Efficacy of image-guided synovial biopsy.

OBJECTIVE: A variety of benign and neoplastic lesions can affect the synovium, including pigmented villonodular synovitis (PVNS) and synovial chondromatosis. Prior to surgical resection, accurate characterization of synovial lesions is necessary for appropriate treatment planning. Additionally, recent advances in potential medical therapies for PVNS could decrease or eliminate the need for surgery in some cases. Such treatment options demand accurate characterization of synovial lesions prior to treatment. METHODS AND MATERIALS: Institutional IRB approval was obtained. We identified 54 synovial biopsies performed at our institution using a comprehensive database search under ultrasound (US) or computed tomography (CT) guidance. Cases were reviewed for pre-procedure imaging, location, biopsy approach, biopsy results, post-procedure complications, and surgical pathology if synovectomy was performed. RESULT: A total of 54 image-guided synovial biopsies were performed, 36 using CT guidance and 18 using US guidance. Six different anatomic locations were biopsied (the hip, knee, shoulder, elbow, ankle, and temporomandibular joint). Synovial tissue was obtained in 89% of cases (48/54). CT-guided biopsies had a positive yield of 86% (31/36) and US-guided biopsies had a positive yield of 94% (17/18). Surgical pathology was obtained in 30 of the cases and image-guided biopsy concordance was 90% (27/30). Of the patients taken for synovectomy, biopsy concordance of suspected neoplastic lesions was 100% (23/23). In cases of suspected neoplasm, the concordance between image-guided biopsy and surgical pathology was 96% (22/23). There were no reported complications. CONCLUSION: Image-guided biopsy of synovial lesions is safe and effective for establishing a definitive diagnosis prior to surgical or other intervention.

Real-world diagnostic potential of bacterial biomarkers of canine periodontitis.

Introduction: The objective of this study was to investigate the diagnostic potential of bacterial biomarkers by comparing the performance of molecular detection assays with clinical assessments of dog's oral health performed by veterinarians. Methods: Supragingival and subgingival plaque samples were collected from 127 client-owned dogs, pre-booked for procedures under general anesthesia, visiting veterinary practices in the United States. DNA was extracted and bacterial biomarkers quantified using quantitative polymerase chain reaction. Gingivitis and periodontitis were recorded by a trained clinician using the Weighted Gingivitis Periodontitis Score which involved assessing the buccal surfaces of 18 teeth while under general anesthesia. Intraoral dental radiographs of the left and right mandibular first molar teeth were also obtained. These data were then used to establish the diagnostic performance of the molecular assay to detect periodontitis. Results: An initial conscious, visual oral examination performed by the veterinarian identified 67.7% of dogs as having periodontitis, but examination under general anesthesia indicated a higher proportion (86.6%). Analysis of supragingival plaque samples collected by veterinarians from conscious and unconscious dogs demonstrated the assay had an accuracy of 77.7 to 80.9%, a sensitivity of 77.6 to 81.0%, and a specificity of 80.0%. Discussion: Use of this molecular screening tool in conscious dogs has the potential to improve early periodontal disease detection and support veterinary decision making, ultimately improving the oral health of dogs and consequently their quality of life.

3-O sulfation of syndecan-1 mediated by the sulfotransferase HS3ST3a1 enhances myeloma aggressiveness.

Multiple myeloma is a hematological neoplasm derived from plasma cells invariably developing in the bone marrow (BM). The persisting clinical challenge in MM resides in its high ability to resist drugs as shown by the frequent relapses observed in patients regardless of the treatment applied. In a mouse model of MM, we identified a subpopulation of cells harboring increased resistance to current MM drugs. These cells bound a proliferation inducing ligand (APRIL), a key MM promoting/survival factor. APRIL binding involved the heparan sulfate (HS) chain present on syndecan-1 (SDC-1), and correlated with reactivity to the anti-HS antibody 10e4. 10e4+cells had a high proliferation activity, and were able to form colonies in 3-D cultures. 10e4+ cells were the only cells able to develop in BM after intravenous injection. They also resisted drugs in vivo, since their number increased after treatment in BM. Notably, 10e4+ cells differentiated into 10e4- cells upon in vitro and in vivo expansion. Expression of one sulfotransferase, HS3ST3a1, allowed modification of syndecan-1 to confer reactivity to 10e4 and binding to APRIL. HS3ST3a1 deletion inhibited tumorigenesis in BM. Notably, the two populations coexisted at a variable frequency in the BM of MM patients at diagnosis. In total, our results indicate that 3-O-sulfation on SDC-1 carried out by HS3ST3a1 defines aggressive MM cells, and that targeting of this enzyme could possibly be used to better control drug resistance.

The effects of conformation and intermolecular hydrogen bonding on the structure and IR spectra of flutamide; a study based on the matrix isolation technique, ab initio and DFT calculations.

In this study, stable conformers of flutamide referred to as an anticancer drug were searched through a relaxed potential energy surface scan carried out at the B3LYP/6-31G(d) level of theory. This was followed by geometry optimization and thermochemistry calculations performed with the HF-SCF, MP2, B3LYP methods and the 6-31G(d), 6-311++G(d,p), aug-cc-pvTZ basis sets for each of the determined minimum energy conformers. The results revealed that flutamide has at least five stable conformers and two of them provide the major contribution to the observed matrix isolation infrared (IR) spectra of the molecule. The effects of conformational variety and intermolecular hydrogen bonding interactions on the observed IR spectra of flutamide were interpreted in the light of the vibrational spectral data obtained for the most stable monomer and dimer forms of the molecule at the same levels of theory. Pulay's "Scaled Quantum Mechanical-Force Field (SQM-FF)" method was used in the refinement of the calculated harmonic wavenumbers, IR intensities and potential energy distributions. This scaling method which proved its superiority to both anharmonic frequency calculations and other scaling methods helped us to correctly interpret the remarkable differences between the matrix IR spectra of flutamide in argon and the condensed phase IR spectra of the molecule in solvents such as KBr, H2O, D2O, ethanol and methanol.

[Targeted therapy in lung cancer: molecular testing using cytological specimens]. / Thérapies ciblées du cancer pulmonaire: tests moléculaires à partir d'échantillons cytologiques.

Important advances in lung cancer treatment have been made over the last decade. Several drugs designed to target molecular pathways involved in cancer-cell growth and survival have been shown to be effective in a selected fraction (<20%) of non-small cell lung cancer patients. Somatic mutations in several genes (i.e.: EGFR and KRAS) can predict patient's response to targeted therapies. Those mutations are commonly detected on histopathological samples (core-needle biopsy/ surgical resection). However, when tissue biopsies are not available, molecular testing has to be performed on cytological specimens. Issues raised by molecular testing on cytological specimen are discussed in this article.

Visceral leishmaniasis in a kidney transplant recipient: parasitic interstitial nephritis, a cause of renal dysfunction.

Visceral leishmaniasis (VL) due to Leishmania infantum is an endemic parasitic infection in the Mediterranean area. It most commonly affects immunosuppressed individuals, especially HIV patients and less frequently organ transplant recipients. Renal involvement seems to be frequent and is mostly associated with tubulointerstitial nephritis, as described in autopsy reports. In the 61 cases of renal transplant recipients with VL reported in the literature, renal dysfunction was noted at clinical presentation and was more frequently observed as a complication of antiparasitic therapy. However, no pathological analysis of the allograft lesions was reported. We present the case of a Swiss renal transplant recipient who developed VL after vacations in Spain and Tunisia, complicated by acute parasitic nephritis in the renal allograft 3 months after a well-conducted treatment of liposomal amphotericin B.

Diagnostic precision of image-guided multisampling core needle biopsy of suspected lymphomas in a primary care hospital.

We evaluated the diagnostic quality of image-guided multisampling core needle biopsy (CNB) in patients investigated for suspected lymphoma in a primary care hospital. A total of 112 patients were consecutively assessed during a 3-year period. There were 80 lymphoid site biopsies and 32 non-lymphoid site biopsies. Eight to nine cores were obtained from different parts of the biopsy site. Two cores were systematically frozen, allowing for further morphological, immunochemistry and molecular studies. The diagnostic yield of CNB for malignancy was 100%. Only 47% (41/87) of patients with initial suspicion of lymphoma were finally diagnosed with Lymphoma. The diagnostic yield of CNB for lymphoma typing was 98% (62/63), according to the WHO classification. The diagnostic yield of CNB for complete lymphoma subtyping/grading was 86% (54/63). The diagnostic yield of CNB for a definite diagnosis of benignity was only 47% (8/17). In a primary care setting, multisampling CNB is a minimally invasive, and very accurate procedure for confirming malignancy in patients with suspected lymphoma, presenting with superficial/deep-seated, lymphoid/non-lymphoid site targets. With a very high diagnostic yield for lymphoma typing and a high diagnostic yield for complete lymphoma subtyping/grading a therapeutic decision can be taken in most patients.

The source of APRIL up-regulation in human solid tumor lesions.

Abundant mRNA expression for a proliferation-inducing ligand (APRIL) from tumor necrosis factor (TNF) family is observed in many solid tumors. Here, we analyzed in situ the cellular source of APRIL in human solid tumors with anti-APRIL antibodies. In most cases, neutrophils present in the tumor stroma constituted the main source of APRIL. In cutaneous lesions such as melanoma or basal cell carcinoma, tumor-adjacent keratinocytes also produced APRIL. APRIL production by tumor cells themselves was a rare event, only observed in urothelial bladder cancer and squamous cell carcinoma. Detailed analysis revealed that APRIL dissociated from producing cells, and secreted APRIL was retained in the tumor lesions. A direct binding onto tumor cells via heparan sulfate proteoglycans (HSPG) was observed in in vitro experiments and confirmed in situ. Taken together, our analysis indicates a potential role for HSPG/APRIL interactions in the development of solid tumors.

Myelopoiesis dysregulation associated to sustained APRIL production in multiple myeloma-infiltrated bone marrow.

Multiple myeloma (MM) is a non-curable tumor developing in the bone marrow (BM). The BM microenvironment rich in hematopoietic precursors is suspected to have a role in MM development. Here we show that a proliferation-inducing ligand (APRIL) mediated in vivo MM promotion. In MM-infiltrated BM, APRIL originated from differentiating myeloid cells with an expression peak in precursor cells. Notably, APRIL expression stayed stable in BM despite MM infiltration. The pool of APRIL-producing cells changed upon MM infiltration. Although CD16(+) mature myeloid cells constituted about half of the APRIL-producing cells in healthy BM, CD16(-) Elastase(+) myeloid precursor cells were predominant in MM-infiltrated BM. Myeloid precursor cells secreted all the APRIL they produced, and binding of secreted APRIL to MM cells, strictly dependent of heparan sulfate carried by CD138, resulted in an in situ internalization by tumor cells. This indicated APRIL consumption by MM in BM. Taken together, our data show that myelopoiesis dysregulation characterized by an increased proportion of precursor cells occurs in MM patients. Such dysregulation correlates with a stable expression of the MM-promoting factor APRIL in infiltrated BM.

Structure-activity modifications of the HIV-1 inhibitors (+)-calanolide A and (-)-calanolide B.

The delta 7,8 olefinic linkages within (+)-calanolide A(1) and (-)-calanolide B(2) were catalytically reduced to determine impact on the anti-HIV activity of the parent compounds. In addition, a series of structure modifications of the C-12 hydroxyl group in (-)-calanolide B was made to investigate the importance of that substituent to the HIV-1 inhibitory activity of these coumarins. A total of 14 analogs were isolated or prepared and compared to (+)-calanolide A and (-)-calanolide B in the NCI primary anti-HIV assay. While none of the compounds showed activity superior to the two unmodified leads, some structure-activity requirements were apparent from the relative anti-HIV potencies of the various analogs.

HIV-inhibitory natural products. 11. Comparative studies of sulfated sterols from marine invertebrates.

A total of 22 sulfated sterols isolated from marine sponges, ophiuroids (brittle stars), and asteroids (sea stars) were comparatively evaluated for their antiviral activity against HIV-1 and HIV-2. In general, sterols with sulfate groups at position 2, 3, or 6 were the most active, with EC50 values of 3-13 microM against HIV-1 (RF) and 2-8 microM against HIV-2 (CBL20). Those compounds which were sulfated on the sterol D ring were completely inactive against both HIV-1 and HIV-2. Overall, sulfated sterols active against HIV-1 were also active against HIV-2.

Novel LCMV-specific H-2k restricted CTL clones recognize internal viral gene products and cause CNS disease.

H-2k (C3H/Hej) cytotoxic T lymphocytes (CTL) specific for lymphocytic choriomeningitis virus (LCMV) were cloned. Three clones recognizing internal viral antigens were studied. One such CTL clone recognized neither the glycoprotein nor nucleoprotein encoded by the viral short RNA segment, but reacted with a protein encoded by the long RNA segment, either the viral polymerase, or the Z protein. This one clone, in addition to primary CTL harvested from immunized C3H mice, failed to lyse target cells expressing the Z protein, suggesting recognition was to the viral polymerase. Two other clones recognized the viral nucleoprotein, amino acids 93-100, as determined by protein deletion and peptide mapping studies. When introduced directly into the central nervous systems of LCMV-infected histocompatible mice, all clones were active in vivo and capable of causing immunopathologically mediated death.

Use of recombinant vectors derived from herpes simplex virus 1 mutant tsK for short-term expression of transgenes encoding cytoplasmic and membrane anchored proteins in postmitotic polarized cortical neurons and glial cells in vitro.

We constructed three recombinant vectors derived from the herpes simplex virus type 1 mutant tsK, each of which contained a different transgene under the control of the herpes simplex virus type 1 immediate early 3 promoter inserted into the thymidine kinase locus: the prokaryotic enzymes beta-galactosidase and chloramphenicol acetyl transferase, and a fusion gene consisting of human tissue inhibitor of metalloproteinases linked to the last exon of Thy-1, which encodes for a glycosyl-phosphatidyl-inositol membrane anchor. Infection of postmitotic neocortical and hippocampal neurons in low-density primary cultures with these vectors, achieved reliable expression of all three foreign gene products in various neocortical cell types, e.g. pyramidal neurons, non-pyramidal neurons, and glial cells. The percentage of neurons expressing transgenes ranged from 1 to 46% depending on the multiplicity of infection (highest assayed = 5); the percentage of glial cells expressing transgenes ranged from 0.5 to 98% (highest multiplicity assayed = 3.4). Expression of transgenes could be detected for up to three days in approximately 20% of neurons infected at a multiplicity of infection of 1. Infection of neurons with tk K-derived recombinant vectors inhibited their protein synthesis by 40-50% at a multiplicity of infection of 10, but no effect was observed at a multiplicity of infection of 1. Infection of glial cells with the same vectors at a multiplicity of infection of 1 inhibited protein synthesis by more than 90%. Analysis of neuronal viability at different times post-infection indicated that more than 98% of neurons expressing transgenes 48 h post-infection were viable. Thus, low-density neuronal cultures can be used to assess the efficiency of herpes simplex virus type 1-derived gene transfer vectors and transgene expression in developing cortical postmitotic cells, before and after they establish polarity. In addition, we show that two cytoplasmic enzymes, beta-galactosidase and chloramphenicol acetyl transferase, are able to diffuse freely in the cytoplasm reaching even growth cones in young neurons, while the chimeric protein tissue inhibitor of metalloproteinases/Thy-1 is correctly targeted to the plasma membrane via a glycosyl-phosphatidylinositol anchor. This model system should be useful for investigation of cellular and molecular aspects of the development and establishment of neuronal polarity, as well as for analysis of signals involved in protein targeting in postmitotic neurons.

A rapid and simple method for determining the DNA sequences of fragments inserted into vaccinia virus.

Recombinant virus vectors such as vaccinia virus, adenovirus and herpesvirus are frequently used to express a variety of foreign products. A rapid method allowing the precise identification of recombinants would be useful to confirm the nature of a newly produced recombinant and, in particular, to discriminate between recombinants bearing near-identical foreign products. Using vaccinia virus, we describe a method that in one day provides sequence analysis of the recombinant viral DNA.

Identification of two protein binding sites within the varicella-zoster virus major immediate early gene promoter.

Binding sites for cellular proteins in the promoter of the varicella-zoster virus (VZV) major immediate early (IE) gene were investigated. Protein binding was detected at sequence motifs possessing homology to the CCAAT element and an ATF/AP-1-like binding site, and recognition of the ATF/AP-1 site was apparently facilitated by occupation of the CCAAT site. Gene expression directed by the VZV major IE promoter was stimulated by the adenovirus 5, 289 amino acid EIA gene product. The implications of the results for VZV gene expression and replication are discussed.

Chromite ore processing residue in Hudson County, New Jersey.

Chromite ore processing residue occurs at over 130 sites in Hudson County, New Jersey. Many of these sites are in urban residential areas. This waste is a result of 70 years of chromate and bichromate chemical manufacturing. At least 15% of the sites contain total chromium concentrations greater than 10,000 mg/kg, with hexavalent content ranging from about 1 to 50%. Continuing leaching of this waste results in yellow-colored surface water runoff and yellow deposits on the soil surface and inside basement walls. The chemistry, environmental fate, health effects, and human exposure potentials for this waste are described.

Internal quality assurance in a clinical virology laboratory. I. Internal quality assessment.

AIMS: In April 1991 an internal quality assessment scheme (IQAS) was introduced into the virology section of the Clinical Microbiology and Public Health Laboratory, Cambridge. The IQAS was established to identify recurring technical and procedural problems, to check the adequacy of current techniques, and to calculate the frequency of errors. METHODS: Between April 1991 and December 1993, 715 anonymous clinical serum samples were submitted to the laboratory to test 3245 individual procedures of diagnostic viral serology. RESULTS: A total of 485 (14.9%) procedural and 61 (1.9%) technical discrepancies were observed, the technical discrepancies mainly being recorded in complement fixation tests. Twenty two (0.7% of total procedures) of the technical discrepancies were diagnostically significant. CONCLUSIONS: Evaluation criteria developed with the introduction of IQAS to viral serology, and technical and procedural discrepancies are assessed. As yet, IQAS has not been introduced to other sections of the diagnostic virology laboratory (virus isolation, electron microscopy, immunofluorescence, and enzyme linked immunosorbent assays for viral and chlamydial antigens).

Internal quality assurance in a clinical virology laboratory. II. Internal quality control.

AIMS: In April 1991 additional quality control procedures were introduced into the virology section of the Clinical Microbiology and Public Health Laboratory, Cambridge. Internal quality control (IQC) samples were gradually included in the serological assays performed in the laboratory and supplemented kit controls and standard sera. METHODS: From April 1991 to December 1993, 2421 IQC procedures were carried out with reference sera. RESULTS: The IQC samples were evaluated according to the Westgard rules. Violations were recorded in 60 of 1808 (3.3%) controls and were highest in the IQC samples of complement fixation tests (25/312 (8%) of controls submitted for complement fixation tests). CONCLUSIONS: The inclusion of IQC samples in the serological assays performed in the laboratory has highlighted batch to batch variation in commercial assays. The setting of acceptable limits for the IQC samples has increased confidence in the validity of assay results.

The development of an antigen capture polymerase chain reaction assay to detect and type human enteroviruses.

Antigen capture polymerase chain reaction (AC-PCR) is a technique that combines the advantages of PCR with those of antibody mediated methods, to detect and type human enteroviruses. Virus particles are captured by specific antisera and RNA is released by heat denaturation to generate the substrate for reverse transcription and PCR. Use of this technique results in purification of human enteroviruses from tissue culture and 10% faecal samples in a serotype-specific manner allowing both rapid detection and a direct correlation between serological and genetic typing methods. The sensitivity of AC-PCR was comparable with that of PCR protocols employing a conventional organic solvent based extraction procedure.

Risks for transmission of hepatitis C virus during artificial insemination.

OBJECTIVE: To identify risks of hepatitis C virus transmission by semen from infected donors. DESIGN: Case report. SETTING: Assisted fertility clinic. PATIENTS: Hepatitis C virus-infected semen donor and recipients of his donations. INTERVENTION: Testing for hepatitis C virus by serology and polymerase chain reaction. MAIN OUTCOME MEASURES: Detection of hepatitis C virus antibodies and viral RNA. RESULTS: Hepatitis C virus RNA was detected in the semen donation before but not after purification; none of the recipients of the donors samples were found to have antibodies to hepatitis C virus. CONCLUSIONS: Hepatitis C virus RNA can be detected in semen donations from infected donors; purification of donations before insemination significantly reduces the amount of viral RNA in the semen pellet.