Human Outbreak of Trichinellosis Caused by Trichinella papuae Nematodes, Central Kampong Thom Province, Cambodia.

In September 2017, a severe trichinellosis outbreak occurred in Cambodia after persons consumed raw wild pig meat; 33 persons were infected and 8 died. We collected and analyzed the medical records for 25 patients. Clinical signs and symptoms included myalgia, facial or peripheral edema, asthenia, and fever. We observed increased levels of creatine phosphokinase and aspartate aminotransferase-, as well as eosinophilia. Histopathologic examination of muscle biopsy specimens showed nonencapsulated Trichinella larvae. A Trichinella excretory/secretory antigen ELISA identified Trichinella IgM and IgG. Biopsy samples were digested and larvae were isolated and counted. PCR for the 5S rDNA intergenic spacer region and a multiplex PCR, followed by sequencing identified the parasite as Trichinella papuae. This species was identified in Papua New Guinea during 1999 and in several outbreaks in humans in Thailand. Thus, we identified T. papuae nematodes in humans in Cambodia.

Prevalence of Abnormal Cervical Cytology in HIVNegative Women Participating in a Cervical Cancer Screening Program in Calmette Hospital, Cambodia.

Prevalence of Abnormal Cervical Cytology in HIVNegative Women Participating in a Cervical Cancer Screening Program in Calmette Hospital, Cambodia.

Multiphoton imaging of tumor biomarkers with conjugates of single-domain antibodies and quantum dots.

An ideal multiphoton fluorescent nanoprobe should combine a nanocrystal with the largest possible two-photon absorption cross section (TPACS) and the smallest highly specific recognition molecules bound in an oriented manner. CdSe/ZnS quantum dots (QDs) conjugated to 13-kDa single-domain antibodies (sdAbs) derived from camelid IgG or streptavidin have been used as efficient two-photon excitation (TPE) probes for carcinoembryonic antigen (CEA) imaging on normal human appendix and colon carcinoma tissue. The TPACS for some conjugates was higher than 49,000 GM (Goeppert-Mayer units), considerably exceeding that of organic dyes being close to the theoretical value of 50,000 GM calculated for CdSe QDs. The ratio of sdAb-QD emission to the autofluorescence for 800 nm TPE was 40 times higher than that for 457.9 nm one-photon excitation. TPE ensures a clear discrimination of CEA-overexpressing tumor areas from normal tissue. Oriented sdAb-QD conjugates are bright specific labels for detecting low concentrations of antigens using multiphoton microscopy. FROM THE CLINICAL EDITOR: This study demonstrates carcinoembryonic antigen imaging on normal human appendix and colon carcinoma tissue utilizing CdSe/ZnS quantum dots conjugated to streptavidin or to 13-kDa single-domain antibodies as efficient two-photon excitation probes.

Juvenile frontotemporal dementia with parkinsonism associated with tau mutation G389R.

Frontotemporal lobe degeneration includes a large spectrum of neurodegenerative disorders. Patients with frontotemporal dementia with parkinsonism linked to chromosome 17 exhibit heterogeneity in both clinical and neuropathological features. Here, we report the case of a young patient with a G389R mutation. This teenager girl was 17 years old when she progressively developed severe behavioral disturbances. First, she was considered to be suffering from atypical depression. After 2 years, she was referred to the department of neurology. By this time, the patient exhibited typical frontotemporal dementia with mild extrapyramidal disorders. The main behavioral features included apathy and reduced speech output. MRI and SPECT showed a frontotemporal atrophy and hypofixation, respectively. She died 7 years after onset. Three relatives on her father side had also died after early onset dementia. Genetic testing revealed a heterozygous guanine to cytosine mutation at the first base of codon 389 (Exon 13) of MAPT, the tau gene, resulting in a glycine to arginine substitution, in the patient and her non-affected father. Postmortem neuropathological and biochemical data indicate a Pick-like tau pathology but with phosphoserine 262-positive immunoreactivity. This case is remarkable because of the extremely early onset of the disease.

Efficacy of ω-3 polyunsaturated fatty acids for the treatment of refractory hydroa vacciniforme.

Hydroa vacciniforme (HV) is a rare photodermatosis. Several therapies, with sometimes severe side effects, have been used in isolated cases. We report a case of refractory HV successfully treated with dietary fish oil rich in ω-3 polyunsaturated fatty acids.

Oriented conjugates of single-domain antibodies and quantum dots: toward a new generation of ultrasmall diagnostic nanoprobes.

Common strategy for diagnostics with quantum dots (QDs) utilizes the specificity of monoclonal antibodies (mAbs) for targeting. However QD-mAbs conjugates are not always well-suited for this purpose because of their large size. Here, we engineered ultrasmall nanoprobes through oriented conjugation of QDs with 13-kDa single-domain antibodies (sdAbs) derived from llama IgG. Monomeric sdAbs are 12 times smaller than mAbs and demonstrate excellent capacity for refolding. sdAbs were tagged with QDs through an additional cysteine residue integrated within the C terminal of the sdAb. This approach allowed us to develop sdAbs-QD nanoprobes comprising four copies of sdAbs coupled with a QD in a highly oriented manner. sdAbs-QD conjugates specific to carcinoembryonic antigen (CEA) demonstrated excellent specificity of flow cytometry quantitative discrimination of CEA-positive and CEA-negative tumor cells. Moreover, the immunohistochemical labeling of biopsy samples was found to be comparable or even superior to the quality obtained with gold standard protocols of anatomopathology practice. sdAbs-QD-oriented conjugates as developed represent a new generation of ultrasmall diagnostic probes for applications in high-throughput diagnostic platforms. FROM THE CLINICAL EDITOR: The authors report the development of sdAbs-QD-oriented conjugates, comprised of single domain antibodies that are 12 times smaller than regular mAb-s and quantum dots. These ultrasmall diagnostic probes represent a new generation of functionalized ODs for applications in high-throughput diagnostic platforms.

Preliminary investigation of a mice model of Klebsiella pneumoniae subsp. ozaenae induced pneumonia.

In the present study, we comparatively assessed the pathophysiological mechanisms developed during lung infection of BALB/C female mice infected by an original wild type Klebsiella pneumoniae subsp. ozaenae strain (CH137) or by a referent subspecies K. pneumoniae. subsp. pneumoniae strain (ATCC10031). The mice infected with 2.106 CFU K. p. subsp. pneumoniae (n = 10) showed transient signs of infection and all of them recovered. All of those infected with 1.106 CFU K. p. subsp. ozaenae (n = 10) developed pneumonia within 24 h and died between 48 and 72 h. Few macrophages, numerous polymorphonuclear cells and lymphocytes were observed in their lungs in opposite to K. p. subsp. pneumoniae. In bronchoalveolar lavage, a significant increase in MIP-2, IL-6, KC and MCP-1 levels was only observed in K. p. subsp. ozaenae infected mice whereas high levels of TNF-α were evidenced with the two subspecies. Our findings indicated a lethal effect of a wild type K. p. subsp. ozaenae strain by acute pneumonia reflecting an insufficient alveolar macrophage response. This model might be of a major interest to comparatively explore the pathogenicity of K. p. subsp ozaenae strains and to further explore the physiopathological mechanisms of gram-negative bacteria induced human pneumonia.

Emerging applications of fluorescent nanocrystals quantum dots for micrometastases detection.

The occurrence of metastases is one of the main causes of death in many cancers and the main cause of death for breast cancer patients. Micrometastases of disseminated tumour cells and circulating tumour cells are present in more than 30% of breast cancer patients without any clinical or even histopathological signs of metastasis. Low abundance of these cell types in clinical diagnostic material dictates the necessity of their enrichment prior to reliable detection. Current micrometastases detection techniques are based on immunocytochemical and molecular methods suffering from low efficiency of tumour cells enrichment and observer-dependent interpretation. The use of highly fluorescent semiconductor nanocrystals, also known as "quantum dots" and nanocrystal-encoded microbeads tagged with a wide panel of antibodies against specific tumour markers offers unique possibilities for ultra-sensitive micrometastases detection in patients' serum and tissues. The nanoparticle-based diagnostics provides an opportunity for highly sensitive parallel quantification of specific proteins in a rapid and low-cost method, thereby providing a link between the primary tumour and the micrometastases for early diagnosis.

Enterovirus-related activation of the cardiomyocyte mitochondrial apoptotic pathway in patients with acute myocarditis.

AIMS: We examined the impact of enterovirus (EV) cardiac replication activity on the endomyocardial mitochondrial pathway in patients with acute myocarditis. METHODS AND RESULTS: Levels of apoptotic cardiomyocytes were determined by TUNEL and ligation-mediated polymerase chain reaction (PCR) assays and EV replication activity was assessed by immunostaining of EV VP1 capsid protein in ventricular myocytes of patients with acute myocarditis (n = 25), and healthy heart controls (n = 15). Ratio of cytosolic/mitochondrial cytochrome c concentrations was determined by ELISA assay, levels of active caspase-9 were determined by western blot analysis and Bax/Bcl2 mRNA ratio was assessed by real-time reverse transcription-polymerase chain reaction (RT-PCR) in the same cardiac tissues. Patients with EV-associated acute myocarditis (n = 15) exhibited a significantly higher number of apoptotic cardiomyocytes than those with non-EV-associated acute myocarditis (n = 10) and controls (n = 15) (P < 0.001). Endomyocardial ratio of cytosolic/mitochondrial cytochrome c concentrations and levels of active caspase-9 protein were significantly increased in EV than in non-EV-related myocarditis patients (P < 0.001). Moreover, Bax/Bcl2 mRNA ratio was significantly increased in EV than in non-EV-related myocarditis patients (P < 0.001). CONCLUSION: Our findings evidence an EV-related activation of the cardiomyocyte mitochondrial apoptotic pathway in patients with acute myocarditis. Moreover, our results indicate that this EV-induced pro-apoptotic mechanism could be partly related to an up-regulation of Bax expression, and suggest that inhibition of this cell death process may constitute the basis for novel therapies.

Semiconductor quantum dots for multiplexed bio-detection on solid-state microarrays.

Understanding cellular systems requires identification and analysis of their multiple components and determination of how they act together and are regulated. Microarray technology is one of the few tools that is able to solve such problems. It is based on high-throughput recognition of a target to the probe and has the potential to simultaneously measure the presence of numerous molecules in multiplexed tests, all contained in a small drop of test fluid. Microarrays allow the parallel analysis of genomic or proteomic content in healthy versus disease-affected or altered tissues or cells. The signal read-out from the microarrays is done with organic dyes which often suffer of photobleaching, low brightness and background fluorescence. Recent data show that the use of fluorescent nanocrystals named "quantum dots" (QDs) allows to push these limits away. QDs are sufficiently bright to be detected as individual particles, extremely resistant against photobleaching and provide unique possibilities for multiplexing, thus supplying the microarray technology with a novel read-out option enabling the sensitivity of detection to reach the single-molecule level. This paper reviews QDs applications to microarray-based detection and demonstrates how the combination of microarray and QDs technologies may increase sensitivity and highly parallel capacities of multiplexed microarrays. Such a combination should provide the breakthrough results in drug discovery, cancer diagnosis and establish new therapeutic approaches through the identification of binding target molecules and better understanding of cell signalling pathways.

Modeling and quantifying biochemical changes in C6 tumor gliomas by Fourier transform infrared imaging.

The purpose of the study was to investigate molecular changes associated with glioma tissues using FT-IR microspectroscopic imaging (FT-IRM). A multivariate statistical analysis allowed one to successfully discriminate between normal, tumoral, peri-tumoral, and necrotic tissue structures. Structural changes were mainly related to qualitative and quantitative changes in lipid content, proteins, and nucleic acids that can be used as spectroscopic markers for this pathology. We have developed a spectroscopic model of glioma to quantify these chemical changes. The model constructed includes individual FT-IR spectra of normal and glioma brain constituents such as lipids, DNA, and proteins (measured on delipidized tissue). Modeling of FT-IR spectra yielded fit coefficients reflecting the chemical changes associated with a tumor. Our results demonstrate the ability of FT-IRM to assess the importance and distribution of each individual constituent and its variation in normal brain structures as well as in the different pathological states of glioma. We demonstrated that (i) cholesterol and phosphatidylethanolamine contributions are highest in corpus callosum and anterior commissure but decrease gradually towards the cortex surface as well as in the tumor, (ii) phosphatidylcholine contribution is highest in the cortex and decreases in the tumor, (iii) galactocerebroside is localized only in white, but not in gray matter, and decreases in the vital tumor region while the necrosis area shows a higher concentration of this cerebroside, (iv) DNA and oleic acid increase in the tumor as compared to gray matter. This approach could, in the future, contribute to enhance diagnostic accuracy, improve the grading, prognosis, and play a vital role in therapeutic strategy and monitoring.

FTIR protein secondary structure analysis of human ascending aortic tissues.

The advent of moderate dilatations in ascending aortas is often accompanied by structural modifications of the main components of the aortic tissue, elastin and collagen. In this study, we have undertaken an approach based on FTIR microscopy coupled to a curve-fitting procedure to analyze secondary structure modifications in these proteins in human normal and pathological aortic tissues. We found that the outcome of the aortic pathology is strongly influenced by these proteins, which are abundant in the media of the aortic wall, and that the advent of an aortic dilatation is generally accompanied by a decrease of parallel beta-sheet structures. Elastin, essentially composed of beta-sheet structures, seems to be directly related to these changes and therefore indicative of the elastic alteration of the aortic wall. Conventional microscopy and confocal fluorescence microscopy were used to compare FTIR microscopy results with the organization of the elastic fibers present in the tissues. This in-vitro study on 6 patients (three normal and three pathologic), suggests that such a spectroscopic marker, specific to aneurismal tissue characterization, could be important information for surgeons who face the dilemma of moderate aortic tissue dilatation of the ascending aortas.

Active Coxsackieviral B infection is associated with disruption of dystrophin in endomyocardial tissue of patients who died suddenly of acute myocardial infarction.

OBJECTIVES: In this study, we evaluated the potential direct role of enterovirus (EV) cardiac infections in the pathogenesis of myocardial infarction (MI). BACKGROUND: Enteroviruses (Picornaviridae) have been suspected to play a role in the development of acute MI. METHODS: The presence of EV ribonucleic acid (RNA) sequences and capsid viral protein 1 (VP1) and the virus-mediated focal disruption of dystrophin were retrospectively investigated by reverse transcriptase-polymerase chain reaction and immunohistochemistry assays in endomyocardial tissues of patients who died suddenly of acute MI by comparison with similar samples of control patients matched for gender, residence area, and year of death. RESULTS: Enterovirus infection markers were detected in 20 (40%) of 50 patients who died suddenly of MI, 2 (4%) of 50 matched subjects without cardiac disease (p < 0.001), and 4 (8%) of 50 matched patients exhibiting a noncoronary chronic cardiopathy (p < 0.001). All of the EV RNA-positive patients exhibited VP1, which provided evidence of viral protein synthesis activity. The VP1 gene sequences amplified after cloning from myocardial or coronary samples of 8 of the MI patients and showed a strong homology with sequences of coxsackievirus B2 and B3 serotypes. Moreover, in the endomyocardial tissue of these 8 patients, immunohistochemical analyses demonstrated that there was disruption of the sarcolemmal localization of dystrophin in the same tissue areas that were infected by coxsackieviruses. CONCLUSIONS: Our findings demonstrate a significantly higher proportion of active coxsackievirus B cardiovascular infections in patients who suddenly died of MI compared with matched control subjects, suggesting that these EVs may significantly contribute to the pathogenesis of acute MI by a focal disruption of the dystrophin-glycoprotein complex.

Indolent CD8-positive lymphoid proliferation of the ear: a distinct primary cutaneous T-cell lymphoma?

The authors report 4 cases of cutaneous lymphoproliferation unusual by their histology and their clinical presentation. Each presented with a history of a slow growing nodule on the ear. Despite the indolent clinical evolution, the histology suggested a high-grade lymphoma. All lesions consisted of a dense, diffuse proliferation of monomorphous medium-sized T cells throughout the dermis and subcutis. There was no epidermotropism and a grenz zone was clearly present in each case. The tumor cells displayed irregular blastlike nuclei, with small nucleoli and clear chromatin and had a CD3+, CD8+, CD4+, TIA1+, granzyme B(-)immunophenotype with a loss of other T-cell antigens. The 3 cases with available material for polymerase chain reaction studies displayed a monoclonal T-cell rearrangement of the T-cell receptor-gamma chain. These cases do not correspond to a recognized cutaneous T-cell lymphoma as described in the recent WHO/EORTC classification. The apparent striking propensity for the ear suggests that they might represent a specific entity. Further cases are needed to confirm this hypothesis. It is important for such indolent lesions to be known to avoid over treatment.

Discriminating healthy from tumor and necrosis tissue in rat brain tissue samples by Raman spectral imaging.

The purpose of this study was to investigate molecular changes associated with glioma tissues by Raman microspectroscopy in order to develop its use in clinical practice. Spectroscopic markers obtained from C6 glioma tissues were compared to conventional histological and histochemical techniques. Cholesterol and phospholipid contents were highest in corpus callosum and decreased gradually towards the cortex surface as well as in the tumor. Two different necrotic areas have been identified: a fully necrotic zone characterized by the presence of plasma proteins and a peri-necrotic area with a high lipid content. This result was confirmed by Nile Red staining. Additionally, one structure was detected in the periphery of the tumor. Invisible with histopathological hematoxylin and eosin staining, it was revealed by immunohistochemical Ki-67 and MT1-MMP staining used to visualize the proliferative and invasive activities of glioma, respectively. Hierarchical cluster analysis on the only cluster averaged spectra showed a clear distinction between normal, tumoral, necrotic and edematous tissues. Raman microspectroscopy can discriminate between healthy and tumoral brain tissue and yield spectroscopic markers associated with the proliferative and invasive properties of glioblastoma. Development of in vivo Raman spectroscopy could thus accurately define tumor margins, identify tumor remnants, and help in the development of novel therapies for glioblastoma.

The NC1 domain of type XIX collagen inhibits in vivo melanoma growth.

Type XIX collagen is a minor collagen that localizes to basement membrane zones, together with types IV, XV, and XVIII collagens. Because several NC1 COOH-terminal domains of other chains from basement membrane collagens were reported to exhibit antitumor activity, we decided to study the effects of the NC1(XIX) collagen domain on tumor progression using an experimental in vivo model of mouse melanoma. We observed a 70% reduction in tumor volume in NC1(XIX)-treated mice compared with the corresponding controls. Histologic examination of the tumors showed a strong decrease in tumor vascularization in treated mice. In vitro, NC1(XIX) inhibited the migrating capacity of tumor cells and their capacity to invade Matrigel. It also inhibited the capacity of human microvascular endothelial cells to form pseudotubes in Matrigel. This effect was accompanied by a strong inhibition of membrane type-1 matrix metalloproteinase (matrix metalloproteinase-14) and vascular endothelial growth factor expression. Collectively, our data indicate that the NC1 domain of type XIX collagen exerts antitumor activity. This effect is mediated by a strong inhibition of the invasive capacities of tumor cells and antiangiogenic effects. NC1(XIX) should now be considered as a new member of the basement membrane collagen-derived matrikine family with antitumor and antiangiogenic activity.

Brain tissue characterisation by infrared imaging in a rat glioma model.

Pathological changes associated with the development of brain tumor were investigated by Fourier transform infrared microspectroscopy (FT-IRM) with high spatial resolution. Using multivariate statistical analysis and imaging, all normal brain structures were discriminated from tumor and surrounding tumor tissues. These structural changes were mainly related to qualitative and quantitative changes in lipids (tumors contain little fat) and were correlated to the degree of myelination, an important factor in several neurodegenerative disorders. Lipid concentration and composition may thus be used as spectroscopic markers to discriminate between healthy and tumor tissues. Additionally, we have identified one peculiar structure all around the tumor. This structure could be attributed to infiltrative events, such as peritumoral oedema observed during tumor development. Our results highlight the ability of FT-IRM to identify the molecular origin that gave rise to the specific changes between healthy and diseased states. Comparison between pseudo-FT-IRM maps and histological examinations (Luxol fast blue, Luxol fast blue-cresyl violet staining) showed the complementarities of both techniques for early detection of tissue abnormalities.

Self-assembly of charged microclusters of CdSe/ZnS core/shell nanodots and nanorods into hierarchically ordered colloidal arrays.

A thermodynamically driven self-organization of microclusters of semiconductor nanocrystals with a narrow size distribution into periodic two-dimensional (2D) arrays is an attractive low-cost technique for the fabrication of 2D photonic crystals. We have found that CdSe/ZnS core/shell quantum dots or quantum rods, transferred in aqueous phase after capping with the bifunctional surface-active agent DL-cysteine, form on a poly-L-lysine coated surface homogeneously sized micro-particles, droplet-like spheroid clusters and hexagon-like colloidal crystals self-organized into millimetre-sized 2D hexagonal assemblies. The presence of an organic molecular layer around the micro-particles prevents immediate contact between them, forming an interstitial space which may be varied in thickness by changing the origin of the molecular layer capping nanocrystals. Due to the high refractive index of CdSe and the low refractive index of the interstitial spaces, these structures are expected to have deep gaps in their photonic band, forming hierarchically ordered 2D arrays of potentially photonic materials.

[Autopsy room and the July 30th 2004 memorandum]. / Salle d'autopsie et circulaire du 30 juillet 2004.

With the July 30th 2004 memorandum, for the first time a text is specifically dedicated to the architecture of the autopsy room. This memorandum reaffirms certain technical specifications stated in the May 7th 2001 decree applicable to hospital mortuaries. It supplements or modifies certain elements, particularly liquid waste processing, which will require new arrangements in death chambers and new expenditures for hospital administrations. It includes the principle of precaution and requires a new approach to handling human corpses in the autopsy room.

In vivo overexpression of tumstatin domains by tumor cells inhibits their invasive properties in a mouse melanoma model.

Our previous studies demonstrated that a synthetic peptide encompassing residues 185-203 of the noncollagenous (NC1) domain of the alpha3 chain of type IV collagen, named tumstatin, inhibits in vitro melanoma cell proliferation and migration. In the present study, B16F1 melanoma cells were stably transfected to overexpress the complete tumstatin domain (Tum 1-232) or its C-terminal part, encompassing residues 185-203 (Tum 183-232). Tumstatin domain overexpression inhibited B16F1 in vitro cell proliferation, anchorage-independent growth, and invasive properties. For studying the in vivo effect of overexpression, representative clones were subcutaneously injected into the left side of C57BL6 mice. In vivo tumor growth was decreased by -60% and -56%, respectively, with B16F1 cells overexpressing Tum 1-232 or Tum 183-232 compared to control cells. This inhibitory effect was associated with a decrease of in vivo cyclin D1 expression. We also demonstrated that the overexpression of Tum 1-232 or Tum 183-232 induced an in vivo down-regulation of proteolytic cascades involving matrix metalloproteinases (MMPs), especially the production or activation of MMP-2, MMP-9, MMP-13, as well as MMP-14. The plasminogen activation system was also altered in tumors with a decrease of urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA) and a strong increase of plasminogen activator inhibitor-1 (PAI-1). Collectively, our results demonstrate that tumstatin or its C-terminal antitumor fragment, Tum 183-232, inhibits in vivo melanoma progression by triggering an intracellular transduction pathway, which involves a cyclic AMP (cAMP)-dependent mechanism.