Is Inflammation a Mitochondrial Dysfunction-Dependent Event in Fibromyalgia?

Fibromyalgia (FM) is a complex disorder that affects up to 5% of the general population worldwide. Both mitochondrial dysfunction and inflammation have been implicated in the pathophysiology of FM. We have investigated the possible relationship between mitochondrial dysfunction, oxidative stress, and inflammation in FM. We studied 30 women diagnosed with FM and 20 healthy women. Blood mononuclear cells (BMCs) from FM patients showed reduced level of coenzyme Q10 (CoQ10) and mtDNA contents and high level of mitochondrial reactive oxygen species (ROS) and serum tumor necrosis factor (TNF)-alpha and transcript levels. A significant negative correlation between CoQ10 and TNF-alpha levels (r= -0.588; p < 0.01), and a positive correlation between ROS and TNF-alpha levels (r = 0.791; p < 0.001) were observed accompanied by a significant correlation of visual analogical scale with serum TNF-alpha and transcript levels (r = 0.4507; p < 0.05 and r = 0.7089; p < 0.001, respectively). TNF-alpha release was observed in an in vitro (BMCs) and in vivo (mice) CoQ10 deficiency model. Oral CoQ10 supplementation restored biochemical parameters and induced a significant improvement in clinical symptoms ( p < 0.001). These results lead to the hypothesis that inflammation could be a mitochondrial dysfunction-dependent event implicated in the pathophysiology of FM in several patients indicating at mitochondria as a possible new therapeutic target.Unión Europea FIS PI10/00543Servicio Andaluz de Salud Junta de Andalucía SAS 111242Junta de Andalucía CTS-572

Phosphorylated AKT and MAPK expression in primary tumours and in corresponding metastases and clinical outcome in colorectal cancer patients receiving irinotecan-cetuximab

Clinical observations suggested that a non negligible proportion of patients, ranging from 40% to 70%, does not seem to benefit from the use of anti-EGFR targeted antibodies even in the absence of a mutation of the K- RAS gene. The EGFR pathway activation via the Ras-Raf-MAP-kinase and the protein-serine/threonine kinase AKT could determine resistance to anti-EGFR treatment.We tested the interaction between phosphorylated AKT and MAPK expression in colorectal tumours and corresponding metastases and global outcome in K-RAS wild type patients receiving irinotecan-cetuximab.Seventy-two patients with histologically proven metastatic colorectal cancer, treated with Irinotecan and Cetuximab based chemotherapy, were eligible for our analysis.In metastases pAKT correlated with RR (9% vs. 58%, p\u2009=\u20090.004), PFS (2.3 months vs. 9.2 months p\u2009&lt;\u20090.0001) and OS (6.1 months vs. 26.7 months p\u2009&lt;\u20090.0001) and pMAPK correlated with RR (10% vs. 47%, p\u2009=\u20090.002), PFS (2.3 months vs. 8.6 months p\u2009&lt;\u20090.0001) and OS (7.8 months vs. 26 months p\u2009=\u20090.0004). At multivariate analysis pAKT and pMAPK in metastases were able to independently predict PFS. pAKT in metastases independently correlated with RR as wellpAKT and pMAPK expression in metastases may modulate the activity of EGFR-targeted antibodies. We could speculate that in patients with pAKT and pMAPK metastases expression targeting these factors may be crucial

Autophagy in periodontitis patients and gingival fibroblasts: unraveling the link between chronic diseases and inflammation

Authors are indebted with Ms Monica Glebocki for extensive editing of the manuscriptBackground: Periodontitis, the most prevalent chronic inflammatory disease, has been related to cardiovascular diseases. Autophagy provides a mechanism for the turnover of cellular organelles and proteins through a lysosome-dependent degradation pathway. The aim of this research was to study the role of autophagy in peripheral blood mononuclear cells from patients with periodontitis and gingival fibroblasts treated with a lipopolysaccharide of Porphyromonas gingivalis. Autophagy-dependent mechanisms have been proposed in the pathogenesis of inflammatory disorders and in other diseases related to periodontitis, such as cardiovascular disease and diabetes. Thus it is important to study the role of autophagy in the pathophysiology of periodontitis. Methods: Peripheral blood mononuclear cells from patients with periodontitis (n = 38) and without periodontitis (n = 20) were used to study autophagy. To investigate the mechanism of autophagy, we evaluated the influence of a lipopolysaccharide from P. gingivalis in human gingival fibroblasts, and autophagy was monitored morphologically and biochemically. Autophagosomes were observed by immunofluorescence and electron microscopy. Results: We found increased levels of autophagy gene expression and high levels of mitochondrial reactive oxygen species production in peripheral blood mononuclear cells from patients with periodontitis compared with controls. A significantly positive correlation between both was observed. In human gingival fibroblasts treated with lipopolysaccharide from P. gingivalis, there was an increase of protein and transcript of autophagy-related protein 12 (ATG12) and microtubule-associated protein 1 light chain 3 alpha LC3. A reduction of mitochondrial reactive oxygen species induced a decrease in autophagy whereas inhibition of autophagy in infected cells increased apoptosis, showing the protective role of autophagy. Conclusion: Results from the present study suggest that autophagy is an important and shared mechanism in other conditions related to inflammation or alterations of the immune system, such as periodontiti

Correlazioni molecolari tra melanoma maligno primitivo e rispettive metastasi

Introduzione: Il melanoma è una complessa patologia che origina da diversi pathway molecolari. Una corretta classificazione molecolare del melanoma è utile per una adeguata strategia terapeutica del singolo paziente. Poiché il melanoma è un tumore estremamente etrerogeneo, l’obiettivo di questo studio è stato di individuare la prevalenza e la distribuzione delle mutazioni patogenetiche BRAF, NRAS, c-KIT, ErbB4 tra il tumore primitivo e le rispettive metastasi. Materiali e metodi: Lo studio comprende in tutto 138 casi di melanoma maligno, di cui 56 primitivi ed 82 metastasi sincrone e asincrone. Le lesioni secondarie includono differenti sedi metastatiche: 34 linfonodali, 27 cutanee e sottocutanee, 17 viscerali, 2 cerebrale e 2 ossee. Tutti i campioni di tessuto, fissati in formalina ed inclusi in paraffina, sono stati sottoposti ad analisi mutazionale mediante spettrometria di massa (Sequenom®). Il metodo si basa sulla misura del tempo di volo dei prodotti della reazione di estensione, precedentemente amplificati mediante una multiplex PCR. Risultati: I melanomi primitivi e metastatici sono risultati mutati rispettivamente nel 48% e nel 50% dei casi. Il BRAF risulta, tra gli altri, il gene maggiormente mutato e la mutazione V600E è stata riscontrata con una frequenza del 81,1% nel tumore primitivo e del 77,7% nel tumore metastatico. Mettendo a confronto le valutazioni dello stato mutazionale dei 4 geni studiati nel tumore primitivo e nelle corrispondenti metastasi linfonodali, cutanee e sottocutanee e viscerali, abbiamo riscontrato una concordanza pari al 65% (22 su 34), 78% (11 su 23) e 60% (11 su 15) rispettivamente. La percentuale di concordanza si abbassa fino al 53% analizzando l’assetto molecolare di melanomi primitivi con le rispettive metastasi linfonodali e ad altre metastasi a distanza. Conclusioni: Possiamo dunque affermare che è presente una concordanza non assoluta tra melanoma primitivo e metastatico e che comunque esiste la possibilità, non trascurabile, di un cambiamento nell’assetto molecolare della neoplasia, per cui è importante, per una mirata scelta terapeutica, valutare lo stato mutazionale nelle metastasi quando il materiale tumorale risulta disponibile. E’ inoltre necessario indagare anche le alterazioni molecolari più rare, significative da un punto di vista clinico e terapeutico, con una metodica ad elevata sensibilità e che permetta di valutare una ampio spettro di geni

Correlazioni molecolari tra melanoma maligno primitivo e rispettive metastasi

Introduzione: Il melanoma è una complessa patologia che origina da diversi pathway molecolari. Una corretta classificazione molecolare del melanoma è utile per una adeguata strategia terapeutica del singolo paziente. Poiché il melanoma è un tumore estremamente etrerogeneo, l’obiettivo di questo studio è stato di individuare la prevalenza e la distribuzione delle mutazioni patogenetiche BRAF, NRAS, c-KIT, ErbB4 tra il tumore primitivo e le rispettive metastasi. Materiali e metodi: Lo studio comprende in tutto 138 casi di melanoma maligno, di cui 56 primitivi ed 82 metastasi sincrone e asincrone. Le lesioni secondarie includono differenti sedi metastatiche: 34 linfonodali, 27 cutanee e sottocutanee, 17 viscerali, 2 cerebrale e 2 ossee. Tutti i campioni di tessuto, fissati in formalina ed inclusi in paraffina, sono stati sottoposti ad analisi mutazionale mediante spettrometria di massa (Sequenom®). Il metodo si basa sulla misura del tempo di volo dei prodotti della reazione di estensione, precedentemente amplificati mediante una multiplex PCR. Risultati: I melanomi primitivi e metastatici sono risultati mutati rispettivamente nel 48% e nel 50% dei casi. Il BRAF risulta, tra gli altri, il gene maggiormente mutato e la mutazione V600E è stata riscontrata con una frequenza del 81,1% nel tumore primitivo e del 77,7% nel tumore metastatico. Mettendo a confronto le valutazioni dello stato mutazionale dei 4 geni studiati nel tumore primitivo e nelle corrispondenti metastasi linfonodali, cutanee e sottocutanee e viscerali, abbiamo riscontrato una concordanza pari al 65% (22 su 34), 78% (11 su 23) e 60% (11 su 15) rispettivamente. La percentuale di concordanza si abbassa fino al 53% analizzando l’assetto molecolare di melanomi primitivi con le rispettive metastasi linfonodali e ad altre metastasi a distanza. Conclusioni: Possiamo dunque affermare che è presente una concordanza non assoluta tra melanoma primitivo e metastatico e che comunque esiste la possibilità, non trascurabile, di un cambiamento nell’assetto molecolare della neoplasia, per cui è importante, per una mirata scelta terapeutica, valutare lo stato mutazionale nelle metastasi quando il materiale tumorale risulta disponibile. E’ inoltre necessario indagare anche le alterazioni molecolari più rare, significative da un punto di vista clinico e terapeutico, con una metodica ad elevata sensibilità e che permetta di valutare una ampio spettro di geni.Introduction: Melanoma is a complex disease that arises through multiple etiologic pathways. A correct molecular classification of melanoma can be useful for an appropriate therapeutic strategy of the individual patient. Although melanoma is a tumor extremely heterogeneous, the aim of this study is to investigate the prevalence and distribution of pathogenetic mutations in BRAF, NRAS, c-KIT, ErbB4 genes among primary and metastatic melanoma tissues. Material and methods: In all, the study includes 138 cases of malignant melanoma, including 56 primary and 82 metastases synchronous and asynchronous. Secondary lesions include different metastatic sites: 34 lymph nodes, 27 cutaneous and subcutaneous, 17 visceral, 2 brain and 2 bone. All tissue samples, fixed in formalin and embedded in paraffin, underwent to mutation analysis by mass spectrometry (Sequenom ®). This method is based on evaluation of time of flight of mini-sequencing products, that earlier were amplificated by multiplex-PCR (Plymerase chain reaction). Results: The primary and metastatic melanomas result to be mutated in 48% and in 50% of cases respectively. The BRAF is the more mutated gene and the V600E mutation was found with a frequency of 81.1% in the primary tumor and by 77.7% in metastatic tumor. The evaluation of mutation status of 4 genes studied in the primary tumor and corresponding lymph node metastases, cutaneous and subcutaneous and visceral, showed a concordance of 65% (22 of 34), 78% (11 of 23) and 60 % (11 of 15) respectively. The concordance rate of the molecular profiling of primary melanomas compared to lymph node metastases and other type metastases decrease up to 53%. Conclusion: There is a correlation, not absolute, among primary melanoma and metastases and that exists the possibility, not inconsiderable, of a change in the molecular profiling of the tumor. It is important for a targeted therapeutic choice, assess the mutational status even in metastatic lesions when the material is available. It is also essential, from a clinical and therapeutic point of view, to investigate not common molecular alterations, with a highly sensitive method to allow the assessment of a wide spectrum of genes

Reduced threshold for inhibitory homeostatic responses in migraine motor cortex? A tDCS/TMS study

BACKGROUND AND OBJECTIVE: Neurophysiological studies in migraine have reported conflicting findings of either cortical hyper- or hypoexcitability. In migraine with aura (MwA) patients, we recently documented an inhibitory response to suprathreshold, high-frequency repetitive transcranial magnetic stimulation (hf-rTMS) trains applied to the primary motor cortex, which is in contrast with the facilitatory response observed in the healthy subjects. The aim of the present study was to support the hypothesis that in migraine, because of a condition of basal increased cortical responsivity, inhibitory homeostatic like mechanisms of cortical excitability could be induced by high magnitude stimulation. For this purpose, the hf-rTMS trains were preconditioned by transcranial direct current stimulation (tDCS), a noninvasive brain stimulation technique able to modulate the cortical excitability state. METHODS: Twenty-two MwA patients and 20 patients with migraine without aura (MwoA) underwent trains of 5-Hz repetitive transcranial magnetic stimulation at an intensity of 130% of the resting motor threshold, both at baseline and after conditioning by 15 minutes of cathodal or anodal tDCS. Motor cortical responses to the hf-rTMS trains were compared with those of 14 healthy subjects. RESULTS: We observed abnormal inhibitory responses to the hf-rTMS trains given at baseline in both MwA and MwoA patients as compared with the healthy subjects (P < .00001).The main result of the study was that cathodal tDCS, which reduces the cortical excitability level, but not anodal tDCS, which increases it, restored the normal facilitatory response to the hf-rTMS trains in both MwA and MwoA. CONCLUSIONS: The present findings strengthen the notion that, in migraine with and without aura, the threshold for inducing inhibitory mechanisms of cortical excitability might be lower in the interictal period. This could represent a protective mechanism counteracting cortical hyperresponsivity. Our results could be helpful to explain some conflicting neurophysiological findings in migraine and to get insight into the mechanisms underlying recurrence of the migraine attacks