In Vivo Optical Characterization of Middle Ear Effusions and Biofilms During Otitis Media.

Otitis media (OM), a common ear infection, is characterized by the presence of an accumulated middle ear effusion (MEE) in a normally air-filled middle ear cavity. While assessing the MEE plays a critical role in the overall management of OM, identifying and examining the MEE is challenging with the current diagnostic tools since the MEE is located behind the semi-opaque eardrum. The objective of this cross-sectional, observational study is to non-invasively visualize and characterize MEEs and bacterial biofilms in the middle ear. A portable, handheld, otoscope-integrated optical coherence tomography (OCT) system combined with novel analytical methods has been developed. In vivo middle ear OCT images were acquired from 53 pediatric subjects (average age of 3.9 years; all awake during OCT imaging) diagnosed with OM and undergoing a surgical procedure (ear tube surgery) to aspirate the MEE and aerate the middle ear. In vivo middle ear OCT acquired prior to the surgery was compared with OCT of the freshly extracted MEEs, clinical diagnosis, and post-operative evaluations. Among the subjects who were identified with the presence of MEEs, 89.6% showed the presence of the TM-adherent biofilm in in vivo OCT. This study provides an atlas of middle ear OCT images exhibiting a range of depth-resolved MEE features, which can only be visualized and assessed non-invasively through OCT. Quantitative metrics of OCT images acquired prior to the surgery were statistically correlated with surgical evaluations of MEEs. Measurements of MEE characteristics will provide new readily available information that can lead to improved diagnosis and management strategies for the highly prevalent OM in children.

Artificial intelligence-based computer-aided system for knee osteoarthritis assessment increases experienced orthopaedic surgeons' agreement rate and accuracy.

PURPOSE: The aims of this study were to (1) analyze the impact of an artificial intelligence (AI)-based computer system on the accuracy and agreement rate of board-certified orthopaedic surgeons (= senior readers) to detect X-ray features indicative of knee OA in comparison to unaided assessment and (2) compare the results to those of senior residents (= junior readers). METHODS: One hundred and twenty-four unilateral knee X-rays from the OAI study were analyzed regarding Kellgren-Lawrence grade, joint space narrowing (JSN), sclerosis and osteophyte OARSI grade by computerized methods. Images were rated for these parameters by three senior readers using two modalities: plain X-ray (unaided) and X-ray presented alongside reports from a computer-assisted detection system (aided). After exclusion of nine images with incomplete annotation, intraclass correlations between readers were calculated for both modalities among 115 images, and reader performance was compared to ground truth (OAI consensus). Accuracy, sensitivity and specificity were also calculated and the results were compared to those from a previous study on junior readers. RESULTS: With the aided modality, senior reader agreement rates for KL grade (2.0-fold), sclerosis (1.42-fold), JSN (1.37-fold) and osteophyte OARSI grades (3.33-fold) improved significantly. Reader specificity and accuracy increased significantly for all features when using the aided modality compared to the gold standard. On the other hand, sensitivity only increased for OA diagnosis, whereas it decreased (without statistical significance) for all other features. With aided analysis, senior readers reached similar agreement and accuracy rates as junior readers, with both surpassing AI performance. CONCLUSION: The introduction of AI-based computer-aided assessment systems can increase the agreement rate and overall accuracy for knee OA diagnosis among board-certified orthopaedic surgeons. Thus, use of this software may improve the standard of care for knee OA detection and diagnosis in the future. LEVEL OF EVIDENCE: Level II.

Surgical treatment of posterior cruciate ligament lesions does not cause growth disturbances in pediatric patients.

PURPOSE: The aim of the present study was to describe epidemiology, management and outcome of pediatric and adolescent patients with posterior cruciate ligament (PCL) injuries. METHODS: Sixteen patients of less than 18 years of age with 7 PCL avulsion fractures and 9 PCL tears were included over a 10-year period. Trauma mechanism, additional injuries and treatment methods were analyzed. Follow-up examination included range of motion and ability to perform squats. Pedi-IKDC and Lysholm score were obtained and posterior shift was measured in kneeling view radiographs and compared to the contralateral side. Patients were grouped into pediatric patients with open physes at the time surgery and adolescent patients with closing or closed physes. In case of open physes, growth disturbances were assessed. RESULTS: Six of the treated patients (median age 12.5 years, range 10-13) had open physes at time of surgery. Five of those sustained avulsion fractures and treatment consisted of open reduction and screw fixation in four cases and graft reconstruction in one case. One patient sustained a PCL tear and underwent graft reconstruction. Follow-up at a median of 71.5 months (range 62-100) did not reveal any growth disturbances. Median Pedi-IKDC was 71.9 (range 51.7-92.1), median Lysholm score was 81.5 (range 66-88) and median posterior shift difference was 2.5 mm (range 0-11). The remaining 10 patients (median age 16 years, range 14-17) had closing/closed physis at the time of operation. Two patients presented with avulsion fractures treated with open reduction and screw fixation and 8 patients sustained PCL tears treated with graft reconstruction. At a median follow-up of 69.5 months (range 11-112), median Pedi-IKDC was 86.8 (range 36.8-97.7), median Lysholm score was 84.0 (range 45-95) and median posterior shift difference was 4 mm (range 0-15). CONCLUSIONS: In our small number of pediatric patients with PCL injuries, open reduction and epiphyseal screw fixation of displaced avulsed fractures and steep tunnel drilling in case of PCL reconstruction did not cause growth disturbances. Nevertheless, long-term functional impairment should be expected and close follow-up has to be recommended. LEVEL OF EVIDENCE: Therapeutic, Level IV.

Outcome following excision of the radial head in children with open physes for impaired elbow motion.

BACKGROUND: Only few reports have described the outcome of pediatric patients following radial head resection. Therefore, the aim of the present study was to assess clinical and radiologic outcome of patients with open physes following resection of the radial head. MATERIALS AND METHODS: Skeletally immature patients treated with resection of the radial head were included. Range of motion (ROM) of the elbow joint was compared with preoperative values. Grip strength, pronation and supination strength, and carrier angle were compared with the unaffected side. Radiographs were assessed for signs of arthrosis, radial migration, and perifocal ossification. Disabilities of the Arm, Shoulder and Hand and Mayo Elbow Performance scores were obtained. RESULTS: The study included 7 patients (mean age, 11 years), 5 with post-traumatic and 2 with congenitally impaired elbow joint motion. Mean follow-up was 47 months. Pronation/supination ROM improved significantly (P = .018). Extension/flexion ROM did not improve significantly (P = .122). Although grip strength (P = .027) and pronation strength (P = .028) of the affected side were significantly lower compared with the contralateral side, supination strength did not differ significantly (P = .176). The carrying angle was increased in 3 patients. Significant radial migration occurred (mean, 3 mm; standard deviation [SD], 3 mm; P = .018). Arthrosis was found in 3 patients. The mean Disabilities of the Arm, Shoulder and Hand score was 16.1 (range 8.8-30.8; SD, 10.1) and mean Mayo Elbow Performance Score was 88 (range, 70-100; SD, 12). CONCLUSIONS: Radial head excision may be considered for selected patients with open physes in cases of severe impairment of pronation/supination. However, sequelae such as radial migration, arthrosis, and elevation of the carrying angle should be expected.

Activation of Notch-1 in oral epithelial cells by P. gingivalis triggers the expression of the antimicrobial protein PLA2-IIA.

P. gingivalis (Pg) is an oral pathogen with the ability to induce oral dysbiosis and periodontal disease. Nevertheless, the mechanisms by which mucosal responses to the oral microbiota in the presence of specific pathogens such as Pg could abrogate the host-microbe symbiotic relationship leading to periodontitis remain unclear. Herein, we identified the Notch-1/PLA2-IIA axis as a new molecular pathway through which Pg could be specifically modulating oral epithelial antimicrobial and inflammatory responses. Pg activated Notch-1, and inhibition or silencing of Notch-1 completely abrogated Pg-induced PLA2-IIA in oral epithelial cells (OECs). Activation of Notch-1 and PLA2-IIA production were associated with Pg-produced gingipains. Other oral Gram-positive and Gram-negative species failed to induce similar responses. Pg enhanced OEC antimicrobial activity through PLA2-IIA. Increased Notch-1 activation correlated with higher PLA2-IIA gingival expression and changes in the abundance of specific oral bacteria phyla during periodontal disease. Oral bacterial species exhibited differential antimicrobial susceptibility to PLA2-IIA. These findings support previous evidence suggesting an important role for epithelial Notch-1 activation and PLA2-IIA production during health and disease at mucosal surfaces, and provide new mechanistic information concerning the regulation of epithelial antimicrobial and pro-inflammatory responses modulated by oral pathogenic bacteria associated with periodontal disease.

Direct Analysis of Pathogenic Structures Affixed to the Tympanic Membrane during Chronic Otitis Media.

Objective To characterize otitis media-associated structures affixed to the mucosal surface of the tympanic membrane (TM) in vivo and in surgically recovered in vitro samples. Study Design Prospective case series without comparison. Setting Outpatient surgical care center. Subjects and Methods Forty pediatric subjects scheduled for tympanostomy tube placement surgery were imaged intraoperatively under general anesthesia. Postmyringotomy, a portable optical coherence tomography (OCT) imaging system assessed for the presence of any biofilm affixed to the mucosal surface of the TM. Samples of suspected microbial infection-related structures were collected through the myringotomy incision. The sampled site was subsequently reimaged with OCT to confirm collection from the original image site on the TM. In vitro analysis based on confocal laser scanning microscope (CLSM) images of fluorescence in situ hybridization-tagged samples and polymerase chain reaction (PCR) provided microbiological characterization and verification of biofilm activity. Results OCT imaging was achieved for 38 of 40 subjects (95%). Images from 38 of 38 (100%) of subjects observed with OCT showed the presence of additional microbial infection-related structures. Thirty-four samples were collected from these 38 subjects. CLSM images provided evidence of clustered bacteria in 32 of 33 (97%) of samples. PCR detected the presence of active bacterial DNA signatures in 20 of 31 (65%) of samples. Conclusion PCR and CLSM analysis of fluorescence in situ hybridization-stained samples validates the presence of active bacteria that have formed into a middle ear biofilm that extends across the mucosal layer of the TM. OCT can rapidly and noninvasively identify middle ear biofilms in subjects with severe and persistent cases of otitis media.

Hypoxia-inducible transcription factors, HIF1A and HIF2A, increase in aging mucosal tissues.

Hypoxia (i.e. oxygen deprivation) activates the hypoxia-signalling pathway, primarily via hypoxia-inducible transcription factors (HIF) for numerous target genes, which mediate angiogenesis, metabolism and coagulation, among other processes to try to replenish tissues with blood and oxygen. Hypoxia signalling dysregulation also commonly occurs during chronic inflammation. We sampled gingival tissues from rhesus monkeys (Macaca mulatta; 3-25 years old) and total RNA was isolated for microarray analysis. HIF1A, HIF1B and HIF2A were significantly different in healthy aged tissues, and both HIF1A and HIF3A were positively correlated with aging. Beyond these transcription factor alterations, analysis of patterns of gene expression involved in hypoxic changes in tissues showed specific increases in metabolic pathway hypoxia-inducible genes, whereas angiogenesis pathway gene changes were more variable in healthy aging tissues across the animals. With periodontitis, aging tissues showed decreases in metabolic gene expression related to carbohydrate/lipid utilization (GBE1, PGAP1, TPI1), energy metabolism and cell cycle regulation (IER3, CCNG2, PER1), with up-regulation of transcription genes and cellular proliferation genes (FOS, EGR1, MET, JMJD6) that are hypoxia-inducible. The potential clinical implications of these results are related to the epidemiological findings of increased susceptibility and expression of periodontitis with aging. More specifically the findings describe that hypoxic stress may exist in aging gingival tissues before documentation of clinical changes of periodontitis and, so, may provide an explanatory molecular risk factor for an elevated capacity of the tissues to express destructive processes in response to changes in the microbial biofilms characteristic of a more pathogenic microbial challenge.

Noninvasive in vivo optical coherence tomography tracking of chronic otitis media in pediatric subjects after surgical intervention.

In an institutional review board-approved study, 25 pediatric subjects diagnosed with chronic or recurrent otitis media were observed over a period of six months with optical coherence tomography (OCT). Subjects were followed throughout their treatment at the initial patient evaluation and preoperative consultation, surgery (intraoperative imaging), and postoperative follow-up, followed by an additional six months of records-based observation. At each time point, the tympanic membrane (at the light reflex region) and directly adjacent middle-ear cavity were observed in vivo with a handheld OCT probe and portable system. Imaging results were compared with clinical outcomes to correlate the clearance of symptoms in relation to changes in the image-based features of infection. OCT images of most all participants showed the presence of additional infection-related biofilm structures during their initial consultation visit and similarly for subjects imaged intraoperatively before myringotomy. Subjects with successful treatment (no recurrence of infectious symptoms) had no additional structures visible in OCT images during the postoperative visit. OCT image findings suggest surgical intervention consisting of myringotomy and tympanostomy tube placement provides a means to clear the middle ear of infection-related components, including middle-ear fluid and biofilms. Furthermore, OCT was demonstrated as a rapid diagnostic tool to prospectively monitor patients in both outpatient and surgical settings.

Ponseti method is superior to surgical treatment in clubfoot - Long-term, randomized, prospective trial.

Treatment of idiopathic clubfoot has shifted from extensive surgical release to conservative Ponseti method; however, randomized prospective trials to compare both methods are lacking. The goal of this study was to compare the long-term results of Ponseti method with those of surgery in terms of morphology, function, X-rays, overall health and gait. In 2001, a single center, prospective, randomized, and controlled trial with a parallel design was started. Twenty-four feet of 15 patients were randomly assigned to two groups: 12 feet each to a Ponseti and surgical group. Mean follow-up was 10 years. International Clubfoot Study Group score (ICFSG), the Pediatric Outcomes Data Collection Instrument (PODCI) and the Functional Rating System (FRS) were used to compare morphology, functional, radiological and subjective outcomes, and the Oxford Foot Model (OFM) was used to evaluate foot kinematics. No significant difference was seen between groups in the initial Pirani scores (p=0.618). Ponseti group had more favorable long-term outcomes according to the FRS (p=0.005), higher ICFSG, scores for morphology (p=0.021), function (p=0.005) and X-rays (p=0.02), and better parent reported functional status according to the PODCI (p=0.018). Surgically treated clubfeet were stiffer - ankle dynamic range of motion (p=0.015) and the maximal hindfoot dorsiflexion during gait were limited as measured by OFM (p=0.039). This was compensated by an increase in motion between the hindfoot and forefoot. Furthermore, these children had more difficulty playing sports and experienced more pain, which might have led to inferior results in the happiness domain of the PODCI. These results suggest that the Ponseti method provides superior long-term outcomes for idiopathic clubfoot deformity.

Cytokine gene expression profiles during initiation, progression and resolution of periodontitis.

AIM: Variations in the expression of cytokines during the progression of periodontitis remain ill-defined. We evaluated the expression of 19 cytokine genes related to T-cell phenotype/function during initiation, progression and resolution of periodontitis, and related these to the expression of soft and bone tissue destruction genes (TDGs). MATERIALS AND METHODS: A ligature-induced periodontitis model was used in rhesus monkeys (M. mulatta) (n = 18). Gingival tissues were taken at baseline pre-ligation, 2 weeks and 1 month (Initiation) and 3 months (progression) post ligation. Ligatures were removed and samples taken 2 months later (resolution). Total RNA was isolated and the Rhesus Gene 1.0 ST (Affymetrix) used for gene expression analysis. Significant expression changes were validated by qRT-PCR. RESULTS: Disease initiation/progression was characterized by overexpression of Th17/Treg cytokine genes (IL-1ß, IL-6, TGFß and IL-21) and down-regulation of Th1/Th2 cytokine genes (IL-18 and IL-25). Increased IL-2 and decreased IL-10 levels were seen during disease resolution. Several Th17/Treg cytokine genes positively correlated with TDGs, whereas most Th1/Th2 genes exhibited a negative correlation. CONCLUSION: Initiation, progression and resolution of periodontitis involve over- and underexpression of cytokine genes related to various T-helper subsets. In addition, variations in individual T-helper response subset/genes during disease progression correlated with protective/destructive outcomes.

Chemistry of ring-substituted 4-(benzothiazol-2-yl)phenylnitrenium ions from antitumor 2-(4-aminophenyl)benzothiazoles.

Ring-substituted derivatives of 2-(4-aminophenyl)benzothiazole, 1a, 1b-g, are under development as antitumor agents. One derivative, 1f, has reached phase 1 clinical trials as the prodrug 2f, Phortress (NSC 710305). These amines are activated by CYP450 1A1, apparently into hydroxylamines 8a-g that are likely metabolized into esters that ionize into nitrenium ions responsible for cellular damage. Previously we showed that 9a, the acetic acid ester of 8a, generates the long-lived (530 ns) nitrenium ion 11a by hydrolysis or photolysis in water. In this study, azide trapping shows that 9b-g generate 11b-g via rate-limiting N-O heterolysis. Ion lifetimes, estimated from azide/solvent selectivities, range from 250 to 1150 ns with identical lifetimes for 11a and 11f. Differences in biological activity of the amines are likely not due to differences in the chemistry of the cations but to differences in metabolic activation/deactivation of individual amines. Unlike the nitrenium ions, lifetimes of the esters are strongly dependent on the 3'-Me substituent. Esters containing 3'-Me (9b, 9f, 9g) have lifetimes of 5-10 s compared to 400-800 s for esters without 3'-Me (9a, 9c, 9d, 9e). This restricts 3'-Me esters to cells/tissues in which activation occurs, concentrating their effects in tumor cells if metabolism is restricted to those cells.

Reactive electrophilic metabolites of aromatic amine and amide carcinogens.

Aromatic and heterocyclic amines are a major class of chemical mutagens and carcinogens. The toxicity of these compounds is a consequence of their metabolic activation. The best-characterized enzymatic pathways for aromatic amine activation lead to the formation of reactive esters such as N-acetoxyarylamines, which are believed to be precursors of short-lived nitrenium ions. In the 1960s, nitrenium ions were invoked as likely intermediates in the formation of arylamine-derived DNA adducts. More recently, nitrenium ion chemistry has been studied by methods such as trapping with azide ion, laser flash photolysis, and the preparation of highly stabilized of examples (e.g., dianisylnitrenium ion). In this review, we discuss the development of our understanding of nitrenium ion chemistry, with emphasis on their generation in biological systems and their reactions with critical targets such as DNA.

Thrombin generation in pediatric patients with Crohn's disease.

BACKGROUND: Patients with inflammatory bowel disease (IBD) have an increased risk of thromboembolic complications. The pathogenesis of IBD is not really clear and a high thrombin activity might contribute to the pathogenesis. We measured thrombin generation by means of calibrated automated thrombography (CAT), a new tool better reflecting overall hemostasis, in children with Crohn's disease (CD) during active and inactive disease and compared it to conventional markers of activity. We wanted to see whether children with CD have a higher potential for thrombin generation and if there is a correlation between hypercoagulability and disease activity. METHODS: Plasma samples were collected from 22 patients with CD and from 61 healthy children. Thrombin generation was measured by means of CAT. The disease activity was estimated using the Pediatric Crohn's Disease Activity Index (PCDAI). In addition, F1+2, TAT, tissue factor pathway inhibitor (TFPI), fibrinogen, prothrombin (FII), antithrombin (AT), erythrocyte sedimentation rate (ESR), platelet count, α2-globulin, and orosomucoide were measured. RESULTS: In all patients we found a significantly higher endogenous thrombin potential (ETP) and higher peak values during active disease. In accordance with this we also found significantly higher mean ETP values during active disease compared with the control group. We observed a significantly positive correlation between PCDAI and thrombin generation parameters. CONCLUSIONS: Our study clearly shows that the active state of CD in children is associated with the potential for high thrombin generation, but this seems to be caused mainly by the inflammatory process and not by a preexisting propensity for high thrombin generation.

Characterization of the 4-(benzothiazol-2-yl)phenylnitrenium ion from a putative metabolite of a model antitumor drug.

The 4-(benzothiazol-2-yl)phenylnitrenium ion 11 is generated from hydrolysis or photolysis of O-acetoxy-N-(4-(benzothiazol-2-yl)phenyl)hydroxylamine 8, a model metabolite of 2-(4-aminophenyl)benzothiazole 1 and its ring-substituted derivatives that are being developed for a variety of medicinal applications, including antitumor, antibacterial, antifungal, and imaging agents. Previously, we showed that 11 had an aqueous solution lifetime of 530 ns, similar to the 560 ns lifetime of the 4-biphenylylnitrenium ion 12 derived from the well-known chemical carcinogen 4-aminobiphenyl. We now show that the analogy between these two cations extends well beyond their lifetimes. The initial product of hydration of 11 is the quinolimine 16, which can be detected as a long-lived reactive intermediate that hydrolyzes in a pH-dependent manner into the final hydrolysis product, the quinol 15. This hydrolysis behavior is equivalent to that previously described for a large number of ester metabolites of carcinogenic arylamines, including 4-aminobiphenyl. The major azide trapping product (90% of azide products) of 11, 20, is generated by substitution on the carbons ortho to the nitrenium ion center of 11. This product is a direct analogue of the major azide adducts, such as 22, generated from trapping of the nitrenium ions of carcinogenic arylamines. The azide/solvent selectivity for 11, k(az)/k(s), is also nearly equivalent to that of 12. A minor product of the reaction of 11 with N(3)(-), 21, contains no azide functionality but may be generated by a process in which N(3)(-) attacks 11 at the nitrenium ion center with loss of N(2) to generate a diazene 25 that subsequently decomposes into 21 with loss of another N(2). The adduct derived from attack of 2'-deoxyguanosine (d-G) on 11, 28, is a familiar C-8 adduct of the type generated from the reaction of d-G with a wide variety of arylnitrenium ions derived from carcinogenic arylamines. The rate constant for reaction of d-G with 11, k(d-G), is very similar to that observed for the reaction of d-G with 12. The similar lifetimes and chemical reactivities of 11 and 12 can be rationalized by B3LYP/6-31G(d) calculations on the two ions that show that they are of nearly equivalent stability relative to their respective hydration products. The calculations also help to rationalize the different regiochemistry observed for the reaction of N(3)(-) with 11 and its oxenium ion analogue, 13. Since 8 is the likely active metabolite of 1 and a significant number of derivatives of 1 are being developed as pharmaceutical agents, the similarity of the chemistry of 11 to that of carcinogenic arylnitrenium ions is of considerable importance. Consideration should be given to this chemistry in continued development of pharmaceuticals containing the 2-(4-aminophenyl)benzothiazole moiety.

Indirect and direct detection of the 4-(benzothiazol-2-yl)phenylnitrenium ion from a putative metabolite of a model anti-tumor drug.

2-(4-Aminophenyl)benzothiazoles related to 1 are potentially important pharmaceuticals. Metabolism apparently involves oxidation and esterification to 3. In water, hydrolysis and photolysis of 3 generates the nitrenium ion 4 that can be detected indirectly by N(3)(-) trapping and directly by UV-vis spectroscopy following laser flash photolysis. The transient, with lambda(max) 570 nm, and a lifetime of 530 ns, reacts with N(3)(-) at a diffusion-controlled rate and generates the quinol 6 by reaction with water.

Hydrolysis and photolysis of 4-Acetoxy-4-(benzothiazol-2-yl)-2,5-cyclohexadien-1-one, a model anti-tumor quinol ester.

4-Acetoxy-4-(benzothiazol-2-yl)-2,5-cyclohexadien-1-one, 1, a quinol derivative that exhibits significant anti-tumor activity against human breast, colon, and renal cancer cell lines, undergoes hydrolysis in aqueous solution to generate an oxenium ion intermediate, 3, that is selectively trapped by N(3)(-) in an aqueous environment. The 4-(benzothiazol-2-yl) substituent slows the rate of ionization of 1 compared to analogues with 4-phenyl or 4-(p-tolyl) substituents, 4a or 4b. However, once generated, 3 is somewhat more selective than the 4-phenyl-substituted cation 5a. Calculations performed at the B3LYP/6-31G(d) level agree that the 4-(benzothiazol-2-yl) substituent does significantly stabilize 3. The structure of the major isolated azide adduct, 4-(6-azidobenzothiazol-2-yl)phenol, 9, confirms that the positive charge is highly delocalized in 3. The results of hydrolysis of 1 show that the 4-(benzothiazol-2-yl) substituent has a significant inductive electron-withdrawing effect as well as a significant resonance effect that is electron-donating. Photolysis of 1 in aqueous solution generates the quinol 2 as one of several photolysis products. The presence of the quinol suggests that photolysis also leads, in part, to generation of 3, but photoionization of 1 is significantly less efficient than is the case for the esters 4a and 4b. This study proves that 3 is generated by ionization of 1 in an aqueous environment. A significant number of other 2-benzothiazole derivatives that are not quinols, including ring-substituted derivatives of 2-(4-aminophenyl)benzothiazole 15, are under development as anti-tumor agents as well. The possible generation of the reactive intermediate 17 by hydrolysis of the putative metabolite 16 is under investigation.

Multicenter U.S. bilateral MED-EL cochlear implantation study: speech perception over the first year of use.

OBJECTIVE: Binaural hearing has been shown to support better speech perception in normal-hearing listeners than can be achieved with monaural stimulus presentation, particularly under noisy listening conditions. The purpose of this study was to evaluate whether bilateral electrical stimulation could confer similar benefits for cochlear implant listeners. DESIGN: A total of 26 postlingually deafened adult patients with short duration of deafness were implanted at five centers and followed up for 1 yr. Subjects received MED-EL COMBI 40+ devices bilaterally; in all but one case, implantation was performed in a single-stage surgery. Speech perception testing included CNC words in quiet and CUNY sentences in noise. Target speech was presented at the midline (0 degrees), and masking noise, when present, was presented at one of three simulated source locations along the azimuth (-90, 0, and +90 degrees). RESULTS: Benefits of bilateral electrical stimulation were observed under conditions in which the speech and masker were spatially coincident and conditions in which they were spatially separated. Both the "head shadow" and "summation" effects were evident from the outset. Benefits consistent with "binaural squelch" were not reliably observed until 1 yr after implantation. CONCLUSIONS: These results support a growing consensus that bilateral implantation provides functional benefits beyond those of unilateral implantation. Longitudinal data suggest that some aspects of binaural processing continue to develop up to 1 yr after implantation. The squelch effect, often reported as absent or rare in previous studies of bilateral cochlear implantation, was present for most subjects at the 1 yr measurement interval.

Synthesis and decomposition of an ester derivative of the procarcinogen and promutagen, PhIP, 2-amino-1-methyl-6-phenyl-1H-imidazo[4,5-b]pyridine: unusual nitrenium ion chemistry.

The food-derived heterocyclic amine (HCA) carcinogen 2-amino-1-methyl-6-phenyl-1H-imidazo[4,5-b]pyridine, PhIP, is often generated in the highest concentration of the HCAs formed during broiling and frying of meat and fish. Although it is considered to be an important contributor to human cancer risk from exposure to HCAs, the chemistry of PhIP metabolites that presumably react with DNA to initiate carcinogenesis has received only cursory attention. We have synthesized the ester derivative N-pivaloxy-2-amino-1-methyl-6-phenyl-1H-imidazo[4,5-b]pyridine, 1b, and investigated its chemistry in aqueous solution. Although 1b was too unstable to isolate, we could characterize it by NMR methods in DMF-d7, a solvent in which it is stable at -40 degrees C. It decomposed rapidly in aqueous solution, but its conjugate acid, 1bH+, is not reactive. The nitrenium ion, 2, was trapped by N(3)(-) to form the unusual tetrazole adduct, 16. In the absence of N3-, the expected hydration products of 2 were not detected, but the reduction product, 12, was detected. Although such products are often taken as evidence of triplet nitrenium ions, the efficient trapping of 2 by N(3)(-) indicates that it is a ground state singlet species. The product 12 appears to be generated by reduction of an initially formed hydration product of 2. An alternative addition-elimination mechanism for the formation of 12 does not fit the available kinetic data. The selectivity of 2, measured as kaz/ks, the ratio of the second-order rate constant for its reaction with N(3)(-) and the first-order rate constant for its reaction with the aqueous solvent, is (2.3 +/- 0.6) x 10(4) M(-1), a value that is in the middle of the range of k(az)/k(s) of 10-10(6) M(-1) observed for nitrenium ions derived from other HCAs. The mutagenicity of aromatic amines (AAs) and HCAs, measured as the log of histidine revertants per nanomole of amine, log m, in Salmonella typhimurium TA 98 and TA 100 correlates with log(k(az)/k(s)) for a wide variety of carbocyclic and heterocyclic amine mutagens including PhIP. Previously developed linear regression models for mutagenicity that include log(k(az)/k(s)) as an independent variable predict log m for PhIP with good accuracy in both TA 98 and TA 100. Quantitative carcinogenicity data are less strongly correlated with log(k(az)/k(s)), so prediction of the carcinogenicity of PhIP and other HCAs or AAs based primarily on log(k(az)/k(s)) is less successful.

Unusual reactions of the model carcinogen N-acetoxy-N-acetyl-2-amino-alpha-carboline.

The aqueous solution reactions of the title compound, 1, were examined for comparison to those previously reported for another model carcinogen N-pivaloyloxy-2-amino-alpha-carboline, 2. Both of these are models for the ultimate carcinogenic metabolites of 2-amino-alpha-carboline (AalphaC), a food-derived heterocyclic amine mutagen and carcinogen. The present study was undertaken to determine the effect of the N-acetyl group on the chemistry of such compounds. The N-acetyl group slows down N-O bond cleavage by a factor of (5.5 x 10(3))-fold. This allows other reactions not observed in 2, or in other model carcinogens, to be observed. Among these are acyl-transfer reactions to the aqueous solvent, both uncatalyzed and catalyzed by N3-. In addition, the conjugate acid of 1, 1H+, is subject to a spontaneous decomposition not previously observed in other esters of heterocyclic hydroxylamines or hydroxamic acids. This reaction yields the hydroxylamine, 5, and does so without the intermediacy of the hydroxamic acid, 3, and with 18O exchange from the solvent into the hydroxylamine O. This unique reaction may be caused by an intramolecular proton donation by the pyridyl N-H to the amide carboxyl that catalyzes an intramolecular nucleophilic attack by the carboxyl O of 1H+. A nitrenium ion pathway can still be detected for 1, but, unlike 2 and related esters, this reaction is in competition with other processes throughout the pH range of the study.

Correlations of nitrenium ion selectivities with quantitative mutagenicity and carcinogenicity of the corresponding amines.

There is a correlation (r(adj)(2) = 0.5491-0.6338) of quantitative mutagenicity, log m, for a series of heterocyclic (HCAs) and carbocyclic (AAs) aromatic amines in Salmonella typhimurium TA 98 (18 amines) and TA 100 (15 amines) vs log S, the log of the azide/solvent selectivity of the corresponding nitrenium ion. Monocyclic aromatic amines, MAAs, are less mutagenic than other amines of similar log S. Multiple variable linear regression analysis led to a two parameter regression model, significant at the 95% confidence level for both variables, that includes log S and a ring index variable, I(rings), that is 0 for MAAs and 1 for all other amines. These models have r(adj)(2) = 0.8448 for TA 98, and 0.8927 for TA 100. Inclusion of a third variable, Clog P, increases r(adj)(2) to 0.8913 for TA 98 and 0.9011 for TA 100. This model is significant at the 95% confidence level for all variables for TA 98, but only for two of the three variables for TA 100. The confidence level is 80% for Clog P in TA 100. Quantitative carcinogenicity data in mice and rats are more weakly correlated with log S (r(adj)(2) = 0.5357 for 12 amines in mice, r(adj)(2) = 0.4216 for 10 amines in rats). Several two parameter regression models, all containing Clog P and one containing log S, adequately correlate the mouse data.