Deciphering the role of the gut microbiome in autoimmune thyroid disease

The aetiology of hyperthyroid Graves’ disease (GD) is incompletely understood. I hypothesized that the gut microbiome affects tolerance to the thyrotropin receptor (TSHR) leading to GD and associated Graves’ orbitopathy (GO). My work comprises two observational studies and two interventional trials, applied to a GD/GO mouse model and GD/GO patients. I applied metataxonomics (16S rRNA gene sequencing) to samples from TSHRimmunised mice from two independent laboratories and observed significant differences in alpha-diversity, beta-diversity and taxonomic profiles. I also compared TSHR-treated and control mice in one centre and identified disease-associated taxonomies (i.e. reduced Bacteroidetes and enriched Firmicutes), correlating with orbital-adipogenesis in diseased but not controls. Changes in gut microbiota taxonomy (e.g. reduced Bacteroides/increased Roseburia spp. and increased Firmicutes:Bacteroidetes ratio) were also observed in GD (n=59) and GO (n=46) patients compared with controls (n=41), and associated with hyperthyroidism or GO severity. Moreover, GD/GO patients-predicted metagenomic pathways included increased “Bacterial epithelial invasion” and “glycosaminoglycan synthesis”. The role of the gut-microbiota in TSHR-induced GD/GO was confirmed by manipulating it in early life using antibiotics which enriched Bacteroides spp. and reduced/ablated disease symptoms. The faecal material transplant from GO patients, despite showing similarities with the GO patients gut microbiota, did not exacerbate murine GO, which also remained unaffected by probiotics. In contrast, in a randomised trial, GD/GO patients receiving probiotics (in addition to anti-thyroid therapy) displayed a more stable gut microbiota composition and sustained improvement in thyroid hormone levels compared with placebo. My results illustrate significant perturbation in the gut microbiota in TSHR-induced murine GD/GO and patients with spontaneous disease. Furthermore, the similarities in differential abundance and disease-associated taxonomies noted in both species support their relevance to disease. Future studies are needed to dissect the mechanistic role of the gut microbiome in activating the immune system and determining the onset of GD/GO

Allergic Diseases

The present Edition "Allergic diseases - highlights in the clinic, mechanisms and treatment" aims to present some recent aspects related to one of the most prevalent daily clinical expression disease. The effort of a group of outstanding experts from many countries reflects a set of scientific studies very promising for a better clinical care and also to the treatment and control of the allergy. This book provides a valuable reference text in several topics of the clinical allergy and basic issues related to the immune system response. The inflammatory reaction understanding in allergic disease is clearly evidenced, as well as new strategies for further researches

Advantages, Disadvantages and Application of Different Technics in PCR Technology

The basic analysis beside the conventional PCR reaction is a plus/minus assay, where the analyses indicates the presence or absence of PCR amplification of a fragment of a DNA molecule. Real-time PCR can monitor the amplification of the PCR product in “real time” with certain fluorogenic reporters. Two the most well-known reporters are SYBR green, which is actually a fluorescent dye that intercalarily binds to DNA, and the TaqMan probe system, which consists of an oligonucleotide probe that is complementary to a certain part of the DNA of interest. The aim of this study was to evaluate, compare and define the specifies of conventional PCR with SYBR green real-time PCR assays and TaqMan probe system. Real-time SYBR green has several disadvantages and advantages compared to the TaqMan probe system. The biggest advantage of the SYBR green is that any pair of primers can be used in this method, with the fact that the primers can be be adapted to real-time PCR technology. Also, the method is much more robust and financially profitable than the TaqMan assay system. The disadvantage of this method is linked to the advantage, because the method is less specific than TaqMan probes that are narrowly bound to a certain part of the DNA. Therefore, it is necessary to perform melting curve analysis after real-time PCR with SYBR green method. SYBR Green is relatively cost benefit and easy to use and technically based on binding the fluorescent dye to double stranded deoxyribonucleic acid (dsDNA) where TaqMan method has more expensive and based on dual labeled oligonucleotide and exonuclease activity of Taq polymerase enzyme. Specificity is the most important concern with the usage of any non-specific dsDNA-binding Dyes such as SYBR Green whiles more specificity showed by labeled oligonucleotide method such as TaqMan. The results of this study indicated that the developed conventional PCR and SYBR green real-time PCR assays are high sensitivity, specificity, accuracy and could be applied as an effective screening tool in different field of laboratory diagnostics