Evaluations of artificial intelligence and machine learning algorithms in neurodiagnostics.

This article evaluates the ethical implications of utilizing artificial intelligence (AI) algorithms in neurological diagnostic examinations. Applications of AI technology have been utilized to aid in the determination of pharmacological dosages of gadolinium for brain lesion detection, localization of seizure foci, and the characterization of large vessel occlusion in ischemic stroke patients. Multiple subtypes of AI/machine learning (ML) algorithms are analyzed, as AI-assisted neurology utilizes supervised, unsupervised, artificial neural network (ANN), and deep neural network (DNN) learning models. As ANN and DNN analyses can be applied to data with an unknown clinical diagnosis, these algorithms are evaluated according to Bayesian statistical analyses. Bayesian neural network analyses are incorporated, as these algorithms indicate that the predictive accuracy and model performance are dependent upon accurate configurations of the model's hyperparameters and neural inputs. Thus, mathematical evaluations of AI algorithms are comprehensively explored to examine their clinical utility, as underperformance of AI/ML models may have deleterious consequences that affect patient outcomes due to misdiagnosis and false-negative test results.

Optimization of physicochemical properties of pyrrolidine GPR40 AgoPAMs results in a differentiated profile with improved pharmacokinetics and reduced off-target activities.

GPR40 AgoPAMs are highly effective antidiabetic agents that have a dual mechanism of action, stimulating both glucose-dependent insulin and GLP-1 secretion. The early lipophilic, aromatic pyrrolidine and dihydropyrazole GPR40 AgoPAMs from our laboratory were highly efficacious in lowering plasma glucose levels in rodents but possessed off-target activities and triggered rebound hyperglycemia in rats at high doses. A focus on increasing molecular complexity through saturation and chirality in combination with reducing polarity for the pyrrolidine AgoPAM chemotype resulted in the discovery of compound 46, which shows significantly reduced off-target activities as well as improved aqueous solubility, rapid absorption, and linear PK. In vivo, compound 46 significantly lowers plasma glucose levels in rats during an oral glucose challenge yet does not demonstrate the reactive hyperglycemia effect at high doses that was observed with earlier GPR40 AgoPAMs.

Muscle-invasive Urothelial Cancer: Association of Mutational Status with Metastatic Pattern and Survival.

Background Muscle-invasive urothelial cancer (MIUC) is characterized by substantial genetic heterogeneity and high mutational frequency. Correlation between frequently mutated genes with clinical behavior has been recently demonstrated. Nonetheless, correlation between mutational status of MIUC and metastatic pattern is unknown. Purpose To investigate the association of mutational status of MIUC with metastatic pattern, metastasis-free survival (MFS), and overall survival (OS). Materials and Methods This single-center retrospective study evaluated consecutive patients with biopsy-proven MIUC who underwent serial cross-sectional imaging (CT, MRI, or fluorine 18 fluorodeoxyglucose PET/CT) between April 2010 and December 2018. Mutational status was correlated with location of metastases using the χ2 or Fisher exact test. Mutational status and metastatic pattern were correlated with MFS and OS using univariable Cox proportional hazard models. High-risk (presence of TP53, RB1, or KDM6A mutation) and low-risk (presence of ARID1A, FGFR3, PIK3CA, STAG2, and/or TSC1 mutation and absence of TP53, RB1, or KDM6A mutation) groups were determined according to existing literature and were correlated with MFS and OS by using multivariable Cox proportional hazard models. Results One hundred three patients (mean age, 72 years ± 11 [standard deviation]; 81 men) were evaluated. Seventeen of 103 (16%) patients had metastatic disease at diagnosis; 38 of 103 (37%) developed metastatic disease at a median of 5.9 months (interquartile range, 0.8-28 months). TP53 mutation (seen in 58 of 103 patients, 56%) was associated with lymphadenopathy (relative risk [RR]: 1.7; 95% confidence interval [CI]: 1.2, 2.4; P = .002) and osseous metastases (RR: 1.9; 95% CI: 1.6, 2.3; P = .02); RB1 mutation (seen in 19 of 103 patients, 18.4%) was associated with peritoneal carcinomatosis (RR: 5.9; 95% CI: 3.8, 9.2; P = .03). ARID1A mutation was associated with greater OS (hazard ratio [HR]: 3.1; 95% CI: 1.2, 10; P = .01). At multivariable Cox analysis, the high-risk group (TP53, RB1, and/or KDM6A mutations) was independently associated with shorter MFS (HR: 3.5, 95% CI: 1.3, 12; P = .009) and shorter OS (HR: 3.1; 95% CI: 1.2, 10; P = .02). Conclusion Mutational status of muscle-invasive urothelial cancer has implications on metastatic pattern, metastasis-free survival, and overall survival. © RSNA, 2020 Online supplemental material is available for this article. See also the editorial by Choyke in this issue.

DNA methylation in breast cancers: Differences based on estrogen receptor status and recurrence.

DNA methylation plays a role in the etiology of primary breast cancers. We analyzed paired primary and second breast tumors to elucidate the role of methylation in recurrence. Methylation profiles from paired primary and second breast tumors of 23 women were assessed using the HumanMethylation450 BeadChip. Twelve women had estrogen receptor positive (ERpos) primary and second tumors, five had estrogen receptor negative (ERneg) primary and second tumors, and six had an ERpos primary tumor but an ERneg second tumor. Stratifying tumors by occurrence revealed that the greater methylation previously associated with ERpos tumors, is more pronounced in primary tumors than in second tumors. Further, ERneg second tumors are more methylated than ERpos second tumors among women who had ERpos primary tumors. Pathway analyses using gene lists generated from comparisons of methylation in ERpos primary tumors from the paired sets with ERpos tumors from six women without recurrences, identified differences between groups based on the ER status of the second tumor. Hypermethylated genes of significantly enriched pathways were differentially associated with survival. DNA methylation profiles of ERpos primary breast tumors support the development and use of tumor methylation profiles for stratifying women with breast cancer both for prognosis and therapy.

TROP2 methylation and expression in tamoxifen-resistant breast cancer.

BACKGROUND: The DNA methyltransferase 1 inhibitor, 5-Aza-2'-deoxycytidine (5-Aza-dC) is a potential treatment for breast cancer. However, not all breast tumors will respond similarly to treatment with 5-Aza-dC, and little is known regarding the response of hormone-resistant breast cancers to 5-Aza-dC. METHODS: We demonstrate that 5-Aza-dC-treatment has a stronger effect on an estrogen receptor-negative, Tamoxifen-selected cell line, TMX2-28, than on the estrogen receptor-positive, MCF7, parental cell line. Using data obtained from the HM450 Methylation Bead Chip, pyrosequencing, and RT-qPCR, we identified a panel of genes that are silenced by promoter methylation in TMX2-28 and re-expressed after treatment with 5-Aza-dC. RESULTS: One of the genes identified, tumor associated calcium signal transducer 2 (TACSTD2), is altered by DNA methylation, and there is evidence that in some cancers decreased expression may result in greater proliferation. Analysis of DNA methylation of TACSTD2 and protein expression of its product, trophoblast antigen protein 2 (TROP2), was extended to a panel of primary (n = 34) and recurrent (n = 34) breast tumors. Stratifying tumors by both recurrence and ER status showed no significant relationship between TROP2 levels and TACSTD2 methylation. Knocking down TACSTD2 expression in MCF7 increased proliferation however; re-expressing TACSTD2 in TMX2-28 did not inhibit proliferation, indicating that TACSTD2 re-expression alone was insufficient to explain the decreased proliferation observed after treatment with 5-Aza-dC. CONCLUSIONS: These results illustrate the complexity of the TROP2 signaling network. However, TROP2 may be a valid therapeutic target for some cancers. Further studies are needed to identify biomarkers that indicate how TROP2 signaling affects tumor growth and whether targeting TROP2 would be beneficial to the patient.

ELF5 and DOK7 regulation in anti-estrogen treated cells and tumors.

BACKGROUND: Most women with primary breast cancers that express estrogen receptor alpha (ER or ESR1) are treated with endocrine therapies including the anti-estrogen tamoxifen, but resistance to these anti-endocrine therapies often develops. This study characterizes the expression of hormone receptors, and the mRNA and DNA methylation levels of docking protein 7 (DOK7), and E74-like factor 5 (ELF5), in 21 novel tamoxifen-resistant cell lines and extends the findings to primary and recurrent human breast tumors. METHODS: Twenty-one tamoxifen-selected cell lines were developed through cloning by limiting dilution of an MCF-7 cell culture treated with 1 µM tamoxifen for 6 months. The parent (MCF-7) and tamoxifen-selected cell lines were characterized for protein expression of ER, progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) using immunohistochemistry (IHC). The mRNA levels of ER, DOK7, and ELF5 were assessed using quantitative RT-PCR. Promoter methylation levels of DOK7 and ELF5 were determined by pyrosequencing of bisulfite-modified DNA. The relationship between hormone receptor status and promoter methylation of DOK7 and ELF5 was further examined using available methylation array data (Illumina HM450) from a set of paired primary and second breast tumors from 24 women. RESULTS: All 21 of the novel tamoxifen-selected cell lines are ER-positive, and HER2-negative, and 18 of the cell lines are PR-negative while the MCF-7 cells were scored as ER-positive, modestly PR-positive and HER2 negative. Expression of DOK7 and ELF5 is significantly up-regulated in half of the tamoxifen-selected cell lines as compared to the parental MCF-7. In contrast, the previously established ER-negative TMX2-28 cell line has decreased expression of both DOK7 and ELF5 and increased DNA methylation in the transcriptional start site region of these genes. ELF5 methylation was lower in second versus primary tumors in women who received anti-estrogen treatment, in PR-negative versus PR-positive tumors, and in the subset of PR-positive first tumors from the group of women who had second PR-negative tumors as compared to those who had second PR-positive tumors. CONCLUSIONS: The distinct ELF5 methylation of PR-positive primary tumors from women who had a PR-negative recurrence indicates the possibility of stratification of women for tailored treatment in the early stages of disease.

Randomized, Double-Blinded Trial of Magnesium Sulfate Tocolysis versus Intravenous Normal Saline for Preterm Nonsevere Placental Abruption.

Objective To evaluate the efficacy and safety of magnesium sulfate in the resolution of vaginal bleeding and contractions in nonsevere placental abruption. Study Design Thirty women between 24 and 34 weeks of gestation diagnosed with nonsevere placental abruption were randomized to receive magnesium sulfate tocolysis or normal saline infusion. The primary outcome was the proportion of women undelivered at 48 hours with resolution of vaginal bleeding and uterine contractions. Maternal and neonatal outcomes were also compared. Results Fifteen (50%) women received magnesium sulfate tocolysis and 15 (50%) received intravenous saline. There was no difference in the number of women who were undelivered at 48 hours with resolution of vaginal bleeding and contractions in the magnesium sulfate (80.0%) and saline (66.7%; p-value = 0.68) groups. There were no differences between groups in the gestational age at randomization, time to uterine quiescence, time on study drug, length of hospitalization, days from randomization to delivery, incidence of side effects, or admissions to the neonatal intensive care unit. Conclusions Magnesium sulfate tocolysis did not provide a significant difference in pregnancy prolongation in the management of preterm nonsevere placental abruption. Recruitment goals were not met due to the introduction of the use of magnesium sulfate for neuroprotection.

We choose: Adolescent girls and young women's choice for an HIV prevention product in a cross-over randomized clinical trial conducted in South Africa, Uganda, and Zimbabwe.

With new pre-exposure prophylaxis (PrEP) modalities for HIV prevention becoming available, understanding how adolescent girls and young women (AGYW) navigate through PrEP options is essential, including factors underlying their choice. Through 16 focus group discussions (FGDs) and 52 in-depth interviews (IDIs) from REACH, an open-label crossover study in which AGYW were allocated 1:1 (between 06 February 2019 and 18 March 2020) to receive oral PrEP for six months and the dapivirine ring for six months, in a randomized sequence, followed by a 6-month period where either product (or neither) could be chosen, we explored decision-making process and product choice, using a mixed inductive-deductive analytical approach. Key themes included the desire to remain HIV-negative and weighing product attributes through experiential learning. Product triability appeared important in informing product choice as individual circumstances changed or assuaging side effects with a given product. Approved biomedical prevention innovations may also benefit from hands-on experience to help with adoption and use during real-world implementation. Furthermore, support from trusted providers will remain critical as AGYW contemplate navigating through PrEP options and choice.

Retinal blood flow association with age and weight in infants at risk for retinopathy of prematurity.

This prospective study evaluated the relationship between laser speckle contrast imaging (LSCI) ocular blood flow velocity (BFV) and five birth parameters: gestational age (GA), postmenstrual age (PMA), and chronological age (CA) at the time of measurement, birth weight (BW), and current weight (CW) in preterm neonates at risk for retinopathy of prematurity (ROP).38 Neonates with BW < 2 kg, GA < 32 weeks, and PMA between 27-47 weeks underwent 91 LSCI sessions. Correlation tests and regression analysis were performed to quantify relationships between birth parameters and ocular BFV. Mean ocular BFV index in this cohort was 8.8 +/- 4.0 IU. BFV positively correlated with PMA (r = 0.3, p = 0.01), CA (r = 0.3, p = 0.005), and CW (r = 0.3, p = 0.02). BFV did not correlate with GA nor BW (r=-0.2 and r=-0.05, p > 0.05). Regression analysis with mixed models demonstrated that BFV increased by 1.2 for every kilogram of CW, by 0.34 for every week of CA, and by 0.36 for every week of PMA (p = 0.03, 0.004, 0.007, respectively). Our findings indicate that increased age and weight are associated with increased ocular BFV measured using LSCI in premature infants. Future studies investigating the associations between ocular BFV and ROP clinical severity must control for age and/or weight of the infant.

Retinal blood flow association with age and weight in infants at risk for retinopathy of prematurity.

This prospective study evaluated the relationship between laser speckle contrast imaging (LSCI) ocular blood flow velocity (BFV) and five birth parameters: gestational age (GA), postmenstrual age (PMA) and chronological age (CA) at the time of measurement, birth weight (BW), and current weight (CW) in preterm neonates at risk for retinopathy of prematurity (ROP). 38 Neonates with BW < 2 kg, GA < 32 weeks, and PMA between 27 and 47 weeks underwent 91 LSCI sessions. Correlation tests and regression analysis were performed to quantify relationships between birth parameters and ocular BFV. Mean ocular BFV index in this cohort was 8.8 +/- 4.0 IU. BFV positively correlated with PMA (r = 0.3, p = 0.01), CA (r = 0.3, p = 0.005), and CW (r = 0.3, p = 0.02). BFV did not correlate with GA nor BW (r = - 0.2 and r = - 0.05, p > 0.05). Regression analysis with mixed models demonstrated that BFV increased by 1.2 for every kilogram of CW, by 0.34 for every week of CA, and by 0.36 for every week of PMA (p = 0.03, 0.004, 0.007, respectively). Our findings indicate that increased age and weight are associated with increased ocular BFV measured using LSCI in premature infants. Future studies investigating the associations between ocular BFV and ROP clinical severity must control for age and/or weight of the infant.

Association of Speckle-Based Blood Flow Measurements and Fluorescein Angiography in Infants with Retinopathy of Prematurity.

Purpose: To determine the correlation between blood flow metrics measured by intravenous fluorescein angiography (IVFA) and the blood flow velocity index (BFVi) obtained by laser speckle contrast imaging (LSCI) in infants with retinopathy of prematurity (ROP). Design: Prospective comparative pilot study. Subjects: Seven eyes from 7 subjects with ROP. Methods: Unilateral LSCI and IVFA data were obtained from each subject in the neonatal intensive care unit. Five LSCI-based metrics and 5 IVFA-based metrics were extracted from images to quantify blood flow patterns in the same region of interest. Correlation between LSCI-based and IVFA-based blood flow metrics was compared between 2 subgroups of ROP severity: moderate ROP (defined as stage ≤ 2 without Plus disease) and severe ROP (defined as stage ≥3 or Plus disease). Main Outcome Measures: Pearson and Kendall rank correlation coefficients between IVFA and LSCI metrics; Student t test P values comparing LSCI metrics between "severe" and "moderate" ROP groups. Results: Pearson correlations between IVFA and LSCI included arterial-venous transit time (AVTT) and peak BFVi (pBFVi; r = -0.917; P = 0.004), AVTT and dip BFVi (dBFVi; r = -0.920; P = 0.003), AVTT and mean BFVi (r = -0.927- P = 0.003), and AVTT and volumetric rise index (r = -0.779; P = 0.039). Kendall rank correlation between AVTT and dBFVi was r = -0.619 (P = 0.051). pBFVi was higher in severe ROP than in moderate ROP (8.4 ± 0.6 and 4.4 ± 1.8, respectively; P = 0.0045 using the 2-sample t test with pooled variance and P = 0.0952 using the Wilcoxon rank-sum test). Conclusions: Correlation was found between blood flow metrics obtained by IVFA and noninvasive LSCI techniques. We demonstrate the feasibility of obtaining quantitative metrics using LSCI in infants with ROP in this pilot study; however, further investigation is needed to evaluate its potential use in clinical assessment of ROP severity. Financial Disclosures: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Improving follow-up rates by optimizing patient educational materials in retinopathy of prematurity.

PURPOSE: To examine the impact of newly designed retinopathy of prematurity (ROP) patient educational materials adherent to health literacy guidelines on improving parent understanding of ROP, perceived importance of follow-up care, and subsequent outpatient follow-up attendance rates. METHODS: This was a repeated-measures study of parents of premature infants at risk for developing ROP. ROP educational materials were redesigned to adhere to current NIH and AMA reading level guidelines. Participants completed surveys that assessed understanding of ROP and perceived importance of clinic follow-up before and after receiving either materials currently available on the website of the American Association for Pediatric Ophthalmology and Strabismus (AAPOS), or the newly designed materials. Results were analyzed to evaluate for an improvement in parent knowledge of ROP and follow-up compliance. RESULTS: Parent ROP knowledge scores improved significantly after receiving educational materials for both the AAPOS materials (55.9% vs 83.7% [P < 0.001]) and the new materials (60.9% vs 91.8% [P < 0.001]). Average post-survey ROP knowledge scores were significantly higher among participants that received the new materials compared to the AAPOS materials (91.8% vs 83.7%, [P < 0.001]). Follow-up attendance rates improved in both groups, with a significantly improved rate from pre-study baseline among the new materials group (80.0% vs 68.2%, [P = 0.008). CONCLUSIONS: Implementation of educational materials significantly improved parent understanding of ROP; combined with knowledge assessment, it also improved follow-up compliance. Materials designed to adhere to health literacy guidelines are the most effective resources for improving knowledge of ROP and follow-up attendance.

How a menu of adherence support strategies facilitated high adherence to HIV prevention products among adolescent girls and young women in sub-Saharan Africa: a mixed methods analysis.

INTRODUCTION: Effective use of pre-exposure prophylaxis (PrEP) has been low among adolescent girls and young women (AGYW) in sub-Saharan Africa. The MTN-034/REACH trial offered AGYW a menu of adherence support strategies and achieved high adherence to both daily oral PrEP and the monthly dapivirine vaginal ring. Understanding how these strategies promoted product use could inform the design of adherence support systems in programmatic settings. METHODS: REACH was a randomized crossover trial evaluating the safety of and adherence to the ring and oral PrEP among 247 HIV-negative AGYW (ages 16-21) in South Africa, Uganda and Zimbabwe from January 2019 to September 2021 (NCT03593655). Adherence support included monthly counselling sessions with drug-level feedback (DLF) plus optional daily short message service (SMS) reminders, weekly phone or SMS check-ins, peer support clubs, "peer buddies" and additional counselling. Counsellors documented adherence support choices and counselling content on standardized forms. Through focus groups, serial in-depth interviews (IDIs) and single IDIs (n = 119 total), we explored participants' experiences with adherence support and how it encouraged product use. RESULTS: Participants received counselling at nearly all visits. DLF was provided at 54.3% of sessions and, across sites, 49%-68% received results showing high adherence for oral PrEP, and 73%-89% for the ring. The most popular support strategies were in-person clubs and weekly calls, followed by online clubs, additional counselling and SMS. Preferences differed across sites but were similar for both products. Qualitative results demonstrated that the REACH strategies supported adherence by providing information about HIV and PrEP, continually motivating participants, and supporting the development of behavioural skills and self-efficacy, aligning with the Information, Motivation, and Behavioural Skills (IMB) model. Effectiveness was supported by three foundational pillars: strong interpersonal relationships with counsellors; ongoing, easily accessible support and resources; and establishing trust in the counsellors and study products through counsellor relationships, peer-to-peer exchange and DLF. CONCLUSIONS: Implementation programmes could support effective PrEP use by offering a small menu of counsellor- and peer-based support options that are youth-friendly and developmentally appropriate. The same menu options can support both ring and oral PrEP users, though content should be tailored to the individual products.

Paralemmin-1 is over-expressed in estrogen-receptor positive breast cancers.

BACKGROUND: Paralemmin-1 is a phosphoprotein lipid-anchored to the cytoplasmic face of membranes where it functions in membrane dynamics, maintenance of cell shape, and process formation. Expression of paralemmin-1 and its major splice variant (Δ exon 8) as well as the extent of posttranslational modifications are tissue- and development-specific. Paralemmin-1 expression in normal breast and breast cancer tissue has not been described previously. RESULTS: Paralemmin-1 mRNA and protein expression was evaluated in ten breast cell lines, 26 primary tumors, and 10 reduction mammoplasty (RM) tissues using real time RT-PCR. Paralemmin-1 splice variants were assessed in tumor and RM tissues using a series of primers and RT-PCR. Paralemmin-1 protein expression was examined in cell lines using Western Blots and in 31 ductal carcinomas in situ, 65 infiltrating ductal carcinomas, and 40 RM tissues using immunohistochemistry. Paralemmin-1 mRNA levels were higher in breast cancers than in RM tissue and estrogen receptor (ER)-positive tumors had higher transcript levels than ER-negative tumors. The Δ exon 8 splice variant was detected more frequently in tumor than in RM tissues. Protein expression was consistent with mRNA results showing higher paralemmin-1 expression in ER-positive tumors. CONCLUSIONS: The differential expression of paralemmin-1 in a subset of breast cancers suggests the existence of variation in membrane dynamics that may be exploited to improve diagnosis or provide a therapeutic target.

Discovery of a Partial Glucokinase Activator Clinical Candidate: Diethyl ((3-(3-((5-(Azetidine-1-carbonyl)pyrazin-2-yl)oxy)-5-isopropoxybenzamido)-1H-pyrazol-1-yl)methyl)phosphonate (BMS-820132).

Glucokinase (GK) is a key regulator of glucose homeostasis, and its small-molecule activators represent a promising opportunity for the treatment of type 2 diabetes. Several GK activators have been advanced into clinical trials and have demonstrated promising efficacy; however, hypoglycemia represents a key risk for this mechanism. In an effort to mitigate this hypoglycemia risk while maintaining the efficacy of the GK mechanism, we have investigated a series of amino heteroaryl phosphonate benzamides as ''partial" GK activators. The structure-activity relationship studies starting from a "full GK activator" 11, which culminated in the discovery of the "partial GK activator" 31 (BMS-820132), are discussed. The synthesis and in vitro and in vivo preclinical pharmacology profiles of 31 and its pharmacokinetics (PK) are described. Based on its promising in vivo efficacy and preclinical ADME and safety profiles, 31 was advanced into human clinical trials.

The Validity of Pulp Tests on Crowned Teeth: A Clinical Study.

OBJECTIVE: The aim of this study was to evaluate the sensitivity and specificity of electric pulp testing (EPT) using the bridging technique in comparison to a cold test on crowned teeth. METHODS: Thirty teeth were included in this study. Only one crowned tooth from each subject was included. The adjacent teeth served as controls. The crowned teeth were tested for pulp vitality using a bridging EPT technique and cold test. Vitality was confirmed upon access based on bleeding or lack of bleeding from detected root canal systems. The data was statistically analyzed using the McNamara test (P<0.05). RESULTS: The sensitivities of the cold test and bridging EPT were 87% and 66% respectively. Accuracy for cold and bridging EPT were 87% and 67% respectively. The cold test demonstrated a statistically significant higher accuracy and sensitivity than the bridging EPT. However, no significant difference was detected in the specificity between the two tests. CONCLUSION: Both EPT and cold test should be considered as an adjunctive diagnostic tool when determining pulp status in a crowned tooth. Pulp sensitivity tests are essential but the results should be interpreted in combination with other clinical signs/symptoms.

Crystal structure of acivicin-inhibited gamma-glutamyltranspeptidase reveals critical roles for its C-terminus in autoprocessing and catalysis.

Helicobacter pylori gamma-glutamyltranspeptidase (HpGT) is a general gamma-glutamyl hydrolase and a demonstrated virulence factor. The enzyme confers a growth advantage to the bacterium, providing essential amino acid precursors by initiating the degradation of extracellular glutathione and glutamine. HpGT is a member of the N-terminal nucleophile (Ntn) hydrolase superfamily and undergoes autoprocessing to generate the active form of the enzyme. Acivicin is a widely used gamma-glutamyltranspeptidase inhibitor that covalently modifies the enzyme, but its precise mechanism of action remains unclear. The time-dependent inactivation of HpGT exhibits a hyperbolic dependence on acivicin concentration with k(max) = 0.033 +/- 0.006 s(-1) and K(I) = 19.7 +/- 7.2 microM. Structure determination of acivicin-modified HpGT (1.7 A; R(factor) = 17.9%; R(free) = 20.8%) demonstrates that acivicin is accommodated within the gamma-glutamyl binding pocket of the enzyme. The hydroxyl group of Thr 380, the catalytic nucleophile in the autoprocessing and enzymatic reactions, displaces chloride from the acivicin ring to form the covalently linked complex. Within the acivicin-modified HpGT structure, the C-terminus of the protein becomes ordered with Phe 567 positioned over the active site. Substitution or deletion of Phe 567 leads to a >10-fold reduction in enzymatic activity, underscoring its importance in catalysis. The mobile C-terminus is positioned by several electrostatic interactions within the C-terminal region, most notably a salt bridge between Arg 475 and Glu 566. Mutational analysis reveals that Arg 475 is critical for the proper placement of the C-terminal region, the Tyr 433 containing loop, and the proposed oxyanion hole.

Regenerative Endodontics: A Systematic Analysis of the Failed Cases.

INTRODUCTION: The aim of this systematic review was to analyze failed cases of regenerative endodontic treatment (RET) reported in the literature in terms of etiology, diagnosis, treatment protocols, signs of failure, and additional endodontic interventions. METHODS: Electronic searches were performed in PubMed, Web of Science, and ProQuest Dissertations & Theses databases. All in vivo publications in humans that reported at least 1 failed case of RET were included in this systematic review. Failed RET cases were defined in the current study as any case that required an additional endodontic intervention or extraction after the completion of the initial RET. RESULTS: A total of 28 studies that reported 67 failed cases of RET were included in this review. A total of 37 failed RET cases reported the etiology that resulted in the initiation of RET; 59% of these cases were caused by dental trauma, and 30% were caused by dens evaginatus. A total of 26 (39%) failed RET cases were detected at least 2 years after the initiation of RET. A total of 53 (79%) failed RET cases were presented with signs and/or symptoms of persistent infection. CONCLUSIONS: Persistent infection was the main presentation in 79% of failed RET cases. Furthermore, 39% of failed RET cases were identified after more than 2 years of follow-up. Future studies should include a detailed description of the etiology, preoperative variables, intraoperative protocols, and postoperative follow-up to provide a better understanding of failed cases after RET.

The Effect of Crown Lengthening on the Outcome of Endodontically Treated Posterior Teeth: 10-year Survival Analysis.

INTRODUCTION: The purpose of this study was to investigate the effect of a crown lengthening (CL) procedure and the crown-root ratio after CL on the long-term survival of endodontically treated teeth (ETT). METHODS: Permanent posterior teeth with opposing dentition that had received adequate nonsurgical root canal treatment (NSRCT) and a full-coverage crown between January 1, 2006, and January 1, 2016 were included in this retrospective study. The data collected included dates of the NSRCT, time of extraction if extracted, age, sex, location, the crown-root ratio after CL, and the presence of a lesion. All included ETT were divided into 2 groups: RESULTS: 5-year survival rates of ETT in the control and CL groups were 88.6% and 82.2%, respectively (P > .05). The 10-year survival rates of ETT in the control and CL groups were 74.5% and 51%, respectively (P < .05). ETT that received the CL procedure after NSRCT were almost 2.3 times more likely to get extracted compared with ETT that did not need the CL procedure at the 10-year follow-up (hazard ratio = 2.29, P < .05). Also, ETT with an inadequate crown-root ratio (1:1) after CL showed the lowest survival rate (40%) compared with ETT with an adequate crown-root ratio (<1:1). CONCLUSIONS: A crown-root ratio of 1:1 after osseous CL may affect the long-term survival of ETT. Despite the promising survival rate of ETT with an adequate crown-root ratio after CL, the long-term survival of NSRCT with an inadequate crown-root ratio (1:1) should be considered in the treatment planning phase. Also, it is worth mentioning that the results of the present study should be evaluated in future prospective studies.