A retrospective study of Enterococcus faecalis infective endocarditis: comparison of clinical characteristics and outcomes associated with treatment.

Introduction: A synergistic antibiotic combination of a penicillin and gentamicin (AG) or ceftriaxone (AC) is used in the management of Enterococcus faecalis infective endocarditis (EFIE). We compare the treatment outcomes between AG and AC, including low and high dose ceftriaxone (1 and 2 g 12 hourly). Methods: A retrospective cohort study of patients treated for EFIE at single tertiary centre (2012-2019). Outcome measures examined were 90- and 180-day mortality, treatment associated adverse events and relapse of bacteraemia (within 1 year). Results: 39 patients were enrolled [61.6% given (AC) (n = 24), 24% received ACL (n = 10) and 34% received ACN (n = 14)], 38.4% received AG (n = 15). We noted a difference in the mortality outcomes at 90 and 180 days between those treated with AG and AC overall (6.7% and 33.3%, respectively) although this did not reach statistical significance (P = 0.114, P = 0.061). No significant difference was noted between these groups in incidence of relapsed bacteraemia with two cases noted in the AC cohort (8.3%, 2/24) and none observed (0/15) in the AG cohort (P = 0.662, P = 0.414). A greater number of adverse events was observed in the AG group (11/15, 73.3%) compared to the overall AC group (6/24, 25.0%) (P = 0.009), with no difference between the high and low dose ceftriaxone groups (P = 0.05). Conclusion: Combination treatment of EFIE with AC is associated with a reduced number of adverse events in comparison to AG groups. Although increased mortality was observed in the AC group, this did not reach statistical significance, and reflects the greater comorbidities and reduced capacity for surgical source control in this cohort.

Nuclear F-actin Cytology in Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma.

Oral cavity cancer has a low 5-y survival rate, but outcomes improve when the disease is detected early. Cytology is a less invasive method to assess oral potentially malignant disorders relative to the gold-standard scalpel biopsy and histopathology. In this report, we aimed to determine the utility of cytological signatures, including nuclear F-actin cell phenotypes, for classifying the entire spectrum of oral epithelial dysplasia and oral squamous cell carcinoma. We enrolled subjects with oral potentially malignant disorders, subjects with previously diagnosed malignant lesions, and healthy volunteers without lesions and obtained brush cytology specimens and matched scalpel biopsies from 486 subjects. Histopathological assessment of the scalpel biopsy specimens classified lesions into 6 categories. Brush cytology specimens were analyzed by machine learning classifiers trained to identify relevant cytological features. Multimodal diagnostic models were developed using cytology results, lesion characteristics, and risk factors. Squamous cells with nuclear F-actin staining were associated with early disease (i.e., lower proportions in benign lesions than in more severe lesions), whereas small round parabasal-like cells and leukocytes were associated with late disease (i.e., higher proportions in severe dysplasia and carcinoma than in less severe lesions). Lesions with the impression of oral lichen planus were unlikely to be either dysplastic or malignant. Cytological features substantially improved upon lesion appearance and risk factors in predicting squamous cell carcinoma. Diagnostic models accurately discriminated early and late disease with AUCs (95% CI) of 0.82 (0.77 to 0.87) and 0.93 (0.88 to 0.97), respectively. The cytological features identified here have the potential to improve screening and surveillance of the entire spectrum of oral potentially malignant disorders in multiple care settings.

Large Draining Focal Fibrous Hyperplasia Secondary to Periapical Granuloma.

Periapical granuloma is a pathological diagnosis associated clinically and radiographically with a nonvital tooth and a periapical radiolucency, respectively. It is frequently seen as a sequela of long-standing pulpal necrosis. Often times, a draining fistula is observed near the nonvital tooth. We report an unusual case of a large draining focal fibrous hyperplasia in association with a large periapical granuloma treated at our clinic. The diagnosis was made by the clinical presentation, radiologic and histopathologic findings.

Programmable bio-nanochip-based cytologic testing of oral potentially malignant disorders in Fanconi anemia.

Fanconi anemia (FA) is caused by mutations of DNA repair genes. The risk of oral squamous cell carcinoma (OSCC) among FA patients is 800-folds higher than in the general population. Early detection of OSCC, preferably at it precursor stage, is critical in FA patients to improve their survival. In an ongoing clinical trial, we are evaluating the effectiveness of the programmable bio-nanochip (p-BNC)-based oral cytology test in diagnosing oral potentially malignant disorders (OPMD) in non-FA patients. We used this test to compare cytomorphometric and molecular biomarkers in OSCC cell lines derived from FA and non-FA patients to brush biopsy samples of a FA patient with OPMD and normal mucosa of healthy volunteers. Our data showed that expression patterns of molecular biomarkers were not notably different between sporadic and FA-OSCC cell lines. The p-BNC assay revealed significant differences in cytometric parameters and biomarker MCM2 expression between cytobrush samples of the FA patient and cytobrush samples of normal oral mucosa obtained from healthy volunteers. Microscopic examination of the FA patient's OPMD confirmed the presence of dysplasia. Our pilot data suggests that the p-BNC brush biopsy test recognized dysplastic oral epithelial cells in a brush biopsy sample of a FA patient.

Myositis ossificans circumscripta of the buccinator muscle: First report of a rare complication of mandibular third molar extraction

Myositis ossificans is a self-limiting ossifying process that most often develops following mechanical trauma to skeletal musculature. It chiefly affects the skeletal muscles of extremities of young athletically active adult males. Myositis ossificans is rare in children except for children affected byheritable disorder known as progressive myositis ossificans (fibrodysplasia ossificans progressiva). Children with this disorder develop ossification of muscles and associated soft tissue in early childhood without prior history of trauma. Traumatic form of myositis ossificans also known as myositis ossificans circumscripta (MOC) is rarely encountered in the head and neck musculature. We report a case of MOC within the buccinator which developed as a postoperative complication of mandibular third molar surgery. During extraction of a left mandibular third molar in a 16-year oldmale, a tooth fragment was accidently displaced into the adjacent soft tissue. Retrieval of this tooth fragment caused significant soft tissue trauma. Eighteen months after his third molar surgery, the patient continued to have pain and tenderness anterior to the left mandibular ramus. Radiographic imaging revealed a well-defined ovoid radiopaque mass within the left buccinator muscle. The lesion was surgically removed and the post-surgical course of the patient was uneventful. Histologicalfindings of the mass were characteristic for myositis ossificans.(AU)

Spontaneous bilateral quadriceps tendon rupture.

Spontaneous bilateral quadriceps tendon ruptures are uncommon. We present a 30-year-old man with end-stage renal failure, who sustained this injury, and subsequently had surgical repair of both tendons on separate occasions. He has since regained full range of movement of both knees.

Aggressive osteoblastoma of the mandible closely simulating calcifying epithelial odontogenic tumor. Report of two cases with unusual histopathologic findings.

Aggressive osteoblastoma is a rare bone-forming neoplasm composed of prominent epithelioid cells that demonstrate locally invasive growth with a high rate of recurrence but no metastatic potential. Clinical, radiographic and pathologic features of mandibular aggressive osteoblastoma in a 21-year-old African-American male and a 12-year-old Caucasian female are presented. Both tumors were resected with wide surgical margins and neither patient had adjuvant radiation or chemotherapy. The patients showed no evidence of local recurrence or distant spread either clinically or radiographically after two years of follow-up. These tumors were composed of solid sheets of pleomorphic epithelioid cells, eosinophilic amorphous osteoid with foci of calcification, which closely simulated amyloid. Differentiation of this tumor from histologically similar calcifying epithelial odontogenic tumor and low-grade osteosarcoma proved difficult. Immunohistochemical study with osteocalcin confirmed the osteoblastic nature of these epithelioid cells.

Intra- and intermolecular triplex DNA formation in the murine c-myb proto-oncogene promoter are inhibited by mithramycin.

Mithramycin inhibits transcription by binding to G/C-rich sequences, thereby preventing regulatory protein binding. However, it is also possible that mithramycin inhibits gene expression by preventing intramolecular triplex DNA assembly. We tested this hypothesis using the DNA triplex adopted by the murine c-myb proto-oncogene. The 5'-regulatory region of c-myb contains two polypurine:polypyrimidine tracts with imperfect mirror symmetry, which are highly conserved in the murine and human c-myb sequences. The DNA binding drugs mithramycin and distamycin bind to one of these regions as determined by DNase I protection assay. Gel mobility shift assays, nuclease and chemical hypersensitivity and 2D-gel topological analyses as well as triplex-specific antibody binding studies confirmed the formation of purine*purine:pyrimidine inter- and pyrimidine*purine:pyrimidine intra-molecular triplex structures in this sequence. Mithramycin binding within the triplex target site displaces the major groove-bound oligonucleotide, and also abrogates the supercoil-dependent H-DNA formation, whereas distamycin binding had no such effects. Molecular modeling studies further support these observations. Triplex-specific antibody staining of cells pretreated with mithramycin demonstrates a reversal of chromosomal triplex structures compared to the non-treated and distamycin-treated cells. These observations suggest that DNA minor groove-binding drugs interfere with gene expression by precluding intramolecular triplex formation, as well as by physically preventing regulatory protein binding.

Variable expression of cathepsin B and D correlates with highly invasive and metastatic phenotype of oral cancer.

The expression levels of cathepsins B, D, and L in oral cancer surgical specimens were determined using immunocytochemical analysis. Cathepsins B and D are frequently overexpressed in squamous cell carcinomas, whereas their overexpression was less frequent in verrucous carcinoma and basaloid squamous cell carcinomas. Elevated level of cathepsin B in oral carcinomas was significantly associated with advanced tumor stage (P < .05) and poor histologic malignancy grade (P < .001). Increased expression of cathepsin D correlated significantly with the presence of metastasis (P < .05), poor histologic malignancy grade (P < .001), and high proliferation rate (P < .05). Cathepsin L was less frequently overexpressed in oral cancers than cathepsin B and D. These findings indicate that there is a strong cause/effect relationship between the expression levels of cathepsin B and D in oral cancers and their local invasive and metastatic growth patterns. Thus, cathepsins B and D are useful prognostic markers as well as promising gene therapy targets for oral cancer.

The integral divalent cation within the intermolecular purine*purine. pyrimidine structure: a variable determinant of the potential for and characteristics of the triple helical association.

In vitro assembly of an intermolecular purine*purine.pyrimidine triple helix requires the presence of a divalent cation. The relationships between cation coordination and triplex assembly were investigated, and we have obtained new evidence for at least three functionally distinct potential modes of divalent cation coordination. (i) The positive influence of the divalent cation on the affinity of the third strand for its specific target correlates with affinity of the cation for coordination to phosphate. (ii) Once assembled, the integrity of the triple helical structure remains dependent upon its divalent cation component. A mode of heterocyclic coordination/chelation is favorable to triplex formation by decreasing the relative tendency for efflux of integral cations from within the triple helical structure. (iii) There is also a detrimental mode of base coordination through which a divalent cation may actively antagonize triplex assembly, even in the presence of other supportive divalent cations. These results demonstrate the considerable impact of the cationic component, and suggest ways in which the triple helical association might be positively or negatively modulated.

Inhibition of transcription of the human c-myc protooncogene by intermolecular triplex.

Triplex-forming oligonucleotides (TFOs) have been shown to inhibit both transcription in vitro and the expression of target genes in cell culture by binding to polypurine/polypyrimidine sequences in several human gene promoters. The c-myc protooncogene is overexpressed in a variety of human cancers and appears to play an important role in the proliferation of these cells. In an attempt to assay the ability of triplex-forming oligonucleotides to inhibit expression of a target gene in vivo, we have developed a cellular system involving transfection of a c-myc promoter-driven luciferase reporter plasmid with triplex-forming oligonucleotides targeted to the human c-myc protooncogene. To increase the stability of the TFO, we have used modified phosphorothioate oligonucleotides. Triplex formation with a modified phosphorothioate oligonucleotide occurs with approximately equal binding affinity as that seen using a phosphodiester oligonucleotide. Phosphorothioate-modified TFOs targeted to c-myc inhibit transcription of the c-myc promoter in HeLa cells as demonstrated by a decrease in luciferase expression from a luciferase reporter gene construct. These results suggests that triplex formation may represent a gene-specific means of inhibiting specific protooncogene expression.

Ameloblastic fibrosarcoma in a bull.

This report describes a malignant odontogenic neoplasm in a 7-year-old bull. The mass, involving the right mandible, was locally invasive and destructive. Histologically, it consisted of islands and cords of benign odontogenic epithelium, entrapped in a population of malignant mesenchymal cells. These morphological features are characteristic of ameloblastic fibrosarcoma in man, an odontogenic tumour not previously described in animals.

Androgen receptor status of the oral sebaceous glands.

In recent years, the receptor status of the estrogens and progestins and their relationship to the oral mucosa and its disease processes have been described. However, investigations regarding the androgens have largely been neglected. The androgens have diverse physiologic effects throughput the body and mediate their actions through androgen receptors (ARS) in various target organs. One of these, the sebaceous gland of the skin (dermal sebaceous gland, DSG) has been evaluated extensively in regard to ARS status. However, to our knowledge, the ARS status of the oral sebaceous glands (OSG) has not yet been elucidated. We attempt to evaluate the ARS status of 10 cases of OSG, in addition to other tissue components of the oral mucosa, using immunohistochemical techniques with a monoclonal antibody (MoAb) ARS. Sebaceous glands in all cases examined exhibited a moderate to more intense degree in immunoreactivity for ARS. Other tissue components, however, including the overlying epithelium and vascular endothelium, were essentially nonreactive. Our results indicate that, like the DSG, the OSG may be a target for the androgenetic hormones.

Efficient delivery of triplex forming oligonucleotides to tumor cells by adenovirus-polylysine complexes.

Oligonucleotides (ODNs) show great promise in their ability to specifically inhibit single gene expression but must cross the cell membrane, escape the endosomal vesicle, and possibly traverse the nuclear membrane to arrive at their intracellular target molecules. In an attempt to improve the delivery of phosphodiester triplex forming ODNs to malignant cells, we have constructed adenovirus-polylysine (AdpL)-ODN complexes designed to take advantage of the receptor mediated endocytosis of adenoviruses to transfer the ODNs to the cell nucleus. Treatment of several different types of tumor cells in culture by AdpL-ODN complex resulted in superior uptake and persistence of the ODNs compared to both free ODN and cationic lipid-ODN complexes. Nuclear uptake peaks at 4 h and intact ODN persists in the nucleus with a half-life of 12 h. ODN concentrations of 20-70 microM are achieved at 24 h in all monolayer cell lines evaluated to date. ODNs are detected in 50-100% of the total cell population by immunohistochemistry with apparent uptake into vesicles and nuclear localization. Luciferase expression of a co-delivered reporter plasmid suggests that these ODNs are free in the nucleus. AdpL-ODN complexes will provide a valuable tool for delivering unmodified ODNs to the nucleus of malignant cells.

Influence of GC and AT specific DNA minor groove binding drugs on intermolecular triplex formation in the human c-Ki-ras promoter.

We have used DNase I footprinting and gel shift assays to characterize the interaction of DNA binding drugs mithramycin, distamycin, and berenil with an intermolecular triplex formed by the human c-Ki-ras promoter. A purine-rich triplex-forming oligonucleotide (ODN) forms a stable intermolecular triple helix (triplex) with a homopurine (PR):homopyrimidine (PY) motif in the human c-Ki-ras promoter which contains a 22bp PR:PY region (-328 to -307). This triplex structure is comprised of 15 G.G:C triplets interspersed with 7 T.A:T triplets. Mithramycin binding sites in the human c-Ki-ras promoter encompass most of the triplex target site and three G-C-rich sequences downstream of this triplex-forming region. Mithramycin binding within the c-Ki-ras promoter completely abrogates triplex formation. Furthermore, the addition of mithramycin to pre-formed triplex by c-Ki-ras promoter displaces the major groove bound ODN. Five prominent distamycin binding sites are noted within the c-Ki-ras promoter including the triplex-forming site as well as A-T-rich regions upstream and downstream of the triplex site. Berenil does not bind within the triplex target sequence, and only one berenil binding sequence downstream of the triplex motif was present within the c-Ki-ras promoter fragment. Neither distamycin nor berenil prevents triplex formation, and, furthermore, the addition of either distamycin or berenil to the pre-formed triplex structure did not displace the major-groove-bound third strand. This study demonstrates that GC-specific and AT-specific minor groove ligands differentially affect the intermolecular pur.pur:pyr triplex. A possible biological significance of mithramycin interaction with intramolecular triplex is discussed.