629‒695) with HOIL-1L-UBL, which influences the structural stability of LUBAC.

Application of 3D printed pelvic fracture related urethra and surrounding tissue as preoperative planning model.

OBJECTIVE: Urethral stenosis caused by pelvic fracture urethral injury (PFUI) is a complex urological disease, especially for the redo cased. However, to find the proximal end of the posterior urethra, and to avoid injury to the rectum and to forecast to remove the inferior pubic margin are two key points for a successful surgery. These steps can be challenging for even the most experienced urologists. This study is to describe a new technique for understanding the three-dimensional (3D) anatomy of the urethra, which will also aid in surgical planning and simplify urethroplasty. MATERIALS AND METHODS: Three patients underwent routine urethroscopy, X ray urethrography and contrast CT urethrography. The 3D images were then reconstructed, and the data were transmitted to a 3D printer. 3D models were printed with polyacrylic acid to simulate the anatomical structure and relationship of urethral stenosis with pubic symphysis and rectum. Various diagnosis methods were compared with the condition in surgery. The patients and trainee questionnaires were performed. RESULTS: Three models of urethral CT were obtained. These models were presented to patients and trainee doctors along with routine urethroscopy, urethrography, and urethral CT. The scores of patients and trainee question forms demonstrated that the 3D printed urethral stenosis model of pelvic fracture has obvious advantages in urethral adjacency and ease of understanding. The 3D printed urethras were easy to show the pubic symphysis and simulate its excision and exposure of urethra. The model could show the precise distance from urethra to rectum to prevent the rectum injury in surgery. CONCLUSIONS: 3D printing technology can be applied to the preoperative evaluation of urethral stenosis caused by PFUI. It can be auxiliary to understand the anatomical structure of the posterior urethra, the direction of urethral displacement, protecting the rectum and the forecasting for pubectomy. It is especially helpful for the accurate preoperative planning of some complex urethral stenosis and redo cases.

The effects of carvacrol on development and gene expression profiles in Spodoptera frugiperda.

The fall armyworm, Spodoptera frugiperda, is a highly polyphagous agricultural pest that is widely distributed around the world and causes severe crop yield loss. Carvacrol showed adverse effects on many pests, such as larval death and growth inhibition. While the effects of carvacrol on S. frugiperda larvae are not yet known. In this study, the effects of carvacrol on S. frugiperda, including larval growth inhibition and mortality induction, were observed. The detoxification and digestive enzyme activities of larvae with 1.0 and 2.0 g/kg carvacrol treatments were analyzed. Carvacrol boosted the enzyme activities of carboxylesterase (CarE) and glutathione S-transferase (GST) while decreasing the activities of α-amylase (AMS), lipase (LIP), and trypsin. A total of 3422 differentially expressed genes were identified in the larvae treated with 2.0 g/kg carvacrol, of which the DEGs involved in xenobiotic detoxification, food digestion, and insecticidal targets were further examined. These results suggest that carvacrol could regulate growth and development by affecting the process of food digestion, and exert its toxicity on the larvae through interaction with a variety of insecticidal targets. While the altered expressions of detoxification enzymes might be related to the detoxification and metabolism of carvacrol. Our findings offer a theoretical foundation for the use of carvacrol for S. frugiperda control in the field.

Characterization of the physiological, histopathological, and gene expression alterations in Spodoptera frugiperda larval midguts affected by toosendanin exposure.

The fall armyworm, Spodoptera frugiperda, is a polyphagous pest worldwide and feeds on many grain and cash crops, which threatens the safety of agriculture and forestry production. Toosendanin (TSN) is a commercial insecticidal active ingredient used to manage various pests in the field and showed adverse effects against S. frugiperda, while the effects of TSN on the larval midguts are not yet known. In this study, the effects of 10 and 20 mg/kg TSN exposures on the larval midguts were analyzed. The structural changes of the larval midgut induced by TSN treatments were also determined by hematoxylin-eosin staining. Besides, TSN treatments also changed the enzyme activities of three digestive enzymes (α-amylase, lipase, and trypsin) and two detoxification enzymes (CarE and GST). A total of 2868 differentially expressed genes (DEGs) were identified by RNA-Seq in the larval midguts with 20 mg/kg TSN treatment, and the DEGs responsible for food digestion and detoxification were further examined. Our findings revealed the preliminary modes of action of TSN on the larval midguts of S. frugiperda, which provide a preliminary rationale for controlling S. frugiperda with TSN in the field.

Effects of removing a highly conserved disulfide bond in ubiquitin-associated domain of human HOIP on biochemical characteristics.

Disulfide bond formed between the cysteine pairs plays a key role in maintaining the integrity of the protein structure and function. The ubiquitin-associated (UBA) domain of human HOIP contains three cysteine residues, Cys504, Cys551, and Cys572. Disulfide bonds formed by Cys504 and Cys551 residues are highly conserved, but the effect of disulfide bonds on the biochemical characteristics of UBA has not been elucidated. In addition, due to the presence of isolated Cys572, inactive inclusion bodies may be formed during protein expression or trigger protein aggregation during protein purification. In this study, the co-expression of SUMO fusion protein combined with SUMO protease (ULP enzyme) in Escherichia coli was successfully applied to improve the soluble expression of UBA domain. Introduced three mutants (UBAC551A, UBAC572A and UBAC551,572A) determined the effects of disulfide bonds on the biochemical characteristics of UBA. Circular dichroism and analytical size exclusion chromatography results showed that the target proteins obtained by co-expression could be folded correctly and had biological activity. Both thermal-induced and urea-induced results demonstrated that the elimination of disulfide bonds would significantly reduce the stability of UBA. Fluorescence spectroscopy result showed that the elimination of disulfide bonds slightly increases the binding affinity of UBA to ligands. In summary, soluble, stable and active UBA domain and its mutants were prepared by co-expression system, which will further contribute to the structural and functional research of UBA.

Biochemical and functional characterization of the N-terminal ubiquitin-like domain of human SHARPIN.

SHARPIN, an accessory subunit of the E3 ligase complex LUBAC, participates in the formation of LUBAC through the ubiquitin-like (UBL) domain located in the central region of SHARPIN and interacts with the ubiquitin-associated domain (UBA) of the catalytic subunit HOIP. However, the role of the N-terminal UBL domain of SHARPIN in stable LUBAC formation has not been clarified. In this study, the 1-127 domain, 128-309 domain, and UBL domain of SHARPIN expression vectors were constructed using the molecular biology method. Then the co-expression of SUMO fusion protein combined with SUMO protease (ULP enzyme) in Escherichia coli was successfully applied to improve the soluble expression of target protein. The results of circular dichroism proved that they all belong to the α+ß class of proteins. The results of size exclusion chromatography showed that 128-309 domain could combine with HOIP and HOIL-1L to participate in the stability of LUBAC. Both thermal-induced and urea-induced unfolding experiment results demonstrated that the existence of the N-terminal UBL domain could make the overall structure more stable than the alone UBL domain. Biosensor experiments indicated that the existence of the N-terminal UBL domain strengthened the binding ability of the UBL domain and the UBA domain. These results were conducive to further study the structure and function of SHARPIN.

The dual structural roles of the membrane distal region of the α-integrin cytoplasmic tail during integrin inside-out activation.

Studies on the mechanism of integrin inside-out activation have been focused on the role of ß-integrin cytoplasmic tails, which are relatively conserved and bear binding sites for the intracellular activators including talin and kindlin. Cytoplasmic tails for α-integrins share a conserved GFFKR motif at the membrane-proximal region and this forms a specific interface with the ß-integrin membrane-proximal region to keep the integrin inactive. The α-integrin membrane-distal regions, after the GFFKR motif, are diverse both in length and sequence and their roles in integrin activation have not been well-defined. In this study, we report that the α-integrin cytoplasmic membrane-distal region contributes to maintaining integrin in the resting state and to integrin inside-out activation. Complete deletion of the α-integrin membrane-distal region diminished talin- and kindlin-mediated integrin ligand binding and conformational change. A proper length and suitable amino acids in α-integrin membrane-distal region was found to be important for integrin inside-out activation. Our data establish an essential role for the α-integrin cytoplasmic membrane-distal region in integrin activation and provide new insights into how talin and kindlin induce the high-affinity integrin conformation that is required for fully functional integrins.

Modulation of integrin activation and signaling by α1/α1'-helix unbending at the junction.

How conformational signals initiated from one end of the integrin are transmitted to the other end remains elusive. At the ligand-binding ßI domain, the α1/α1'-helix changes from a bent to a straightened α-helical conformation upon integrin headpiece opening. We demonstrated that a conserved glycine at the α1/α1' junction is crucial for maintaining the bent conformation of the α1/α1'-helix in the resting state. Mutations that facilitate α1/α1'-helix unbending rendered integrin constitutively active; however, mutations that block the α1/α1'-helix unbending abolished soluble ligand binding upon either outside or inside stimuli. Such mutations also blocked ligand-induced integrin extension from outside the cell, but had no effect on talin-induced integrin extension from inside the cell. In addition, integrin-mediated cell spreading, F-actin stress fiber and focal adhesion formation, and focal adhesion kinase activation were also defective in these mutant integrins, although the cells still adhered to immobilized ligands at a reduced level. Our data establish the structural role of the α1/α1' junction that allows relaxation of the α1/α1'-helix in the resting state and transmission of bidirectional conformational signals by helix unbending upon integrin activation.

Cordyceps sinensis protects against liver and heart injuries in a rat model of chronic kidney disease: a metabolomic analysis.

AIM: To test the hypothesis that the traditional Chinese medicine Cordyceps sinensis could improve the metabolic function of extrarenal organs to achieve its anti-chronic kidney disease (CKD) effects. METHODS: Male SD rats were divided into CKD rats (with 5/6-nephrectomy), CKD rats treated with Cordyceps sinensis (4 mg•kg-1•d-1, po), and sham-operated rats. After an 8-week treatment, metabolites were extracted from the hearts and livers of the rats, and then subjected to (1)H-NMR-based metabolomic analysis. RESULTS: Oxidative stress, energy metabolism, amino acid and protein metabolism and choline metabolism were considered as links between CKD and extrarenal organ dysfunction. Within the experimental period of 8 weeks, the metabolic disorders in the liver were more pronounced than in the heart, suggesting that CKD-related extrarenal organ dysfunctions occurred sequentially rather than simultaneously. Oral administration of Cordyceps sinensis exerted statistically significant rescue effects on the liver and heart by reversely regulating levels of those metabolites that are typically perturbed in CKD. CONCLUSION: Oral administration of Cordyceps sinensis significantly attenuates the liver and heart injuries in CKD rats. The (1)H NMR-based metabolomic approach has provided a systematic view for understanding of CKD and the drug treatment, which can also be used to elucidate the mechanisms of action of other traditional Chinese medicines.

Achieving long-term survival in extensive-stage SCLC: a case report and mini literature review.

Managing extensive-stage SCLC (ES-SCLC) has long been challenging for clinicians and oncologists due to its aggressive nature and poor prognosis. We report a case of a 41-year-old female with ES-SCLC who survived for six years, defying the disease's typically poor prognosis. Through a heavy treatment strategy involving chemotherapy, targeted therapy, and immunotherapy, the patient experienced robust responses and avoided distant metastasis, including brain involvement. The long-term survival case in SCLC highlights the need for further research into personalized strategies and prognostic biomarkers. This case holds significant value for both clinicians and researchers as it challenges the conventional strategies for ES-SCLC and sets the stage for future evidence-based studies aimed at extending survival in SCLC.

Exercise motivation, physical exercise, and mental health among college students: examining the predictive power of five different types of exercise motivation.

Introduction: The mental health (MH) of college students has long been a crucial concern for families, educational institutions, and society. Extensive research has demonstrated the influential role of exercise motivation in shaping MH. However, further investigation is warranted to ascertain which types of exercise motivation may have more influence on the MH of college students. The present study examined the direct effects of five distinct types of exercise motivation, namely health motivation (HM), appearance motivation (APM), fun motivation (FM), ability motivation (ABM), and social motivation (SM) on MH. Additionally, the study explored the potential mediating role of physical exercise (PE) in these relationships. Methods: An cross-sectional study design was employed. A total of 433 Chinese college students participated in the study and completed our questionnaires, which included the Exercise motivation scale (EM scale), the Physical exercise scale (PE scale), and the Mental health scale (MH scale). Results: The findings revealed a significant and positive relationship between all five categories of exercise motivation and the MH of college students. Specifically, FM was found to have the most pronounced impact on MH, followed by HM, ABM, SM, and APM, in descending order of influence. Furthermore, the impacts of HM, FM, ABM, and SM on MH were found to be partially mediated by PE. However, the association between APM and MH was entirely mediated by PE. Discussion: The present study contributes to enhancing the comprehension of the underlying mechanisms behind different exercise motivations in relation to PE and MH. Additionally, it offers practical implications for developing intervention strategies for improving the MH of college students.

Efficacy and safety of a modified intravenous recombinant tissue plasminogen activator regimen in Chinese patients with acute ischemic stroke.

BACKGROUND: Thrombolytic treatment with intravenous (IV) recombinant tissue plasminogen activator (rtPA; 0.90 mg/kg, with a maximum dose of 90 mg) has been recommended as the standard management for acute ischemic stroke (AIS) thrombolysis. However, the dose of IV rtPA in Asia remains controversial. METHODS: This study was designed to verify the safety and efficacy of IV rtPA treatment for AIS with a lower dosage (0.90 mg/kg, with a maximum dose of 50 mg). Patients were divided into 3 dosage groups according to body weight (BW): group 1, <55 kg for 0.90 mg/kg; group 2, 55 to 67 kg for 0.75 to 0.90 mg/kg; and group 3, >67 kg for <0.75 mg/kg. The following data were collected: patient demographics, vascular risk factors, neuroimaging results, time of rtPA administration, National Institutes of Health Stroke Scale score before treatment and at 24 hours, and a modified Rankin Scale (mRS) score at 3 months. RESULTS: Eighty-three AIS patients who were of Han Chinese descent were included in the study. The baseline characteristics of the 3 dosage groups were well matched. In group 1 (BW <55 kg for 0.90 mg/kg; n = 19), 57.1% had a favorable outcome at 3 months, compared with 61.2% of patients in group 2 (BW 55-67 kg for 0.75-0.90 mg/kg; n = 33) and 51.5% in group 3 (BW >67 kg for <0.75 mg/kg; n = 31; P = .362). There were no significantly statistical differences in the incidence of symptomatic intracerebral hemorrhage and mortality rate. CONCLUSIONS: This IV rtPA regimen (0.90 mg/kg, with a maximum dose of 50 mg) not only shows sufficient favorable outcome in clinical practice in Chinese patients with AIS but also good health economic savings. This regimen could be suitable for many developing countries.

The COVID-19 related stress and social network addiction among Chinese college students: A moderated mediation model.

Based on social cognitive theory and gender differences, this study verified a moderated mediation model to explore the relationship between the COVID-19 related stress (CRS) and social network addiction (SNA) and evaluate the mediating role of fear of missing out (FoMO) and the moderating role of gender. A questionnaire survey was conducted, including 702 Chinese university students.This study used PROCESS to test the hypothesis model.The results showed that the CRS significantly and positively affected the SNA of college students and FoMO played a complementary mediating role. Moreover, the analysis of the moderated mediation model showed that gender moderated the relationship between FoMO and SNA; the effect of FoMO was stronger on the SNA of male college students than that of females. The results not only enhanced our understanding of the internal influencing mechanism of the relationship between CRS and SNA but also considered gender differences. In addition, some suggestions were proposed.

The Correlation Between Histologic, Immunophenotypic, and Molecular Characteristics of Pulmonary Sarcomatoid Carcinoma Reveals That Sarcomatoid Change Is Potentially Derived From Epithelial Carcinoma Cells Undergoing Epithelial-Mesenchymal Transition.

Pulmonary sarcomatoid carcinoma (PSC) is characterized by biphasic tumors with epithelial and mesenchymal phenotype. Little is known about the correlation between histologic, immunophenotypic features and the genetic profile of PSC. We analyzed the expression of epithelial-mesenchymal transition-related markers, adenocarcinoma (ADC) and squamous cell carcinoma lineage-specific markers of 205 PSC cases. The alteration of 5 targeted genes was detected by amplification-refractory mutation system-polymerase chain reaction. The intensity of cytokeratin staining was stronger in epithelial carcinoma (EC) than that of the sarcomatoid component (SC) of pleomorphic carcinoma, while vimentin was positive in only 16.3% (17/104) of EC of pleomorphic carcinoma. There is no significant difference between thyroid transcription factor 1 (TTF-1) expression in the SC (46.5%, 33/71) of pleomorphic carcinoma with ADC components and pure PSC (44.2%, 42/95) without p40 expression ( P =0.858). Four cases with ALK rearrangement were confirmed to co-express ALK fusion protein in both the SC and EC. The incidence of EGFR/ALK/KRAS mutation was similar between pleomorphic carcinoma with ADC components (40.6%, 26/64) and TTF-1 + pure PSC (38.2%, 13/34) ( P =0.583). However, higher proportions of TTF-1 + /p40 - PSC patients (44.8%, 39/87) had EGFR/ALK/KRAS mutation than those with TTF-1 - /p40 - PSC (16.7%, 4/24) ( P =0.031). The incidence of EGFR mutation was significantly higher in TTF-1-positive (18.4%, 16/87) than TTF-1-negative (2.7%, 2/74) PSC ( P =0.002). No EGFR and ALK abnormality were observed in 24 pleomorphic carcinoma cases with squamous cell carcinoma components or pure PSC with p40 expression. Our study reveals a close correlation between SC and EC components of pleomorphic carcinoma in terms of immunophenotypic and genetic features, which suggests that pleomorphic carcinoma is potentially derived from the sarcomatoid change of EC cells undergoing epithelial-mesenchymal transition.

Backstepping control for a two-link manipulator with appointed-time convergence.

In this paper, a novel backstepping control law is investigated, which guarantees appointed-time convergence for a two-link manipulator. In contrast to other fixed-time controllers or predefined-time controllers, the practical convergence time can be precisely obtained instead of being estimated. By utilizing a novel appointed-time prescribed performance function, the trajectory tracking error of the manipulator can be previously constrained. Furthermore, the external disturbances have been suppressed by introducing a disturbance observer with the convergence time as a prior control parameter and a switching sliding mode control law. The stability of the controller is analyzed by the Lyapunov method. Various numerical simulation results are carried out to demonstrate the efficacy of the proposed control scheme.

Conformational and biochemical characterization of a rat epididymis-specific lipocalin 12 expressed in Escherichia coli.

Lipocalin 12 (Lcn12) is a recently identified epididymis-specific protein that might play a significant physiological role in male reproduction. However, the detailed structure and function of Lcn12 remain to be determined. In the present work, we cloned, expressed, and purified the rat Lcn12 (rLcn12) protein in Escherichia coli, introduced the Cys176Ala substitution to eliminate the aggregation problem associated with the wild-type protein. Homology modeling results demonstrated that rLcn12 adopted an eight-stranded, antiparallel ß-barrel conformation containing a conserved disulfide bond between Cys98 and Cys203, which was in accordance with the physicochemical properties elucidated by a combination of mass, circular dichroism, and nuclear magnetic resonance spectrometry. The purified rLcn12 protein exhibited a high binding affinity for all-trans retinoic acid in fluorescence titration experiments, implying that rLcn12 could be involved in retinoic acid transport in the epididymis.

Soluble expression and characterization of a mouse epididymis-specific protein lipocalin6.

Mouse lipocalin6 (mLcn6) was recently identified to be specifically expressed in the epididymis and speculated to may play a role in sperm maturation. However, further studies were hindered due to the bottleneck to obtain enough recombinant mLcn6 proteins. In this article, GB1 tag was successfully applied to improve the soluble expression of mLcn6. Thermal unfolding experiments demonstrate that GB1 can enhance the structural stability of mLcn6. Fluorescence spectroscopy experiments show that mLcn6 prepared according to our procedure has high affinities to both retinoic acid (K(d)=810nM) and retinol (K(d)=210nM). In conclusion, soluble, stable and active mLcn6 was recombinantly prepared with the help of the GB1 tag, which will facilitate the structural and functional studies of mLcn6.

Genotypic variation in element concentrations in brown rice from Yunnan landraces in China.

The mineral elements present in brown rice play an important physiological role in global human health. We investigated genotypic variation of eight of these elements (P, K, Ca, Mg, Fe, Zn, Cu, and Mn) in 11 different grades of brown rice on the basis of the number and distance coefficients of 282 alleles for 20 simple sequence repeat (SSR) markers. Six-hundred and twenty-eight landraces from the same field in Yunnan Province, one of the largest centers of genetic diversity of rice (Oryza sativa L.) in the world, formed our core collection. The mean concentrations (mg kg(-1)) of the eight elements in brown rice for these landraces were P (3,480) > K (2,540) > Mg (1,480) > Ca (157) > Zn (32.8) > Fe (32.0) > Cu (13.6) > Mn (13.2). Mean P concentrations in brown rice were 6.56 times total soil P, so the grains are important in tissue storage of P, but total soil K is 7.82 times mean K concentrations in brown rice. The concentrations of the eight elements in some grades of brown rice, on the basis of the number and distance coefficients of alleles for 20 SSR markers for the landraces, were significantly different (P < 0.05), and further understanding of the relationship between mineral elements and gene diversity is needed. There was large variation in element concentrations in brown rice, ranging from 2,160 to 5,500 mg P kg(-1), from 1,130 to 3,830 mg K kg(-1), from 61.8 to 488 mg Ca kg(-1), from 864 to 2,020 mg Mg kg(-1), from 0.40 to 147 mg Fe kg(-1), from 15.1 to 124 mg Zn kg(-1), from 0.10 to 59.1 mg Cu kg(-1), and from 6.7 to 26.6 mg Mn kg(-1). Therefore, germplasm evaluations for Ca, Fe, and Zn concentrations in rice grains have detected up to sevenfold genotypic differences, suggesting that selection for high levels of Ca, Fe, and Zn in breeding for mass production is a feasible approach. Increasing the concentrations of Ca, Fe, and Zn in rice grains will help alleviate chronic Ca, Zn, and Fe deficiencies in many areas of the world.

Elemental content in brown rice by inductively coupled plasma atomic emission spectroscopy reveals the evolution of asian cultivated rice.

The phylogenetic relationship for classification traits and eight mineral elements in brown rice (Oryza sativa L.) from Yunnan Province in China was carried out using microwave assisted digestion followed by inductively coupled plasma atomic emission spectroscopy, and the analytical procedures were carefully controlled and validated. In general, the results show that the mean levels of K, Ca, Mg, Fe and Cu in brown rice for 789 accessions of rice landraces was distinctly lower than that of improved cultivars. They further demonstrate that Ca plays an important role in the differentiation of subspecies indica-japonica, especially to enhance adaptation of cold stress, and that five mineral elements in brown rice enhance the eurytopicity from landrace to improved cultivar. Hierarchical cluster analysis, using average linkage from SPSS software based on eight mineral elements in brown rice, showed that Yunnan rice could be grouped into rice landrace and improved cultivar, with the rice landrace being further clustered into five subgroups, and that, interestingly, purple rice does not cluster with either of the groups. Our present data confirm that indica is the closest relative of late rice and white rice, and that they constitute rice landraces together, whereas japonica is the closest relatives of non-nuda, early-mid and glutinous rice. It is further shown that japonica, non-nuda, early-mid, glutinous, white and red rice might be more primitive than indica, nuda, late, non-glutinous and purple rice, respectively.