A nomogram predicting distant metastasis risk for gastric cancer patients with preoperative anemia: a multicenter retrospective study.

BACKGROUND: Anemia represents a well-established risk factor for patients diagnosed with gastric cancer, and is often associated with an unfavorable prognosis. In this context, the timely prediction of distant metastasis risk in patients with anemic gastric cancer assumes paramount importance. METHODS: Information of gastric cancer patients complicated with preoperative anemia in the First Affiliated Hospital of Sun Yat-sen University was collected. The cohort from the First Affiliated Hospital of Guangxi Medical University was used as an external validation set. A Nomogram was established based on the risk factors screened by univariate and multivariate logistic regression analyses. RESULTS: A total of 848 gastric cancer patients with preoperative anemia were enrolled. Pyloric obstruction, carcinoma antigen 125, T stage, N stage, tumor size, and preoperative weight loss were independent predictors of distant metastasis in gastric cancer patients with anemia (p < 0.05), based on which a nomogram was constructed. The accuracy, reliability and clinical value of the nomogram were evaluated by concordance index, receiver operating characteristic curve, decision curve analysis, calibration curve and showed good stability and clinical predictive value. CONCLUSIONS: Preoperative anemic gastric cancer patients, complicated with pyloric obstruction, elevated CA125, advanced T and N stage, larger tumor size, and preoperative weight loss, should be paid more attention to distant metastasis.

The neuropathological mechanism of EV-A71 infection attributes to inflammatory pryoptosis and viral replication via activating the hsa_circ_0045431/ hsa_miR_584/NLRP3 regulatory axis.

Neuropathological damage has been considered to be the main cause of death from EV-A71 infection, but the underlying mechanism has not been elucidated. Pyroptosis, a new form of inflammatory programmed cell death, has been verified to be involved in the pathogenesis of various viruses. circRNAs are a novel type of endogenous noncoding RNA gaining research interest in recent years, especially their special roles in the process of virus infection. Thus, in this study, we combined EV-A71, pyroptosis and circRNA to find a breakthrough in the pathogenesis of EV-A71 infection. Firstly, whether EV-A71 infection leaded to pyroptosis formation was examined by a series detection of cell death, cell viability, LDH release, caspase 1 activity, the expression levels of pyroptosis-related molecules and the concentrations of IL-1ß and IL-18. Secondly, high-throughput sequencing of circRNAs was carried out to excavate the circRNA-miRNA-mRNA regulatory axis which might be associated with pyroptosis formation. Finally, the gain- and loss-of-functional experiments were further conducted to identify their functions. Our results showed that EV-A71 infection caused pyroptosis formation in SH-SY5Y cells. The circRNA sequencing analyzed the differentially expressed circRNAs and their possible functions. It was found that the hsa_circ_0045431/hsa_miR_584/NLRP3 regulatory axis might be involved in pyroptosis formation during EV-A71 infection. Then, hsa_circ_0045431 sponged hsa_miR_584 and hsa_miR_584 directly targeted NLRP3 were validated by IF, dual-luciferase, qRT-PCR and WB assays. Functional experiments were performed to further uncover that the up-regulation of hsa_circ_0045431 and NLRP3 promoted the inflammatory pyroptosis and viral replication, while the up-regulation of hsa_miR_584 suppressed the inflammatory pyroptosis and viral replication, and vice versa. Collectively, our study demystified that EV-A71 infection induced pyroptosis formation by activating hsa_circ_0045431/hsa_miR_584/NLRP3 regulatory axis, which could further effect viral replication. These findings provided novel insights into the pathogenesis of EV-A71 infection, and meanwhile revealed that the hsa_circ_0045431/ hsa_miR_584/NLRP3 regulatory axis can serve as a potential biological therapeutic target for EV-A71 infection.

Value of the combination of a smartphone-compatible infrared camera and a hand-held doppler ultrasound in preoperative localization of perforators in flaps.

This study was conducted to evaluate the effectiveness of the FLIR ONE PRO, a thermal imaging camera for smartphones, combined with handheld Doppler (HHD) in the localization of perforator arteries and to assess the efficacy of the FLIR ONE PRO in distinguishing perforators of the descending branch of the lateral circumflex femoral artery (LCFA) from other perforators of the anterolateral thigh perforator (ALTP) flap. We enrolled 29 free perforator flaps from 22 patients in our study. Before surgery, dynamic infrared thermography was performed using a FLIR ONE PRO to visualize hotspots on the flaps. Subsequently, HHD was used to further determine the perforators under the hotspots, which were ultimately identified and confirmed through intraoperative findings. Additionally, infrared images of the ALTP flap were analyzed using FLIR Tools. The performances of the FLIR ONE PRO and FLIR ONE PRO + HHD groups were evaluated by comparing the intraoperative findings. Using FLIR ONE PRO + HHD, 119 hotspots and 106 perforators were identified during surgery. Using FLIR ONE PRO + HHD, sensitivity and positive predictive value were 97.87% and 88.46%, respectively, in the young (age≤45 years). In the elderly group (age>45 years), these percentages were 93.22% and 82.09%, respectively. In addition, we found that the FLIR ONE PRO could be useful for differentiating perforators in the descending branch of the LCFA from other perforators within 5 min. The results showed a sensitivity of 96.15%, a specificity of 98.9%, a positive predictive value of 96.15%, and a negative predictive value of 98.9%. Compared to using FLIR ONE PRO alone, the combined application of HHD and FLIR ONE PRO had a higher value in perforator localization by increasing the positive predictive value. The FLIR ONE PRO may have significance in the rapid prediction of perforators deriving from the descending branch of the LCFA.

Automatic range of motion measurement via smartphone images for telemedicine examination of the hand.

BACKGROUND: Telemedicine support virtual consultations and evaluations in hand surgery for patients in remote areas during the COVID-19 era. However, traditional physical examination is challenging in telemedicine and it is inconvenient to manually measure the hand range of motion (ROM) from images or videos. Here, we propose an automatic method using the hand pose estimation technique, aiming to measure the hand ROM from smartphone images. METHODS: Twenty-eight healthy volunteers participated in the study. An eight-hand gestures measurement protocol and the Google MediaPipe Hands were used to analyze images and calculate the ROM automatically. Manual goniometry was also performed according to the guideline of the American Medical Association. The correlation between the automatic and manual methods was analyzed by the intraclass correlation coefficient and Pearson correlation coefficient. The clinical acceptance was testified using Bland-Altman plots. RESULTS: A total of 32 parameters of each hand were measured by both methods, and 1792 measurement results were compared. The mean difference between automatic and manual methods is -2.21 ± 9.29° in the angle measurement and 0.48 ± 0.48 cm in the distance measurement. The intraclass correlation coefficient of 75% of parameters was higher than 0.75, the Pearson correlation coefficient of 84% of parameters was over 0.6, and 40.6% of parameters reached well-accepted clinical agreements. CONCLUSIONS: The proposed method provides a helpful protocol for automatic hand ROM measurement based on smartphone images and the MediaPipe Hands pose estimation technique. The automatic measurement is acceptable and comparable with existing methods, showing a possible application in the telemedicine examination of hand surgery.

Validity and Reliability of a Depth Camera-Based Quantitative Measurement for Joint Motion of the Hand.

Purpose: Quantitative measurement of hand motion is essential in evaluating hand function. This study aimed to investigate the validity and reliability of a novel depth camera-based contactless automatic measurement system to assess hand range of motion and its potential benefits in clinical applications. Methods: Five hand gestures were designed to evaluate the hand range of motion using a depth camera-based measurement system. Seventy-one volunteers were enrolled in performing the designed hand gestures. Then, the hand range of motion was measured with the depth camera and manual procedures. System validity was evaluated based on 3 dimensions: repeatability, within-laboratory precision, and reproducibility. For system reliability, linear evaluation, the intraclass correlation coefficient, paired t -test and bias were employed to test the consistency and difference between the depth camera and manual procedures. Results: When measuring phalangeal length, repeatability, within-laboratory precision, and reproducibility were 2.63%, 12.87%, and 27.15%, respectively. When measuring angles of hand motion, the mean repeatability and within-laboratory precision were 1.2° and 3.3° for extension of 5 digits, 2.7° and 10.2° for flexion of 4 fingers, and 3.1° and 5.3° for abduction of 4 metacarpophalangeal joints, respectively. For system reliability, the results showed excellent consistency (intraclass correlation coefficient = 0.823; P < .05) and good linearity with the manual procedures (r = 0.909-0.982, approximately; P < .001). Besides, 78.3% of the measurements were clinically acceptable. Conclusions: Our depth camera-based evaluation system provides acceptable validity and reliability in measuring hand range of motion and offers potential benefits for clinical care and research in hand surgery. However, further studies are required before clinical application. Clinical relevance: This study suggests a depth camera-based contactless automatic measurement system holds promise for assessing hand range of motion in hand function evaluation, diagnosis, and rehabilitation for medical staff. However, it is currently not adequate for all clinical applications.

Precision and accuracy of measuring finger motion with a depth camera: a cross-sectional study of healthy participants.

The purpose of this cross-sectional study was to determine the precision and accuracy of the measurement of finger motion with a depth camera. Fifty-five healthy adult hands were included. Measurements were done with a depth camera and compared with traditional manual goniometer measurements. Repeated measuring showed that the overall repeatability and reproducibility of extension measured with the depth camera were within 3° and 4° and that of flexion were within 13° and 14°. Compared with traditional manual goniometry, biases of extension of all finger joints and flexion of metacarpophalangeal joints were less than 5°, and the average bias of flexion of proximal and distal interphalangeal joints was 29°. We conclude that the measurement of finger extension and flexion of the metacarpophalangeal joints with a depth camera was reliable, but improvement is required in the precision and accuracy of interphalangeal joint flexion.

[Accuracy of key point matrix technology based contactless automatic measurement for joint motion of hand].

Objective: To validate the use of key point matrix technology based contactless automatic measurement for evaluation of joint motion of hand. Methods: Thirty-three volunteers were enrolled to evaluate the extension and flexion of hand joints between May 2021 and November 2021. There were 20 males and 13 females, the age ranged from 16 to 70 years with an average of 30.2 years. The extension angles of 14 joints of 5 fingers (including hyperextension) and the flexion angles of 12 joints of 4 fingers (excluding thumb) of volunteers were measured by key point matrix technology and manual goniometer, respectively. Then 5 participants and repeated measurement experiment were employed to test the system repeatability and accuracy; 28 participants and paired measurement experiment were employed to test the system accuracy. Results: The average repeatability of finger joint motion measured by the key point matrix technology was 1.801° (extension) and 7.823° (flexion), respectively. Compared with manual measurement, the average differences of each finger joint measured by the key point matrix technology were 3.225° in extension and 14.145° in flexion, respectively. The key point matrix technology based contactless automatic evaluation system offered excellent consistency with the manual goniometers ( ICC=0.875). While most of the consistency with manual goniometer of individual joints were at moderate levels (median of ICC, 0.440). The correlation coefficients between the measurement results of the two methods were mainly positive in the extension of the joint ( P<0.05) and negative in the flexion of the joints ( P<0.05). Conclusion: The key point matrix technology based contactless automatic evaluation provides sufficient measurement repeatability and accuracy in evaluation for the joint motion of hand.

Gene expression profiles for an immunoscore model in bone and soft tissue sarcoma.

BACKGROUND: Immune infiltration is a prognostic marker to clinical outcomes in various solid tumors. However, reports that focus on bone and soft tissue sarcoma are rare. The study aimed to analyze and identify how immune components influence prognosis and develop a novel prognostic system for sarcomas. METHODS: We retrieved the gene expression data from 3 online databases (GEO, TCGA, and TARGET). The immune fraction was estimated using the CIBERSORT algorithm. After that, we re-clustered samples by K-means and constructed immunoscore by the least absolute shrinkage and selection operator (LASSO) Cox regression model. Next, to confirm the prognostic value, nomograms were constructed. RESULTS: 334 samples diagnosed with 8 tumor types (including osteosarcoma) were involved in our analysis. Patients were next re-clustered into three subgroups (OS, SAR1, and SAR2) through immune composition. Survival analysis showed a significant difference between the two soft tissue groups: patients with a higher proportion of CD8+ T cells, macrophages M1, and mast cells had favorable outcomes (p=0.0018). Immunoscore models were successfully established in OS and SAR2 groups consisting of 12 and 9 cell fractions, respectively. We found immunosocre was an independent factor for overall survival time. Patients with higher immunoscore had poor prognosis (p<0.0001). Patients with metastatic lesions scored higher than those counterparts with localized tumors (p<0.05). CONCLUSIONS: Immune fractions could be a useful tool for the classification and prognosis of bone and soft tissue sarcoma patients. This proposed immunoscore showed a promising impact on survival prediction.

DNA methylation patterns-based subtype distinction and identification of soft tissue sarcoma prognosis.

ABSTRACT: Soft tissue sarcomas (STSs) are heterogeneous at the clinical with a variable tendency of aggressive behavior. In this study, we constructed a specific DNA methylation-based classification to identify the distinct prognosis-subtypes of STSs based on the DNA methylation spectrum from the TCGA database. Eventually, samples were clustered into 4 subgroups, and their survival curves were distinct from each other. Meanwhile, the samples in each subgroup reflected differentially in several clinical features. Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis was also conducted on the genes of the corresponding promoter regions of the above-described specific methylation sites, revealing that these genes were mainly concentrated in certain cancer-associated biological functions and pathways. In addition, we calculated the differences among clustered methylation sites and performed the specific methylation sites with LASSO algorithm. The selection operator algorithm was employed to derive a risk signature model, and a prognostic signature based on these methylation sites performed well for risk stratification in STSs patients. At last, a nomogram consisted of clinical features and risk score was developed for the survival prediction. This study declares that DNA methylation-based STSs subtype classification is highly relevant for future development of personalized therapy as it identifies the prediction value of patient prognosis.

Novel therapeutic compounds for prostate adenocarcinoma treatment: An analysis using bioinformatic approaches and the CMap database.

INTRODUCTION: Prostate adenocarcinoma is the most frequently diagnosed malignancy, particularly for people >70 years old. The main challenge in the treatment of advanced neoplasm is bone metastasis and therapeutic resistance for known oncology drugs. Novel treatment methods to prolong the survival time and improve the life quality of these specific patients are required. The present study attempted to screen potential therapeutic compounds for the tumor through bioinformatics approaches, in order to provide conceptual treatment for this malignant disease. METHODS: Differentially expressed genes were obtained from the Gene Expression Omnibus database and submitted into the Connectivity Map database for the detection of potentially associated compounds. Target genes were extracted from the search results. Functional annotation and pathway enrichment were performed for the confirmation. Survival analysis was used to measure potential therapeutic effects. RESULTS: It was revealed that 3 compounds (vanoxerine, tolnaftate, and gabexate) may help to prolong the disease-free survival time from tumor metastasis of patients with the tumor. A total of 6 genes [also-keto reductase family 1 member C3 (AKR1C3), collagen type III α 1 chain (COL3A1), lipoprotein lipase (LPL), glucuronidase, ß pseudogene 11 (GUSBP11), apolipoprotein E (APOE), and collagen type I α 1 chain (COL1A1)] were identified to be the potential therapeutic targets for the aforementioned compounds. CONCLUSION: In the present study, it was speculated that 3 compounds may function as the potential therapeutic drugs of bone metastatic prostate adenocarcinoma; however, further studies verifying vitro and in vivo are necessary.

Transcriptome sequencing analysis of SH-SY5Y cells infected with EV71 reveals the potential neuropathic mechanisms.

Enterovirus A71 (EV71) remains the most common causative agent of hand, foot, and mouth disease (HFMD), and the neurological complications induced by EV71 are usually the leading cause of death in children with HFMD. However, the mechanism of nervous system changes caused by EV71 infection is still unclear. Therefore, in the current study, EV71 was inoculated into the human neuroblastoma cell line SH-SY5Y and subsequent transcriptome sequencing was used to examine the alterations of the transcriptome in infected SH-SY5Y cells. It is expected to determine the underlying mechanism of neurological diseases in response to EV71 infection. As a result, a total of 82,406,974, 112,410,808 and 87,780,371 clean reads were found in the control, EV71-12 h and EV71-24 h groups, respectively. Moreover, 160 and 745 differentially expressed genes were identified in the EV71-12 h and EV71-24 h groups, respectively, as compared to the control group. Next, to further explore the pathogenic mechanism triggered by EV71 infection, we mainly focused on the common differentially expressed genes at different time points of EV71 infection. And it was discovered that there were 95 common differentially expressed genes, which were used to conduct GO and pathway analysis. GO enrichment analysis demarcated related biological processes, molecular functions and cellular components, and KEGG pathway analysis enabled annotations of metabolic pathways and revealed interactions among the significantly enriched pathways. The results showed that the enriched GO term "Nervous system development" and enriched pathway "CCKR signaling map" might be important contributors to EV71-induced neuropathological mechanisms. In addition, we also screened 10 up- and down-regulated non-protein coding genes with significantly different expression in our transcriptome profiling, which suggested that these abnormally regulated non-protein-encoding genes might also play important roles in the pathogenesis of EV71 infection. Eventually, RT-qPCR technology was adopted to validate the transcriptome sequencing data and the experiment demonstrated that the RT-qPCR and transcriptome sequencing results were basically consistent. In summary, this is the first transcriptome analysis of SH-SY5Y cells in response to EV71 infection and provides valuable cues for further exploring the mechanism of nervous system changes caused by EV71 infection.

Extremely rare case of intravascular solitary fibrous tumour in the inferior vena cava with review of the literature.

BACKGROUND: Solitary fibrous tumour (SFT) is a mesenchymal tumour of fibroblastic type, and it develops in almost any part of the human body. However, according to previous studies, the occurrence of intravascular SFTs is extremely rare. CASE PRESENTATION: We reported a case of intravascular SFT in a 67-year-old woman who has been experiencing swelling and pain in the right leg for 2 months. Computed tomography venography scan revealed a well-defined mass obstructing the inferior vena cava (IVC). Surgical resection was performed, and histopathologic and immunohistochemical results were consistent with SFT. Further, next-generation sequencing (NGS) analysis was performed, and results revealed two tumour-related gene mutations (deletion of PMS2 and variation of ESR1 [L536P]). The patient did not receive any adjuvant therapy, and no signs of tumour progression were observed during the 6-month follow-up. CONCLUSION: To the best of our knowledge, this study first presented about SFT arising from the IVC and carried out an NGS analysis to validate the molecular mechanism of such condition.

Periodontal manifestations of systemic diseases and developmental and acquired conditions: Consensus report of workgroup 3 of the 2017 World Workshop on the Classification of Periodontal and Peri-Implant Diseases and Conditions.

BACKGROUND: A variety of systemic diseases and conditions can affect the course of periodontitis or have a negative impact on the periodontal attachment apparatus. Gingival recessions are highly prevalent and often associated with hypersensitivity, the development of caries and non-carious cervical lesions on the exposed root surface and impaired esthetics. Occlusal forces can result in injury of teeth and periodontal attachment apparatus. Several developmental or acquired conditions associated with teeth or prostheses may predispose to diseases of the periodontium. The aim of this working group was to review and update the 1999 classification with regard to these diseases and conditions, and to develop case definitions and diagnostic considerations. METHODS: Discussions were informed by four reviews on 1) periodontal manifestions of systemic diseases and conditions; 2) mucogingival conditions around natural teeth; 3) traumatic occlusal forces and occlusal trauma; and 4) dental prostheses and tooth related factors. This consensus report is based on the results of these reviews and on expert opinion of the participants. RESULTS: Key findings included the following: 1) there are mainly rare systemic conditions (such as Papillon-Lefevre Syndrome, leucocyte adhesion deficiency, and others) with a major effect on the course of periodontitis and more common conditions (such as diabetes mellitus) with variable effects, as well as conditions affecting the periodontal apparatus independently of dental plaque biofilm-induced inflammation (such as neoplastic diseases); 2) diabetes-associated periodontitis should not be regarded as a distinct diagnosis, but diabetes should be recognized as an important modifying factor and included in a clinical diagnosis of periodontitis as a descriptor; 3) likewise, tobacco smoking - now considered a dependence to nicotine and a chronic relapsing medical disorder with major adverse effects on the periodontal supporting tissues - is an important modifier to be included in a clinical diagnosis of periodontitis as a descriptor; 4) the importance of the gingival phenotype, encompassing gingival thickness and width in the context of mucogingival conditions, is recognized and a novel classification for gingival recessions is introduced; 5) there is no evidence that traumatic occlusal forces lead to periodontal attachment loss, non-carious cervical lesions, or gingival recessions; 6) traumatic occlusal forces lead to adaptive mobility in teeth with normal support, whereas they lead to progressive mobility in teeth with reduced support, usually requiring splinting; 7) the term biologic width is replaced by supracrestal tissue attachment consisting of junctional epithelium and supracrestal connective tissue; 8) infringement of restorative margins within the supracrestal connective tissue attachment is associated with inflammation and/or loss of periodontal supporting tissue. However, it is not evident whether the negative effects on the periodontium are caused by dental plaque biofilm, trauma, toxicity of dental materials or a combination of these factors; 9) tooth anatomical factors are related to dental plaque biofilm-induced gingival inflammation and loss of periodontal supporting tissues. CONCLUSION: An updated classification of the periodontal manifestations and conditions affecting the course of periodontitis and the periodontal attachment apparatus, as well as of developmental and acquired conditions, is introduced. Case definitions and diagnostic considerations are also presented.

Periodontal manifestations of systemic diseases and developmental and acquired conditions: Consensus report of workgroup 3 of the 2017 World Workshop on the Classification of Periodontal and Peri-Implant Diseases and Conditions.

BACKGROUND: A variety of systemic diseases and conditions can affect the course of periodontitis or have a negative impact on the periodontal attachment apparatus. Gingival recessions are highly prevalent and often associated with hypersensitivity, the development of caries and non-carious cervical lesions on the exposed root surface and impaired esthetics. Occlusal forces can result in injury of teeth and periodontal attachment apparatus. Several developmental or acquired conditions associated with teeth or prostheses may predispose to diseases of the periodontium. The aim of this working group was to review and update the 1999 classification with regard to these diseases and conditions, and to develop case definitions and diagnostic considerations. METHODS: Discussions were informed by four reviews on 1) periodontal manifestions of systemic diseases and conditions; 2) mucogingival conditions around natural teeth; 3) traumatic occlusal forces and occlusal trauma; and 4) dental prostheses and tooth related factors. This consensus report is based on the results of these reviews and on expert opinion of the participants. RESULTS: Key findings included the following: 1) there are mainly rare systemic conditions (such as Papillon-Lefevre Syndrome, leucocyte adhesion deficiency, and others) with a major effect on the course of periodontitis and more common conditions (such as diabetes mellitus) with variable effects, as well as conditions affecting the periodontal apparatus independently of dental plaque biofilm-induced inflammation (such as neoplastic diseases); 2) diabetes-associated periodontitis should not be regarded as a distinct diagnosis, but diabetes should be recognized as an important modifying factor and included in a clinical diagnosis of periodontitis as a descriptor; 3) likewise, tobacco smoking - now considered a dependence to nicotine and a chronic relapsing medical disorder with major adverse effects on the periodontal supporting tissues - is an important modifier to be included in a clinical diagnosis of periodontitis as a descriptor; 4) the importance of the gingival phenotype, encompassing gingival thickness and width in the context of mucogingival conditions, is recognized and a novel classification for gingival recessions is introduced; 5) there is no evidence that traumatic occlusal forces lead to periodontal attachment loss, non-carious cervical lesions, or gingival recessions; 6) traumatic occlusal forces lead to adaptive mobility in teeth with normal support, whereas they lead to progressive mobility in teeth with reduced support, usually requiring splinting; 7) the term biologic width is replaced by supracrestal tissue attachment consisting of junctional epithelium and supracrestal connective tissue; 8) infringement of restorative margins within the supracrestal connective tissue attachment is associated with inflammation and/or loss of periodontal supporting tissue. However, it is not evident whether the negative effects on the periodontium are caused by dental plaque biofilm, trauma, toxicity of dental materials or a combination of these factors; 9) tooth anatomical factors are related to dental plaque biofilm-induced gingival inflammation and loss of periodontal supporting tissues. CONCLUSION: An updated classification of the periodontal manifestations and conditions affecting the course of periodontitis and the periodontal attachment apparatus, as well as of developmental and acquired conditions, is introduced. Case definitions and diagnostic considerations are also presented.

Ecto-5'-nucleotidase: a candidate virulence factor in Streptococcus sanguinis experimental endocarditis.

Streptococcus sanguinis is the most common cause of infective endocarditis (IE). Since the molecular basis of virulence of this oral commensal bacterium remains unclear, we searched the genome of S. sanguinis for previously unidentified virulence factors. We identified a cell surface ecto-5'-nucleotidase (Nt5e), as a candidate virulence factor. By colorimetric phosphate assay, we showed that S. sanguinis Nt5e can hydrolyze extracellular adenosine triphosphate to generate adenosine. Moreover, a nt5e deletion mutant showed significantly shorter lag time (P<0.05) to onset of platelet aggregation than the wild-type strain, without affecting platelet-bacterial adhesion in vitro (P=0.98). In the absence of nt5e, S. sanguinis caused IE (4 d) in a rabbit model with significantly decreased mass of vegetations (P<0.01) and recovered bacterial loads (log(10)CFU, P=0.01), suggesting that Nt5e contributes to the virulence of S. sanguinis in vivo. As a virulence factor, Nt5e may function by (i) hydrolyzing ATP, a pro-inflammatory molecule, and generating adenosine, an immunosuppressive molecule to inhibit phagocytic monocytes/macrophages associated with valvular vegetations. (ii) Nt5e-mediated inhibition of platelet aggregation could also delay presentation of platelet microbicidal proteins to infecting bacteria on heart valves. Both plausible Nt5e-dependent mechanisms would promote survival of infecting S. sanguinis. In conclusion, we now show for the first time that streptococcal Nt5e modulates S. sanguinis-induced platelet aggregation and may contribute to the virulence of streptococci in experimental IE.

The two-component system BfrAB regulates expression of ABC transporters in Streptococcus gordonii and Streptococcus sanguinis.

The putative two-component system BfrAB is involved in Streptococcus gordonii biofilm development. Here, we provide evidence that BfrAB regulates the expression of bfrCD and bfrEFG, which encode two ATP-binding cassette (ABC) transporters, and bfrH, which encodes a CAAX amino-terminal protease family protein. BfrC and BfrE are ATP-binding proteins, and BfrD, BfrF and BfrG are homologous membrane-spanning polypeptides. Similarly, BfrABss, the BfrAB homologous system in Streptococcus sanguinis, controls the expression of two bfrCD-homologous operons (bfrCDss and bfrXYss), a bfrH-homologous gene (bfrH1ss) and another CAAX amino-terminal protease family protein gene (bfrH2ss). Furthermore, we demonstrate that the purified BfrA DNA-binding domain from S. gordonii binds to the promoter regions of bfrCD, bfrEFG, bfrH, bfrCDss, bfrXYss and bfrH1ss in vitro. Finally, we show that the BfrA DNA-binding domain recognizes a conserved DNA motif with a consensus sequence of TTTCTTTAGAAATATTTTAGAATT. These data suggest, therefore, that S. gordonii BfrAB controls biofilm formation by regulating multiple ABC-transporter systems.