Abstract: Amyloidosis is a disorder related to errors in protein folding. We present a clinical case of systemic amyloidosis manifesting as hypotension, tachycardia, pain, weight loss, asthenia, anorexia, dysphagia, and mood deflection in a 49-year-old-year-old woman with a previous clinical history of articular and muscular pain, correlated to suspected seronegative arthritis. The blood test revealed kidney insufficiency, an electrocardiogram identified low voltages of the peripheral leads and T waves anomalies. A serum protein electrophoresis revealed the presence of high levels of monoclonal kappa free chains. The woman started to have a sense of suffocation, and after one week she was found dead in her bed. After the autopsy, the results of Congo red staining of the myocardium were characteristic of amyloid. According to the autoptic and the histological examination, death occurred due to acute cardiac and respiratory arrest secondary to amyloid cardiomyopathy in a patient with undiagnosed systemic amyloidosis.
Amyloidosis , Immunoglobulin Light-chain Amyloidosis , Female , Humans , Middle Aged , Amyloidosis/complications , Amyloidosis/diagnosis , Immunoglobulin Light-chain Amyloidosis/complications , Immunoglobulin Light-chain Amyloidosis/diagnosis , Amyloid/analysis , Congo Red , Pain
Local delivery of cytokines or other immunomodulatory components has been applied as a potential therapy for experimental autoimmune encephalomyelitis (EAE), which is used as a model of human multiple sclerosis. We have used herpes simplex virus-based vectors expressing Th2 cytokines IL-4 and IL-10 and have previously shown a significant abolishment of disease symptoms by the virus expressing IL-4 (R8306), but not by the one expressing IL-10 (R8308). In the present study, the aim was to investigate the local and systemic cytokine response after HSV-based gene therapy. We show that the local expression of IL-4 from an HSV vector delivered to the brain converts the cytokine environment from the disease-promoting Th1-prominent to the disease-limiting IL-4 expressing type. We measured the expression of cytokines IL-4, IL-10, IFN-gamma, IL-12p35, IL-12p40 and the novel IL-23p19 from the brain by quantitative LightCycler RT-PCR. We also investigated the systemic cytokine response from the mouse sera. The results indicate that an increase in the Th2 cytokine IL-4 is observed if the diseased mice are treated with IL-4-expressing virus R8306. Surprisingly, the IL-23 expression of R8306 treated mice was at the same level as in the untreated EAE mice. On the contrary, in the R8308 (IL-10 expression) treated mice, the expression of IL-23 was decreased (P < 0.05). We conclude that the favorable effect of IL-4 on the disease development is more important than the downregulation of the Th1 type cytokines (like IL-23), and that IL-4 would be the key mediator of disease abolishment during gene therapy using these vectors.
Encephalomyelitis, Autoimmune, Experimental/therapy , Genetic Therapy , Interleukin-4/genetics , Simplexvirus/genetics , Animals , Brain/virology , Cytokines/genetics , Female , Genetic Vectors , Mice , Mice, Inbred BALB C , Reverse Transcriptase Polymerase Chain Reaction
The real-time principle of reverse transcriptase polymerase chain reaction (RT-PCR) provides a quantitative and reproducible method to detect low copy number transcripts. The quantitative detection of cytokines from tissue samples is complicated by the low expression rates and the short half-lives of the cytokine proteins. The methods have been insensitive and labor-intensive. The LightCycler technique provides a 30-min PCR system with continuous fluorescent detection, analysis of the melting points of the products and user-friendly software for the analysis of the unknown samples. External copy number standards enable the measurement of amounts of the desired targets. We demonstrate the dynamic range of the RT-PCR system from a 100 to 10(7) mRNA copies of the mouse Th1 cytokines interleukin- (IL-) 12p35, 12p40 and IL-23p19 as well as gamma interferon (IFN-gamma) and the housekeeping gene beta-actin, with the usage of fluorescent hybridization probes. The cytokine quantitation was exemplified in murine nervous system samples. A viral transcript, mRNA of alpha trans-inducing factor (alphaTIF), or VP16 gene, of herpes simplex virus (HSV) type 1 was used to quantitate the viral replication in infected cells and in murine nervous system samples. For this viral transcript the linear dynamic range spanned from ten copies to one million copies (highest tested). For all tested cytokine transcripts, the detection level with the dsDNA binding dye SYBR Green I was one log lower than with the hybridizing fluorescent probes. The viral transcript was detected even with the SYBR Green I system at the level of ten copies. The specificity of the PCR was reached with the use of TaqStart antibody, by careful design of primers and probes, by melting temperature analysis and comparison with the gel electrophoresis and Southern blot analysis.
Cytokines/analysis , DNA, Viral/analysis , Herpesvirus 1, Human/physiology , Polymerase Chain Reaction/methods , Th1 Cells/metabolism , Animals , Chlorocebus aethiops , Cytokines/biosynthesis , Cytokines/genetics , Fluorescent Dyes/metabolism , Herpesvirus 1, Human/genetics , Herpesvirus 1, Human/isolation & purification , Humans , Mice , RNA, Messenger/genetics , Reproducibility of Results , Vero Cells
To determine the usefulness of nasal swabs as a simple method for detection of respiratory viruses, we compared nasal swabs and nasopharyngeal aspirates obtained at the same time from the opposite nostrils of 230 children with upper respiratory infection. The sensitivity of nasal swabs was comparable to that of nasopharyngeal aspirates for the detection of all major respiratory viruses except respiratory syncytial virus.
Nasopharynx/virology , Nose/virology , Respiratory Tract Infections/virology , Specimen Handling/methods , Virus Diseases/virology , Viruses/isolation & purification , Adolescent , Child , Child, Preschool , Female , Humans , Infant , Infant, Newborn , Male , Mucus/virology , Respiratory Syncytial Virus Infections/virology , Respiratory Syncytial Virus, Human/isolation & purification , Sensitivity and Specificity
We investigated the expression kinetics of several cytokines in trigeminal ganglia (TG) and in brains of BALB/c mice during the course of ocular herpes simplex virus type 1 (HSV-1) infection. All mice recovered from the infection within 2 weeks. The quantitative rapid real-time RT-PCR method was used to analyze interleukin-4 (IL-4), interferon-gamma (IFN-gamma), IL-12p35, IL-12p40, and the recently described IL-23 (p19) mRNA in TG, brain, and splenocyte samples. In TG, we found elevated expression of mRNA for IL-23 (p19) from early acute infection (day 3) to the beginning of the latent phase (day 14). The increase was not detected in brain or in the spleen. IL-4 expression occurred in both TG and brain from the beginning of the experiment to the latent phase. During the latent phase (days 14 and 31), IL-4 expression was significantly elevated in the brain when compared with the uninfected controls (p < 0.05). Considerable expression of IFN-gamma mRNA was detected in TG of mice during acute HSV-1 infection. The expression of IL-23 was detected also in the brains of the mice, even though no significant changes were found during the acute HSV-1 infection. This is, to our knowledge, the first report to show elevated expression of IL-23 (p19) mRNA (p < 0.05) during viral infection in TG of mice.
Herpes Simplex/metabolism , Interleukins/metabolism , Trigeminal Ganglion/metabolism , Up-Regulation , Acute Disease , Animals , Brain/metabolism , Brain/virology , Chick Embryo , Eye/metabolism , Eye/virology , Female , Herpes Simplex/genetics , Interferon-gamma/metabolism , Interleukin-12/metabolism , Interleukin-12 Subunit p35 , Interleukin-12 Subunit p40 , Interleukin-23 , Interleukin-23 Subunit p19 , Interleukin-4/metabolism , Interleukins/genetics , Mice , Mice, Inbred BALB C , Protein Subunits/metabolism , RNA, Messenger/metabolism , Trigeminal Ganglion/virology
The microbiological and sensory quality of cabbage casserole and mixed vegetable salad with mayonnaise was assessed after production, on the sell-by date and 7 d later. Samples were taken directly from five different plants, stored at 4°C and analyzed by three different laboratories. Sell-by periods were 8 to 14 d after the day of production. No Bacillus cereus , Clostridium perfringens , Staphylococcus aureus (salad), fecal streptococci or coliforms (casserole) were detected. Casseroles had median aerobic plate counts (APC) of 1.5 × 102, 3.3 × 102 and 4.5 × 103 cfu/g on different analysis times. Yeasts were detected in some casseroles on the sell-by dates. A few more had yeasts and/or molds a week later. Taste, odor, consistency and appearance scores showed a steady decrease during storage. Nine casseroles were deemed unfit for human consumption 7 d after the sell-by dates. Main defects were sliminess and acid, fermented taste and visible mold spots. Salads had a median APC of 5.2 × 103 cfu/g after production, which remained constant during storage. Salads from all companies contained lactobacilli and counts increased slightly during storage. Molds were encountered in samples of only one company and yeasts primarily of another. Median sensory scores decreased slightly during storage. One salad was deemed unfit on the sell-by date and six a week later. Main defects were musty and/or fermented taste and odor and watery consistency. Linear regression equations between taste and log10 microbial counts showed very low or no correlation. Also, the counts of sensorially unfit samples varied from low to high.
