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Background: Mycoplasma genitalium infection can adversely affect female reproductive health, but data are limited about prevalence and characteristics of the infection in female adolescents. We employed a sensitive assay to detect M. genitalium infection, and we describe its characteristics in a clinical sample of women younger than 21 years., Methods: We recruited females aged 13 to 20 years in children's hospital clinics whose clinicians were testing for chlamydia/gonorrhea. Participants completed a questionnaire providing demographics, sexual history, and current symptoms. Urine/endocervical samples were tested for chlamydia/gonorrhea and partitioned for M. genitalium testing using Aptima M. genitalium assay. We reviewed records for the clinic visit to document examination, diagnosis, and results of sexually transmitted infection (STI) testing. We compared prevalence of M. genitalium infection by demographics, sexual history, symptoms, and signs., Results: Of 153 participants mean age 18.07 ± 1.68 years, 58% self-identified as Hispanic, 27% Black, 64% straight/heterosexual, 27% bisexual, 1% gay/lesbian, 29% reported a prior STI diagnosis. Prevalence of M. genitalium was 11.1% (17/153), 13 of 17 were asymptomatic, 2 of 17 had pelvic inflammatory disease (PID), 3 of 17 coinfected with chlamydia or gonorrhea. Prevalence of chlamydia was 6.6% and of gonorrhea 2.6%. A logistic regression model indicated independent associations of bisexual orientation versus all other orientations (adjusted odds ratio [aOR], 4.80; 95% confidence interval [CI], 1.38-16.67), self-reported prior STI (aOR, 3.83; 95% CI, 1.10-13.37), and self-reported prior PID (aOR, 9.12; 95% CI, 1.02-81.72) with higher odds of M. genitalium infection., Conclusions: Findings suggest that in at-risk female populations younger than 21 years, M. genitalium is a prevalent STI and symptomatic adolescents may warrant testing and treatment. Further study of harms and benefits of testing asymptomatic bisexual female adolescents or those with prior STI/PID is needed., Competing Interests: Conflict of Interest and Sources of Funding: None declared., (Copyright © 2023 American Sexually Transmitted Diseases Association. All rights reserved.)

With an almost unremittent progression of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections all around the world, there is a compelling need to introduce rapid, reliable, and high-throughput testing to allow appropriate clinical management and/or timely isolation of infected individuals. Although nucleic acid amplification testing (NAAT) remains the gold standard for detecting and theoretically quantifying SARS-CoV-2 mRNA in various specimen types, antigen assays may be considered a suitable alternative, under specific circumstances. Rapid antigen tests are meant to detect viral antigen proteins in biological specimens (e.g. nasal, nasopharyngeal, saliva), to indicate current SARS-CoV-2 infection. The available assay methodology includes rapid chromatographic immunoassays, used at the point-of-care, which carries some advantages and drawbacks compared to more conventional, instrumentation-based, laboratory immunoassays. Therefore, this document by the International Federation for Clinical Chemistry and Laboratory Medicine (IFCC) Taskforce on COVID-19 aims to summarize available data on the performance of currently available SARS-CoV-2 antigen rapid detection tests (Ag-RDTs), providing interim guidance on clinical indications and target populations, assay selection, and evaluation, test interpretation and limitations, as well as on pre-analytical considerations. This document is hence mainly aimed to assist laboratory and regulated health professionals in selecting, validating, and implementing regulatory approved Ag-RDTs., (© 2021 Walter de Gruyter GmbH, Berlin/Boston.)

Serological testing for the detection of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is emerging as an important component of the clinical management of patients with coronavirus disease 2019 (COVID-19) as well as the epidemiological assessment of SARS-CoV-2 exposure worldwide. In addition to molecular testing for the detection of SARS-CoV-2 infection, clinical laboratories have also needed to increase testing capacity to include serological evaluation of patients with suspected or known COVID-19. While regulatory approved serological immunoassays are now widely available from diagnostic manufacturers globally, there is significant debate regarding the clinical utility of these tests, as well as their clinical and analytical performance requirements prior to application. This document by the International Federation for Clinical Chemistry and Laboratory Medicine (IFCC) Taskforce on COVID-19 provides interim guidance on: (A) clinical indications and target populations, (B) assay selection, (C) assay evaluation, and (D) test interpretation and limitations for serological testing of antibodies against SARS-CoV-2 infection. These evidence-based recommendations will provide practical guidance to clinical laboratories in the selection, verification, and implementation of serological assays and are of the utmost importance as we expand our pandemic response from initial case tracing and containment to mitigation strategies to minimize resurgence and further morbidity and mortality.

Routine biochemical and hematological tests have been reported to be useful in the stratification and prognostication of pediatric and adult patients with diagnosed coronavirus disease (COVID-19), correlating with poor outcomes such as the need for mechanical ventilation or intensive care, progression to multisystem organ failure, and/or death. While these tests are already well established in most clinical laboratories, there is still debate regarding their clinical value in the management of COVID-19, particularly in pediatrics, as well as the value of composite clinical risk scores in COVID-19 prognostication. This document by the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) Task Force on COVID-19 provides interim guidance on: (A) clinical indications for testing, (B) recommendations for test selection and interpretation, (C) considerations in test interpretation, and (D) current limitations of biochemical/hematological monitoring of COVID-19 patients. These evidence-based recommendations will provide practical guidance to clinical laboratories worldwide, underscoring the contribution of biochemical and hematological testing to our collective pandemic response.

The diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection globally has relied extensively on molecular testing, contributing vitally to case identification, isolation, contact tracing, and rationalization of infection control measures during the coronavirus disease 2019 (COVID-19) pandemic. Clinical laboratories have thus needed to verify newly developed molecular tests and increase testing capacity at an unprecedented rate. As the COVID-19 pandemic continues to pose a global health threat, laboratories continue to encounter challenges in the selection, verification, and interpretation of these tests. This document by the International Federation for Clinical Chemistry and Laboratory Medicine (IFCC) Task Force on COVID-19 provides interim guidance on: (A) clinical indications and target populations, (B) assay selection, (C) assay verification, and (D) test interpretation and limitations for molecular testing of SARS-CoV-2 infection. These evidence-based recommendations will provide practical guidance to clinical laboratories worldwide and highlight the continued importance of laboratory medicine in our collective pandemic response.

The incidence of melanoma has continued to increase over the past 30 years. Hence, developing effective therapies to treat both primary and metastatic melanoma are essential. While advances in targeted therapy and immunotherapy have provided novel therapeutic options to treat melanoma, gene therapy may provide additional strategies for the treatment of metastatic melanoma clinically. This review focuses upon the challenges and opportunities that gene therapy provides for targeting melanoma. We begin with a discussion of the various gene therapy targets which are relevant to melanoma. Next, we explore the gene therapy clinical trials that have been conducted for treating melanoma. Finally, challenges faced in gene therapy as well as combination therapies for targeting melanoma, which may circumvent these obstacles, will be discussed. Targeted combination gene therapy strategies hold significant promise for developing the most effective therapeutic outcomes, while reducing the toxicity to noncancerous cells, and would integrate the patient's immune system to diminish melanoma progression. Next-generation vectors designed to embody required safety profiles and "theranostic" attributes, combined with immunotherapeutic strategies would be critical in achieving beneficial management and therapeutic outcomes in melanoma patients., (© 2018 Elsevier Inc. All rights reserved.)

Subtraction hybridization identified genes displaying differential expression as metastatic human melanoma cells terminally differentiated and lost tumorigenic properties by treatment with recombinant fibroblast interferon and mezerein. This approach permitted cloning of multiple genes displaying enhanced expression when melanoma cells terminally differentiated, called melanoma differentiation associated (mda) genes. One mda gene, mda-7, has risen to the top of the list based on its relevance to cancer and now inflammation and other pathological states, which based on presence of a secretory sequence, chromosomal location, and an IL-10 signature motif has been named interleukin-24 (MDA-7/IL-24). Discovered in the early 1990s, MDA-7/IL-24 has proven to be a potent, near ubiquitous cancer suppressor gene capable of inducing cancer cell death through apoptosis and toxic autophagy in cancer cells in vitro and in preclinical animal models in vivo. In addition, MDA-7/IL-24 embodied profound anticancer activity in a Phase I/II clinical trial following direct injection with an adenovirus (Ad.mda-7; INGN-241) in tumors in patients with advanced cancers. In multiple independent studies, MDA-7/IL-24 has been implicated in many pathological states involving inflammation and may play a role in inflammatory bowel disease, psoriasis, cardiovascular disease, rheumatoid arthritis, tuberculosis, and viral infection. This review provides an up-to-date review on the multifunctional gene mda-7/IL-24, which may hold potential for the therapy of not only cancer, but also other pathological states., (© 2018 Elsevier Inc. All rights reserved.)

The External Quality Assessment (EQA) in Brazil is performed by the National Health Ministry for diseases that are under supervision of Public Health Department. In addition to the government program, the Brazilian Society of Clinical Analysis and the Brazilian Society of Medical Pathology are allowed to provide their programs under the Supervision of National Agency for Sanitary Surveillance (ANVISA) that regulates laboratories to perform EQA programs., Competing Interests: None declared.

Epithelial-mesenchymal transition (EMT) is one of the decisive steps regulating cancer invasion and metastasis. However, the molecular mechanisms underlying this transition require further clarification. MDA-9/syntenin (SDCBP) expression is elevated in breast cancer patient samples as well as cultured breast cancer cells. Silencing expression of MDA-9 in mesenchymal metastatic breast cancer cells triggered a change in cell morphology in both 2D- and 3D-cultures to a more epithelial-like phenotype, along with changes in EMT markers, cytoskeletal rearrangement and decreased invasion. Conversely, over expressing MDA-9 in epithelial non-metastatic breast cancer cells instigated a change in morphology to a more mesenchymal phenotype with corresponding changes in EMT markers, cytoskeletal rearrangement and an increase in invasion. We also found that MDA-9 upregulated active levels of known modulators of EMT, the small GTPases RhoA and Cdc42, via TGFβ1. Reintroducing TGFβ1 in MDA-9 silenced cells restored active RhoA and cdc42 levels, modulated cytoskeletal rearrangement and increased invasion. We further determined that MDA-9 interacts with TGFβ1 via its PDZ1 domain. Finally, in vivo studies demonstrated that silencing the expression of MDA-9 resulted in decreased lung metastasis and TGFβ1 re-expression partially restored lung metastases. Our findings provide evidence for the relevance of MDA-9 in mediating EMT in breast cancer and support the potential of MDA-9 as a therapeutic target against metastatic disease.

Abrus agglutinin (AGG), a plant lectin isolated from the seeds of Abrus precatorius, has documented antitumor and immunostimulatory effects in murine models. To examine possible antitumor activity against breast cancer, we established human breast tumor xenografts in athymic nude mice and intraperitoneally administered AGG. AGG inhibited tumor growth and angiogenesis as confirmed by monitoring the expression of Ki-67 and CD-31, respectively. In addition, TUNEL positive cells increased in breast tumors treated with AGG suggesting that AGG mediates anti-tumorigenic activity through induction of apoptosis and inhibition of angiogenesis. On a molecular level, AGG caused extrinsic apoptosis through ROS generation that was AKT-dependent in breast cancer cells, without affecting primary mammary epithelial cells, suggesting potential cancer specificity of this natural compound. In addition, using HUVECs, AGG inhibited expression of the pro-angiogenic factor IGFBP-2 in an AKT-dependent manner, reducing angiogenic phenotypes both in vitro and in vivo. Overall, the present results establish that AGG promotes both apoptosis and anti-angiogenic activities in human breast tumor cells, which might be exploited for treatment of breast and other cancers., (© 2016 UICC.)

Background: Prognosis in pulmonary hypertension (PH) is related to right ventricular (RV) function. Quantification of RV mechanics may offer additive value. The objective of our study is to determine the feasibility and clinical and prognostic value of RV strain analysis by cardiovascular magnetic resonance (CMR) based feature tracking (FT) in PH., Methods: We retrospectively enrolled 116 patients (age 52.2 ± 12 years, 73.6 % women) referred to CMR for PH evaluation who underwent right heart catheterization within 1 month. Using dedicated FT software, peak global longitudinal and circumferential RV strain and strain rates (GLS, GCS, GLSR, and GCSR, respectively) were quantified from standard cine images. Using multivariate regression analysis, we evaluated the associations of strain with a composite endpoint of death, lung transplantation, or functional class deterioration., Results: RV strain analysis was feasible in 110 (95 %) patients. Patients were classified into: Group A (no PH, normal right ventricular ejection fraction [RVEF]; n = 17), Group B (PH, normal RVEF; n = 26), or Group C (PH, abnormal RVEF; n = 67). All strain and strain rate values were reduced in Group C. Furthermore, GCSR was significantly reduced in Group B (-0.92 [-1.0/-0.7]; p < 0.001) compared to Group A (-1.12 [-1.3/-0.9]; p < 0.001). After adjustment for six clinically meaningful covariates, GLS (hazard ratio 1.06; p = 0.026), GLSR (hazard ratio 2.52; p = 0.04), and GCSR (hazard ratio 4.5; p = 0.01) were independently associated with the composite endpoint. GCSR successfully discriminated patients with and without events (p = 0.01)., Conclusions: Quantification of RV strain with CMR-FT is feasible in the majority of patients, correlates with disease severity, and is independently associated with poor outcomes in PH.

The expression of the tumor suppressor Merlin is compromised in nervous system malignancies due to genomic aberrations. We demonstrated for the first time, that in breast cancer, Merlin protein expression is lost due to proteasome-mediated elimination. Immunohistochemical analysis of tumor tissues from patients with metastatic breast cancer revealed characteristically reduced Merlin expression. Importantly, we identified a functional role for Merlin in impeding breast tumor xenograft growth and reducing invasive characteristics. We sought to determine a possible mechanism by which Merlin accomplishes this reduction in malignant activity. We observed that breast and pancreatic cancer cells with loss of Merlin show an aberrant increase in the activity of β-catenin concomitant with nuclear localization of β-catenin. We discovered that Merlin physically interacts with β-catenin, alters the sub-cellular localization of β-catenin, and significantly reduces the protein levels of β-catenin by targeting it for degradation through the upregulation of Axin1. Consequently, restoration of Merlin inhibited β-catenin-mediated transcriptional activity in breast and pancreatic cancer cells. We also present evidence that loss of Merlin sensitizes tumor cells to inhibition by compounds that target β-catenin-mediated activity. Thus, this study provides compelling evidence that Merlin reduces the malignant activity of pancreatic and breast cancer, in part by suppressing the Wnt/β-catenin pathway. Given the potent role of Wnt/β-catenin signaling in breast and pancreatic cancer and the flurry of activity to test β-catenin inhibitors in the clinic, our findings are opportune and provide evidence for Merlin in restraining aberrant activation of Wnt/β-catenin signaling.

Metastasis is the complex process by which primary tumor cells migrate and establish secondary tumors in an adjacent or distant location in the body. Early detection of metastatic disease and effective therapeutic options for targeting these detected metastases remain impediments to effectively treating patients with advanced cancers. If metastatic lesions are identified early, patients might maximally benefit from effective early therapeutic interventions. Further, monitoring patients whose primary tumors are effectively treated for potential metastatic disease onset is also highly valuable. Finally, patients with metastatic disease can be monitored for efficacy of specific therapeutic interventions through effective metastatic detection techniques. Thus, being able to detect and visualize metastatic lesions is key and provides potential to greatly improve overall patient outcomes. In order to achieve these objectives, researchers have endeavored to mechanistically define the steps involved in the metastatic process as well as ways to effectively detect metastatic progression. We presently overview various preclinical and clinical in vitro and in vivo assays developed to more efficiently detect tumor metastases, which provides the foundation for developing more effective therapies for this invariably fatal component of the cancerous process., Competing Interests: Drs. M.G.P. and P.B.F. are cofounders of, serve as consultants to and have ownership interest in CTS, Inc. Dr. M.G.P. is a member of the board of directors of CTS, Inc., Johns Hopkins University, Virginia Commonwealth University, and Columbia University have ownership interest in CTS, Inc., (© 2016 Elsevier Inc. All rights reserved.)

Melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) encodes a tumor suppressor gene implicated in the growth of various tumor types including breast cancer. We previously demonstrated that recombinant adenovirus-mediated mda-7/IL-24 expression in the mammary glands of carcinogen-treated (methylnitrosourea, MNU) rats suppressed mammary tumor development. Since most MNU-induced tumors in rats contain activating mutations in Ha-ras, which arenot frequently detected in humans, we presently examined the effect of MDA-7/IL-24 on Her2/Neu-induced mammary tumors, in which the RAS pathway is induced. We generated tet-inducible MDA-7/IL-24 transgenic mice and crossed them with Her2/Neu transgenic mice. Triple compound transgenic mice treated with doxycycline exhibited a strong inhibition of tumor development, demonstrating tumor suppressor activity by MDA-7/IL-24 in immune-competent mice. MDA-7/IL-24 induction also inhibited growth of tumors generated following injection of Her2/Neu tumor cells isolated from triple compound transgenic mice that had not been treated with doxycycline, into the mammary fat pads of isogenic FVB mice. Despite initial growth suppression, tumors in triple compound transgenic mice lost mda-7/IL-24 expression and grew, albeit after longer latency, indicating that continuous presence of this cytokine within tumor microenvironment is crucial to sustain tumor inhibitory activity. Mechanistically, MDA-7/IL-24 exerted its tumor suppression effect on HER2+ breast cancer cells, at least in part, through PERP, a member of PMP-22 family with growth arrest and apoptosis-inducing capacity. Overall, our results establish mda-7/IL-24 as a suppressor of mammary tumor development and provide a rationale for using this cytokine in the prevention/treatment of human breast cancer.

Melanoma differentiation associated gene-7/Interleukin-24 (MDA-7/IL-24) is a novel member of the IL-10 gene family that selectively induces apoptosis and toxic autophagy in a broad spectrum of human cancers, including breast cancer, without harming normal cells or tissues. The ability to investigate the critical events underlying cancer initiation and progression, as well as the capacity to test the efficacy of novel therapeutics, has been significantly advanced by the development of genetically engineered mice (GEMs) that accurately recapitulate specific human cancers. We utilized three transgenic mouse models to better comprehend the in vivo role of MDA-7/IL-24 in breast cancer. Using the MMTV-PyMT spontaneous mammary tumor model, we confirmed that exogenously introducing MDA-7/IL-24 using a Cancer Terminator Virus caused a reduction in tumor burden and also produced an antitumor "bystander" effect. Next we performed xenograft studies in a newly created MMTV-MDA-7 transgenic model that over-expresses MDA-7/IL-24 in the mammary glands during pregnancy and lactation, and found that MDA-7/IL-24 overexpression delayed tumor growth following orthotopic injection of a murine PDX tumor cell line (mPDX) derived from a tumor formed in an MMTV-PyMT mouse. We also crossed the MMTV-MDA-7 line to MMTV-Erbb2 transgenic mice and found that MDA-7/IL-24 overexpression delayed the onset of mammary tumor development in this model of spontaneous mammary tumorigenesis as well. Finally, we assessed the role of MDA-7/IL-24 in immune regulation, which can potentially contribute to tumor suppression in vivo. Our findings provide further direct in vivo evidence for the role of MDA-7/IL-24 in tumor suppression in breast cancer in immune-competent transgenic mice.

Aims: This study was designed to assess the accuracy of coronary artery calcium scans (CACS) acquired at radiation doses below mammography and low-dose lung scanning, compared with standard-dose CACS., Methods and Results: CACS was performed in 102 consecutive patients at 120 kVp; all were imaged at standard-dose mAs levels ranging from 30 to 80 mAs determined by their weight, with iterative reconstruction (IR) level 3, and at 50% of the standard-dose mAs with IR level 7 to compensate for the expected increased noise with lower mAs. The low- vs. standard-dose mAs was 24.5 ± 8.8 vs. 48.5 ± 17.8 mAs (P < 0.0001), and the radiation exposure was 0.37 ± 0.16 vs. 0.76 + 0.34 mSv (P < 0.0001). The Agatston score correlation between the low and high dose was excellent (r = 0.998, P < 0.0001) over a range of scores from 0 to 2512. The weighted kappa for agreement of standard CAC risk categories was 0.95 (95% CI 0.83-0.97). The mean of the differences between individual low- and standard-dose Agatston scores was 17.4 ± 25.8, lower than the reported variability of two scans performed with the same mAs., Conclusion: There was excellent agreement of CACS-based risk classification at low and standard doses, with lower interscan variability than with reported identical doses. The low-dose CACS radiation exposure was less than the approved screening tools of mammography and low-dose lung scanning., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.)

Background: Malignant glioma is an aggressive cancer requiring new therapeutic targets. MicroRNAs (miRNAs) regulate gene expression post transcriptionally and are implicated in cancer development and progression. Deregulated expressions of several miRNAs, specifically hsa-miR-184, correlate with glioma development., Methods: Bioinformatic approaches were used to identify potential miR-184-regulated target genes involved in malignant glioma progression. This strategy identified a multifunctional nuclease, SND1, known to be overexpressed in multiple cancers, including breast, colon, and hepatocellular carcinoma, as a putative direct miR-184 target gene. SND1 levels were evaluated in patient tumor samples and human-derived cell lines. We analyzed invasion and signaling in vitro through SND1 gain-of-function and loss-of-function. An orthotopic xenograft model with primary glioma cells demonstrated a role of miR-184/SND1 in glioma pathogenesis in vivo., Results: SND1 is highly expressed in human glioma tissue and inversely correlated with miR-184 expression. Transfection of glioma cells with a miR-184 mimic inhibited invasion, suppressed colony formation, and reduced anchorage-independent growth in soft agar. Similar phenotypes were evident when SND1 was knocked down with siRNA. Additionally, knockdown (KD) of SND1 induced senescence and improved the chemoresistant properties of malignant glioma cells. In an orthotopic xenograft model, KD of SND1 or transfection with a miR-184 mimic induced a less invasive tumor phenotype and significantly improved survival of tumor bearing mice., Conclusions: Our study is the first to show a novel regulatory role of SND1, a direct target of miR-184, in glioma progression, suggesting that the miR-184/SND1 axis may be a useful diagnostic and therapeutic tool for malignant glioma., (© The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: The SYNTAX score helps in the treatment decision in multivessel coronary disease. Coronary computed tomography angiography (CTA) can measure the SYNTAX score but has been used in few patients with multivessel disease. Our aim was to assess the feasibility, accuracy and reproducibility of SYNTAX score with CCTA compared with invasive coronary angiography (ICA) in de novo left main and/or 3-vessel disease., Methods: 57 patients with new left main and/or 3-vessel disease on ICA and a CCTA performed within the previous month were included. The SYNTAX score was calculated retrospectively for both modalities. Agreement for the global score, vessel score, different components and inter-readers was evaluated with intraclass correlation coefficient (ICC). The ability to classify SYNTAX score categories (low, intermediate and high) was assessed using weighted kappa (K) coefficient., Results: CCTA-based SYNTAX score showed an acceptable concordance (ICC=0.64) and good correlation (r=0.65, p<0.001) with ICA. ICC per vessel and component ranged from 0 to 0.73. There was good agreement classifying the SYNTAX score categories (K=0.53) and interobserver reproducibility (ICC=0.85). CCTA demonstrated high diagnostic accuracy (0.84) for detecting patients in the high score group. No patient with a high CCTA SYNTAX score had a low risk score by ICA that would suggest benefit from percutaneous revascularization., Conclusions: CCTA showed good correlation, acceptable concordance, and high reproducibility for the quantification of the SYNTAX score in de novo left main and/or 3-vessel coronary disease. A high CCTA SYNTAX score identified a group of patients less likely to benefit from percutaneous coronary intervention., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Gene therapy, which involves replacement of a defective gene with a functional, healthy copy of that gene, is a potentially beneficial cancer treatment approach particularly over chemotherapy, which often lacks selectivity and can cause non-specific toxicity. Despite significant progress pre-clinically with respect to both enhanced targeting and expression in a tumor-selective manner several hurdles still prevent success in the clinic, including non-specific expression, low-efficiency delivery and biosafety. Various innovative genetic approaches are under development to reconstruct vectors/transgenes to make them safer and more effective. Utilizing cutting-edge delivery technologies, gene expression can now be targeted in a tissue- and organ-specific manner. With these advances, gene therapy is poised to become amenable for routine cancer therapy with potential to elevate this methodology as a first line therapy for neoplastic diseases. This review discusses recent advances in gene therapy and their impact on a pre-clinical and clinical level., (© 2014 Wiley Periodicals, Inc.)

Introduction: Melanoma differentiation-associated gene - 9 (MDA-9)/Syntenin has become an increasingly popular focus for investigation in numerous cancertypes. Originally implicated in melanoma metastasis, it has diverse cellular roles and is consistently identified as a regulator of tumor invasion and angiogenesis. As a potential target for inhibiting some of the most lethal aspects of cancer progression, further insight into the function of MDA-9/Syntenin is mandatory., Areas Covered: Recent literature and seminal articles were reviewed to summarize the latest collective understanding of MDA-9/Syntenin's role in normal and cancerous settings. Insights into its participation in developmental processes are included, as is the functional significance of the N- and C-terminals and PDZ domains of MDA-9/Syntenin. Current reports highlight the clinical significance of MDA-9/Syntenin expression level in a variety of cancers, often correlating directly with reduced patient survival. Also presented are assessments of roles of MDA-9/Syntenin in cancer progression as well as its functions as an intracellular adapter molecule., Expert Opinion: Multiple studies demonstrate the importance of MDA-9/Syntenin in tumor invasion and progression. Through the use of novel drug design approaches, this protein may provide a worthwhile therapeutic target. As many conventional therapies do not address, or even enhance, tumor invasion, an anti-invasive approach would be a worthwhile addition in cancer therapy.

Little is known about the molecular pathways regulating poor differentiation and invasion of head and neck squamous cell carcinoma (HNSCC). In the present study, we aimed to determine the role of MDA-9/Syntenin, a metastasis associated molecule in HNSCC tumorigenesis. Elevated MDA-9/Syntenin expression was evident in 67% (54/81) primary HNSCC tumors (p=0.001-0.002) and 69% (9/13) pre-neoplastic tissues (p=0.02-0.03). MDA-9/Syntenin overexpression was associated with the stage (p=0.001), grade (p=0.001) and lymph node metastasis (p=0.0001). Silencing of MDA-9/Syntenin in 3 poorly differentiated HNSCC cell lines induced squamous epithelial cell differentiation, disrupted angiogenesis and reduced tumor growth in vitro and in vivo. We confirmed SPRR1B and VEGFR1 as the key molecular targets of MDA-9/Syntenin on influencing HNSCC differentiation and angiogenesis respectively. MDA-9/Syntenin disrupted SPRR1B expression interacting through its PDZ1 domain and altered VEGFR1 expression in vitro and in vivo. VEGFR1 co-localized with MDA-9/Syntenin in HNSCC cell lines and primary tumor. Downregulation of growth regulatory molecules CyclinD1, CDK4, STAT3, PI3K and CTNNB1 was also evident in the MDA-9/Syntenin depleted cells, which was reversed following over-expression of MDA-9/Syntenin in immortalized oral epithelial cells. Our results suggest that early induction of MDA-9/Syntenin expression influences HNSCC progression and should be further evaluated for potential biomarker development.

Background: The extraordinary invasiveness of human glioblastoma multiforme (GBM) contributes to treatment failure and the grim prognosis of patients diagnosed with this tumor. Consequently, it is imperative to define further the cellular mechanisms that control GBM invasion and identify promising novel therapeutic targets. Melanoma differentiation associated gene-9 (MDA-9/syntenin) is a highly conserved PDZ domain-containing scaffolding protein that promotes invasion and metastasis in vitro and in vivo in human melanoma models. To determine whether MDA-9/syntenin is a relevant target in GBM, we investigated its expression in tumor samples and involvement in GBM invasion and angiogenesis., Materials: We assessed MDA-9/syntenin levels in available databases, patient tumor samples, and human-derived cell lines. Through gain-of-function and loss-of-function studies, we analyzed changes in invasion, angiogenesis, and signaling in vitro. We used orthotopic xenografts with GBM6 cells to demonstrate the role of MDA-9/syntenin in GBM pathogenesis in vivo., Results: MDA-9/syntenin expression in high-grade astrocytomas is significantly higher than normal tissue counterparts. Forced overexpression of MDA-9/syntenin enhanced Matrigel invasion, while knockdown inhibited invasion, migration, and anchorage-independent growth in soft agar. Moreover, overexpression of MDA-9/syntenin increased activation of c-Src, p38 mitogen-activated protein kinase, and nuclear factor kappa-B, leading to elevated expression of matrix metalloproteinase 2 and secretion of interleukin-8 with corresponding changes observed upon knockdown. GBM6 cells that stably express small hairpin RNA for MDA-9/syntenin formed smaller tumors and had a less invasive phenotype in vivo., Conclusions: Our findings indicate that MDA-9/syntenin is a novel and important mediator of invasion in GBM and a key regulator of pathogenesis, and we identify it as a potential target for anti-invasive treatment in human astrocytoma.

Elucidating the mechanism of pathogenesis of breast cancer has greatly benefited from breakthrough advances in both genetically engineered mouse (GEM) models and xenograft transplantation technologies. The vast array of breast cancer mouse models currently available is testimony to the complexity of mammary tumorigenesis and attempts by investigators to accurately portray the heterogeneity and intricacies of this disease. Distinct molecular changes that drive various aspects of tumorigenesis, such as alterations in tumor cell proliferation and apoptosis, invasion and metastasis, angiogenesis, and drug resistance have been evaluated using the currently available GEM breast cancer models. GEM breast cancer models are also being exploited to evaluate and validate the efficacy of novel therapeutics, vaccines, and imaging modalities for potential use in the clinic. This review provides a synopsis of the various GEM models that are expanding our knowledge of the nuances of breast cancer development and progression and can be instrumental in the development of novel prevention and therapeutic approaches for this disease., (© 2014 Elsevier Inc. All rights reserved.)

There is accumulative evidence on the multiple functions of the intestinal microflora in relation to the homeostasis of the host. At first considered as a simple mutualism, today this relationship proves to be essential to the health and to pathologic processes, particularly metabolic (eg, obesity) and gastrointestinal (eg, inflammatory bowel disease and functional disorders). The first studies were conducted on the microbiota from fecal material cultured anaerobically. With the advent of molecular biology, it has become possible to determine qualitative and quantitatively the dominant, subdominant and transients species. In recent years, there were advances in the understanding of the relationship betwen the microbiota and the host, as well as among the microorganisms in their respective niches. These advances result from translational integration of microbiology with specialities such as molecular biology, cell phisiology, immunology and ecology. There are few studies on the spatial distribution of the microflora in the gut. Unravelling the topography of the microflora in mammals is a way to validate new animal models for the study of microflora.

Molecular-genetic imaging of cancer using nonviral delivery systems has great potential for clinical application as a safe, efficient, noninvasive tool for visualization of various cellular processes including detection of cancer, and its attendant metastases. In recent years, significant effort has been expended in overcoming technical hurdles to enable clinical adoption of molecular-genetic imaging. This chapter will provide an introduction to the components of molecular-genetic imaging and recent advances on each component leading to safe, efficient clinical applications for detecting cancer. Combination with therapy, namely, generating molecular-genetic theranostic constructs, will provide further impetus for clinical translation of this promising technology., (© 2014 Elsevier Inc. All rights reserved.)

First identified almost two decades ago as a novel gene differentially expressed in human melanoma cells induced to terminally differentiate, MDA-7/IL-24 has since shown great potential as an anti-cancer gene. MDA-7/IL24, a secreted protein of the IL-10 family, functions as a cytokine at normal physiological levels and is expressed in tissues of the immune system. At supra-physiological levels, MDA-7/IL-24 plays a prominent role in inhibiting tumor growth, invasion, metastasis and angiogenesis and was recently shown to target tumor stem/initiating cells for death. Much of the attention focused on MDA-7/IL-24 originated from the fact that it can selectively induce cell death in cancer cells without affecting normal cells. Thus, this gene originally shown to be associated with melanoma cell differentiation has now proven to be a multi-functional protein affecting a broad array of cancers. Moreover, MDA-7/IL-24 has proven efficacious in a Phase I/II clinical trial in humans with multiple advanced cancers. As research in the field progresses, we will unravel more of the functions of MDA-7/IL-24 and define novel ways to utilize MDA-7/IL-24 in the treatment of cancer.

Melanoma differentiation-associated gene-7/interleukin-24 (mda-7/IL-24) displays a broad range of antitumor properties including cancer-specific induction of apoptosis, inhibition of tumor angiogenesis and modulation of antitumor immune responses. In our study, we elucidated the role of MDA-7/IL-24 in inhibiting growth of breast cancer-initiating/stem cells. Ad.mda-7 infection decreased proliferation of breast cancer-initiating/stem cells without affecting normal breast stem cells. Ad.mda-7 induced apoptosis and endoplasmic reticulum stress in breast cancer-initiating/stem cells similar to unsorted breast cancer cells and inhibited the self-renewal property of breast cancer-initiating/stem cells by suppressing Wnt/β-catenin signaling. Prevention of inhibition of Wnt signaling by LiCl increased cell survival upon Ad.mda-7 treatment, suggesting that Wnt signaling inhibition might play a key role in MDA-7/IL-24-mediated death of breast cancer-initiating/stem cells. In a nude mouse subcutaneous xenograft model, Ad.mda-7 injection profoundly inhibited growth of tumors generated from breast cancer-initiating/stem cells and also exerted a potent "bystander" activity inhibiting growth of distant uninjected tumors. Further studies revealed that tumor growth inhibition by Ad.mda-7 was associated with a decrease in proliferation and angiogenesis, two intrinsic features of MDA-7/IL-24, and a reduction in vivo in the percentage of breast cancer-initiating/stem cells. Our findings demonstrate that MDA-7/IL-24 is not only nontoxic to normal cells and normal stem cells but also can kill both unsorted cancer cells and enriched populations of cancer-initiating/stem cells, providing further documentation that MDA-7/IL-24 might be a safe and effective way to eradicate cancers and also potentially establish disease-free survival., (Copyright © 2013 UICC.)

Purpose: Urothelial cell carcinoma (UCC) rapidly progresses from superficial to muscle-invasive tumors. The key molecules involved in metastatic progression and its early detection require clarification. The present study defines a seminal role of the metastasis-associated gene MDA-9/Syntenin in UCC progression., Experimental Design: Expression pattern of MDA-9/Syntenin was examined in 44 primary UCC and the impact of its overexpression and knockdown was examined in multiple cells lines and key findings were validated in primary tumors., Results: Significantly higher (P=0.002-0.003) expression of MDA-9/Syntenin was observed in 64% (28 of 44) of primary tumors and an association was evident with stage (P=0.01), grade (P=0.03), and invasion status (P=0.02). MDA-9/Syntenin overexpression in nontumorigenic HUC-1 cells increased proliferation (P=0.0012), invasion (P=0.0001), and EGF receptor (EGFR), AKT, phosphoinositide 3-kinase (PI3K), and c-Src expression. Alteration of β-catenin, E-cadherin, vimentin, claudin-1, ZO-1, and T-cell factor-4 (TCF4) expression was also observed. MDA-9/Syntenin knockdown in three UCC cell lines reversed phenotypic and molecular changes observed in the HUC-1 cells and reduced in vivo metastasis. Key molecular changes observed in the cell lines were confirmed in primary tumors. A physical interaction and colocalization of MDA-9/Syntenin and EGFR was evident in UCC cell lines and primary tumors. A logistic regression model analysis revealed a significant correlation between MDA-9/Syntenin:EGFR and MDA-9/Syntenin:AKT expressions with stage (P=0.04, EGFR; P=0.01, AKT). A correlation between MDA-9/Syntenin:β-catenin coexpression with stage (P=0.03) and invasion (P=0.04) was also evident., Conclusions: Our findings indicate that MDA-9/Syntenin might provide an attractive target for developing detection, monitoring, and therapeutic strategies for managing UCC., (©2013 AACR.)

Gene therapy provides a novel platform for therapeutic intervention of several genetic and non-genetic disorders. With the recent developments in the field, a wide variety of viral and non-viral vectors have emerged that can deliver genetic payloads to target cells. However, non-targeted delivery of transgenes often results in undesirable effects, low tumor transduction, and reduced therapeutic index. In this review, we focus on some of the novel approaches that can be used to meet the present challenges in the field and translate the potential of cancer gene therapy from 'bench to bedside' in the near future.

Structure-based modeling combined with rational drug design, and high throughput screening approaches offer significant potential for identifying and developing lead compounds with therapeutic potential. The present review focuses on these two approaches using explicit examples based on specific derivatives of Gossypol generated through rational design and applications of a cancer-specificpromoter derived from Progression Elevated Gene-3. The Gossypol derivative Sabutoclax (BI-97C1) displays potent anti-tumor activity against a diverse spectrum of human tumors. The model of the docked structure of Gossypol bound to Bcl-XL provided a virtual structure-activity-relationship where appropriate modifications were predicted on a rational basis. These structure-based studies led to the isolation of Sabutoclax, an optically pure isomer of Apogossypol displaying superior efficacy and reduced toxicity. These studies illustrate the power of combining structure-based modeling with rational design to predict appropriate derivatives of lead compounds to be empirically tested and evaluated for bioactivity. Another approach to cancer drug discovery utilizes a cancer-specific promoter as readouts of the transformed state. The promoter region of Progression Elevated Gene-3 is such a promoter with cancer-specific activity. The specificity of this promoter has been exploited as a means of constructing cancer terminator viruses that selectively kill cancer cells and as a systemic imaging modality that specifically visualizes in vivo cancer growth with no background from normal tissues. Screening of small molecule inhibitors that suppress the Progression Elevated Gene-3-promoter may provide relevant lead compounds for cancer therapy that can be combined with further structure-based approaches leading to the development of novel compounds for cancer therapy.

Elevated levels of the oncoprotein, osteopontin (OPN), are associated with poor outcome of several types of cancers including melanoma. We have previously reported an important involvement of DNAJB6, a member of heat-shock protein 40 (HSP40) family, in negatively impacting tumor growth. The current study was prompted by our observations reported here which revealed a reciprocal relationship between DNAJB6 and OPN in melanoma specimens. The 'J domain' is the most conserved domain of HSP40 family of proteins. Hence, we assessed the functional role of the J domain in activities of DNAJB6. We report that the J domain of DNAJB6 is involved in mediating OPN suppression. Deletion of the J domain renders DNAJB6 incapable of impeding malignancy and suppressing OPN. Our mechanistic investigations reveal that DNAJB6 binds HSPA8 (heat-shock cognate protein, HSC70) and causes dephosphorylation of glycogen synthase kinase 3β (GSK3β) at Ser 9 by recruiting protein phosphatase, PP2A. This dephosphorylation activates GSK3β, leading to degradation of β-catenin and subsequent loss of TCF/LEF (T cell factor1/lymphoid enhancer factor1) activity. Deletion of the J domain abrogates assembly of this multiprotein complex and renders GSK3β inactive, thus, stabilizing β-catenin, a transcription co-activator for OPN expression. Our in-vitro and in-vivo functional analyses show that silencing OPN expression in the background of deletion of the J domain renders the resultant tumor cells less malignant despite the presence of stabilized β-catenin. Thus, we have uncovered a new mechanism for regulation of GSK3β activity leading to inhibition of Wnt/β-catenin signaling.

DKK1 (dickkopf 1 homologue) is a secreted inhibitor of the Wnt signalling pathway and a critical modulator of tumour promotion and the tumour microenvironment. However, mechanisms regulating DKK1 expression are understudied. DNAJB6 {DnaJ [HSP40 (heat-shock protein 40 kDa)] homologue, subfamily B, member 6} is an HSP40 family member whose expression is compromised during progression of breast cancer and melanoma. Inhibition of the Wnt/β-catenin signalling pathway by up-regulation of DKK1 is one of the key mechanisms by which DNAJB6 suppresses tumour metastasis and EMT (epithelial-mesenchymal transition). Analysis of the DKK1 promoter to define the cis-site responsible for its up-regulation by DNAJB6 revealed the presence of two binding sites for a transcriptional repressor, MSX1 (muscle segment homeobox 1). Our investigations showed that MSX1 binds the DKK1 promoter and inhibits DKK1 transcription. Interestingly, silencing DNAJB6 resulted in up-regulation of MSX1 concomitant with increased stabilization of β-catenin. ChIP (chromatin immunoprecipitation) studies revealed that β-catenin binds the MSX1 promoter and stabilization of β-catenin elevates MSX1 transcription, indicating that β-catenin works as a transcription co-activator for MSX1. Functionally, exogenous expression of MSX1 in DNAJB6-expressing cells promotes the mesenchymal phenotype by suppression of DKK1. Thus we have identified a novel regulatory mechanism of DNAJB6-mediated DKK1 transcriptional up-regulation that can influence EMT. DKK1 is a feedback regulator of β-catenin levels and thus our studies also define an additional negative control of this β-catenin/DKK1 feedback loop by MSX1, which may potentially contribute to excessive stabilization of β-catenin.

HSP40 family member MRJ (DNAJB6) has been in the spot light for its relevance to Huntington's, Parkinson's diseases, limb-girdle muscular dystrophy, placental development, neural stem cells, cell cycle and malignancies such as breast cancer and melanoma. This gene has two spliced variants coding for 2 distinct proteins with significant homology. However, MRJ(L) (large variant) is predominantly localized to the nucleus whereas MRJ(S) (small variant) is predominantly cytoplasmic. Interestingly MRJ(S) translocates to the nucleus in response to heat shock. The classical heat shock proteins respond to crises (stress) by increasing the number of molecules, usually by transcriptional up-regulation. Our studies imply that a quick increase in the molar concentration of MRJ in the nuclear compartment is a novel method by which MRJ responds to stress. We found that MRJ(S) shows NLS (nuclear localization signal) independent nuclear localization in response to heat shock and hypoxia. The specificity of this response is realized due to lack of such response by MRJ(S) when challenged by other stressors, such as some cytokines or UV light. Deletion analysis has allowed us to narrow down on a 20 amino acid stretch at the C-terminal region of MRJ(S) as a potential stress sensing region. Functional studies indicated that constitutive nuclear localization of MRJ(S) promoted attributes of malignancy such as proliferation and invasiveness overall indicating distinct phenotypic characteristics of nuclear MRJ(S)., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Dickkopf1 (DKK1), a secreted inhibitor of the Wnt/β-catenin pathway, is a negative regulator of bone formation. DKK1 acts as a switch that transitions prostate cancer bone metastases from osteolytic to osteoblastic and also is an active indicator of poor outcome for multiple myeloma. However, in other tumor types, DKK1 upregulation or overexpression suppresses tumor growth. Thus, the role of DKK1 in cancer appears to be diverse. This raises a question: Could the increased levels of DKK1 still be tumor protective when observed in high levels in the serum of patients? Here, we summarize the diverse, seemingly contradicting roles of DKK1 and attempt to explain the apparent dichotomy in its activity. We propose that DKK1 is a critical secreted factor that modulates microenvironment. Based on the location and components of the microenvironment DKK1 will support different outcomes., (Copyright © 2011 UICC.)

We showed that expression of MRJ (DNAJB6) protein is lost in invasive ductal carcinoma, and restoration of MRJ(L) restricts malignant behavior of breast cancer and melanoma cells. However, the signaling pathways influenced by MRJ(L) are largely unknown. Our observations revealed that MRJ(L) expression causes changes in cell morphology concomitant with down-regulation of several mesenchymal markers, viz. vimentin, N-cadherin, Twist, and Slug, and up-regulation of epithelial marker keratin 18. Importantly, MRJ(L) expression led to reduced levels of beta-catenin, an epithelial mesenchymal transition marker, and a critical player in the Wnt pathway. We found that MRJ(L) up-regulates expression of DKK1, a well known Wnt/beta-catenin signaling inhibitor, that causes degradation of beta-catenin. Re-expression of DNAJB6 alters the Wnt/beta-catenin signaling in cancer cells, leading to partial reversal of the mesenchymal phenotype. Thus, MRJ(L) may play a role in maintaining an epithelial phenotype, and inhibition of the Wnt/beta-catenin pathway may be one of the potential mechanisms contributing to the restriction of malignant behavior by MRJ(L).

Allele frequencies for 15 short tandem repeats (STR) loci were determined with a sample of 3000 unrelated individuals from the population of Santa Catarina, Southern Brazil. The loci are most commonly used in forensic and paternity testing, being analyzed by the Powerplex 16 (Promega) commercial kit. The data shows that most polymorphic loci were Penta E and FGA. The distributions of the genotypes in the evaluated loci are in Hardy-Weinberg equilibrium. Comparative analyses between our population data and other Brazilian populations are presented. The calculated forensic parameters showed that the loci are useful for the solution of forensic problems in Brazilian Southern region.

Hepatitis C virus (HCV) isolates have been divided into six genotypes (1 to 6). The duration of hepatitis C standard treatment is 48 weeks for patients infected with HCV genotype 1 vs 24 weeks for those infected with genotypes 2 and 3. A total of 1544 HCV isolates from chronic patients living in the southern Brazilian states of Rio Grande do Sul (RS, n=627) and Santa Catarina (SC, n=917) were genotyped by restriction fragment length polymorphism (RFLP) of polymerase chain reaction (PCR) products. In RS, 338 (53.9%; 95% CI 50.0-57.8%), 34 (5.4%; 95% CI 3.8-7.4%) and, 255 (40.7%; 95% CI 36.9-44.6%) samples were from genotypes 1, 2, and 3, respectively. In SC, 468 (51%; 95% CI 47.8-54.2%), 26 (2.9%; 95% CI 1.9-4.1%) and, 423 (46.1%; 95% CI 42.9-49.3%) samples were from genotypes 1, 2, and 3, respectively. Genotyping results were confirmed by direct nucleotide sequencing of PCR products derived from 68 samples, without any discrepancy between PCR-RFLP and nucleotide sequencing methods. In conclusion, almost half of the hepatitis C patients from South of Brazil are infected by genotypes 2 and 3 and, these results have important consequential therapeutic implications as they can be treated for only 24 weeks, not 48.

TT virus (TTV) is a newly described nonenveloped human virus, with a circular, negative-stranded DNA genome, that was first identified in the blood of a patient with posttransfusion hepatitis of unknown etiology. PCR primers and conditions used for TTV DNA amplification may greatly influence the level of TTV detection in serum. Three PCR assays, with different regions of the genome as targets, were used to test TTV DNA in 130 sera from children and adults visiting a hospital in the south of Brazil, most of them for routine procedure. Forty-four percent of adult sera and 73% of sera from children aged 0-10 years were TTV positive with at least one PCR assay. However, the three assays were able to detect only 33%, 35%, and 70% of the total positive samples. Our results showed a high prevalence of TTV infection in the south of Brazil, particularly among young children, and confirmed the necessity of performing several PCR assays to assess the true TTV prevalence in a determined population.

Objective: To report and assess the incidence of cardiac tamponade in systemic lupus erythematosus as a cardiac manifestation of the disease., Methods: We reviewed the medical records of 325 patients diagnosed with systemic lupus erythematosus according to the American Rheumatism Association and their complementary laboratory tests compatible with cardiac tamponade., Results: In the 325 medical records reviewed, we found 108 patients with pericardial effusions corresponding to 33.2% of the total and 54% of the patients studied in the active phase of the disease. Clinical assessment and transthoracic echocardiogram allowed the clinical diagnosis of cardiac tamponade in only 4 (1.23%) patients, 3 of whom were females, white, with ages ranging from 25 to 44 years. The pericardial fluid was hemorrhagic or serosanguineous with high levels of FAN and positivity for LE cells. In the treatment, we successfully used pericardiocentesis associated with high doses of corticosteroids. In clinical and laboratory follow-up performed for a period of 3 years, neither recrudescence of the pericardial effusion nor evolution to constriction occurred., Conclusion: Even though rare (1.23%), cardiac tamponade in patients with systemic lupus erythematosus has a benign evolution when properly treated, according to our experience.

Objective: The intracellular Gram-negative bacterium Chlamydia pneumoniae has been associated with atherosclerosis. The presence of Chlamydia pneumoniae has been investigated in fragments of the arterial wall with a technique for DNA identification., Methods: Arterial fragments obtained from vascular surgical procedures in 58 patients were analyzed. From these patients, 39 were males and the mean age was 65+/-6 years. The polymerase chain reaction was used to identify the bacterial DNA with a pair of primers that codify the major outer membrane protein (MOMP) of Chlamydia pneumoniae. The amplified product was visualized by electrophoresis in the 2% agarose gel stained with ethidium bromide, and it was considered positive when migrating in the band of molecular weight of the positive controls., Results: Seven (12%) out of the 58 patients showed positive results for Chlamydia pneumoniae., Conclusion: DNA from Chlamydia pneumoniae was identified in the arterial wall of a substantial number of patients with atherosclerosis. This association, which has already been described in other countries, corroborates the evidence favoring a role played by Chlamydia pneumoniae in atherogenesis.

The most common externalization of the acquired immunodeficiency syndrome (AIDS) is opportunist infection, and tuberculosis is one of the most frequent agents. The tuberculous pericarditis has been associated with AIDS, but it exceptionally occurs as the first event in the syndrome. We reported four cases in which tuberculous pericarditis was the initial manifestation of AIDS, characterizing by its clinical picture, diagnostic methods, therapeutics and the evolution of this involvement.

To evaluate the involvement of the heart in patients with seronegative spondyloarthropathies by echodopplercardiography, 35 patients including 20 with ankylosing spondylitis, 10 with Reiter's syndrome and 5 with psoriatic arthritis (21 men and 14 women, with ages ranging from 17-68 years and averaging 38.5) were studied. Most were asymptomatic with respect to the cardiovascular system (65.71%) and 12 oligosymptomatic with palpitations as their main complaint. Each patient had an echocardiogram and electrocardiogram. A two-dimensional echocardiogram demonstrated alterations in 19 patients (54.29%), 28.58% asymptomatic and 25.71% symptomatic. This study revealed most of lesions (17/19-84.47%) followed by the Dopplerechocardiography (10/19-52.63%) and the one-dimensional echocardiography (9/19-47.36%). Abnormal aortic valves were found in 10 patients, in 7 thickenning and in 3 calcifications. The mitral valve was involved in 11 patients, in 8 thickenning, in 1 calcification and in 2 valve prolapse. In ankylosing spondylitis aortic valve disease was found in 8 patients. Dopplerechocardiography evidenced the presence of aortic regurgitation in 4 patients and mitral insufficiency in 3. The Q-T interval was increased in 19 patients, there was one first degree auriculoventricular block, one right branch block and one sinus bradicardia. Thus the echocardiogram is an excellent noninvasive method to disclose cardiac disturbances in patients with seronegative spondyloarthropaties.

Purpose: To study, by a non-invasive method, patients with systemic lupus erythematosus (SLE), to evaluate possible cardiac involvement., Methods: A hundred-eight lupic patients, 60 of them during activity, were studied, independently of cardiovascular signs and symptoms, by M-mode and two-D echocardiography and Doppler. Among the patients in the acute phase, 19 had never used steroid therapy before., Results: Echocardiographic evaluation showed cardiac involvement in all patients who were in clinical activity. Seven had myocardial involvement with systolic impairment. In 35 patients, pericardial effusion was found, all in the acute phase. Regarding endocardial involvement, there were valve thickening in 54 patients in group I (acute phase), valve vegetations in eight and one with mitral valve prolapse. There were only six with valve thickening in group II (remission). Pulmonary hypertension was observed in 15 patients in the activity group and in two during remission., Conclusion: Echocardiogram has showed how frequent cardiac involvement is in SLE, especially during disease activity and being independent of previous steroid therapy. As it is a non-invasive method, it could be used in a routine protocol in the evaluation and follow-up of these patients.

In this communication, we summarize site-directed mutagenesis studies of the lac permease from Escherichia coli, a prototypic H(+)-coupled active transport protein. We classify mutant permeases by phenotype, and suggest possible roles for some individual residues in the mechanism of H+/lactose symport. Although high-resolution structural information is not presently available, kinetic analysis of the partial reactions catalysed by the mutant permeases, as well as biophysical studies, suggest an evolving model for the mechanism of H+/lactose symport.

To modify the lac permease of Escherichia coli for fluorescence spectroscopy, six tryptophan residues at positions 10, 33, 78, 151, 171, and 223 were first replaced individually with phenylalanine by using oligonucleotide-directed site-specific mutagenesis. None of the tryptophan residues is critical for activity, as evidenced by the finding that the mutant permease molecules catalyze lactose/H+ symport almost as well as wild-type permease. Subsequently, a permease molecule was designed in which all of the tryptophan residues were replaced with phenylalanine. Remarkably, the lac permease harboring all six mutations catalyzes active lactose transport about 75% as well as wild-type permease. The fluorescence emission spectrum of purified wild-type permease solubilized in octyl beta-D-glucopyranoside and phospholipid exhibits a broad maximum centered at 350 nm, and the peak is almost completely absent from the spectrum of permease devoid of tryptophan. Furthermore, a new maximum centered at about 306 nm is apparent in the spectrum of the modified permease, suggesting that tyrosine fluorescence in the native protein is quenched by internal energy transfer to tryptophan residues. By using site-directed mutagenesis to replace specified residues in the molecule without tryptophan, it should now be possible to utilize tryptophan fluorescence spectroscopy to study static and dynamic aspects of permease structure and function.