Thyroid hormone promotes differentiation of colon cancer stem cells

Tumor formation and maintenance depend on a small fraction of cancer stem cells (CSCs) that can self-renew and generate a wide variety of differentiated cells. CSCs are resistant to chemotherapy and radiation, and can represent a reservoir of cancer cells that often cause relapse after treatment. Evidence suggests that CSCs also give rise to metastases. Thyroid hormone (TH) controls a variety of biological processes including the development and functioning of most adult tissues. Recent years has seen the emergence of an intimate link between TH and multiple steps of tumorigenesis. Thyroid hormone controls the balance between the proliferation and differentiation of CSCs, and may thus be a druggable anti-cancer agent. Here, we review current understanding of the effects of TH on colorectal CSCs, including the cross regulatory loops between TH and regulators of CSC stemness. Targeting TH in the tumor microenvironment may improve treatment strategies

Novel Approach for Evaluation of Bacteroides fragilis Protective Role against Bartonella henselae Liver Damage in Immunocompromised Murine Model

Bartonella henselae is a gram-negative facultative intracellular bacterium and is the causative agent of cat-scratch disease. Our previous data have established that Bacteroides fragilis colonization is able to prevent B. henselae damages through the polysaccharide A (PSA) in an experimental murine model. In order to determine whether the PSA is essential for the protection against pathogenic effects of B. henselae in immunocompromised hosts, SCID mice were co-infected with B. fragilis wild type or its mutant B. fragilis 1PSA and the effects of infection on murine tissues have been observed by High-Frequency Ultrasound (HFUS), histopathological examination, and Transmission Electron Microscopy (TEM). For the first time, echostructure, hepatic lobes length, vascular alterations, and indirect signs of hepatic dysfunctions, routinely used as signs of disease in humans, have been analyzed in an immunocompromised murine model. Our findings showed echostructural alterations in all infected mice compared with the Phosphate Buffer Solution (PBS) control group; further, those infected with B. henselae and co-infected with B. henselae/B. fragilis 1PSA presented the major echostructural alterations. Half of the mice infected with B. henselae and all those co-infected with B. henselae/B. fragilis 1PSA have showed an altered hepatic echogenicity compared with the renal cortex. The echogenicity score of co-infected mice with B. henselae/B. fragilis 1PSA differed significantly compared with the PBS control group (p < 0.05). Moreover the inflammation score of the histopathological evaluation was fairly concordant with ultrasound findings. Ultrastructural analysis performed by TEM revealed no significant alterations in liver samples of SCID mice infected with B. fragilis wild type while those infected with B. fragilis 1PSA showed the presence of collagen around the main vessels compared with the PBS control group. The liver samples of mice infected with B. henselae showed macro-areas rich in collagen, stellate cells, and histiocytic cells. Interestingly, our data demonstrated that immunocompromised SCID mice infected with B. henselaeand co-infected with B. henselae/B. fragilis ΔPSA showed the most severe morpho-structural liver damage. In addition, these results suggests that the HFUS together with histopathological evaluation could be considered good imaging approach to evaluate hepatic alterations

Seroprevalence of Bartonella henselae in patients awaiting heart transplant in Southern Italy

Background Bartonella henselae is the etiologic agent of cat-scratch disease. B. henselae infections are responsible for a widening spectrum of human diseases, although often symptomless, ranging from self-limited to life-threatening and show different courses and organ involvement due to the balance between host and pathogen. The role of the host immune response to B. henselae is critical in preventing progression to systemic disease. Indeed in immunocompromised patients, such as solid organ transplant patients, B. henselae results in severe disseminated disease and pathologic vasoproliferation. The purpose of this study was to determine the seroprevalence of B. henselae in patients awaiting heart transplant compared to healthy individuals enrolled in the Regional Reference Laboratory of Transplant Immunology of Second University of Naples. Methods Serum samples of 38 patients awaiting heart transplant in comparison to 50 healthy donors were examined using immunfluorescence assay. Results We found a B. henselae significant antibody positivity rate of 21% in patients awaiting heart transplant ( p = 0.002). There was a positive rate of 8% ( p > 0.05) for immunoglobulin (Ig)M and a significant value of 13% ( p = 0.02) for IgG, whereas controls were negative both for IgM and IgG antibodies against B. henselae . The differences in comorbidity between cases and controls were statistically different (1.41 ± 0.96 vs 0.42 ± 0.32; p = 0.001). Conclusions Although this study was conducted in a small number of patients, we suggest that the identification of these bacteria should be included as a routine screening analysis in pretransplant patients

Loss of p53 activates thyroid hormone via type 2 deiodinase and enhances DNA damage

: The Thyroid Hormone (TH) activating enzyme, type 2 Deiodinase (D2), is functionally required to elevate the TH concentration during cancer progression to advanced stages. However, the mechanisms regulating D2 expression in cancer still remain poorly understood. Here, we show that the cell stress sensor and tumor suppressor p53 silences D2 expression, thereby lowering the intracellular THs availability. Conversely, even partial loss of p53 elevates D2/TH resulting in stimulation and increased fitness of tumor cells by boosting a significant transcriptional program leading to modulation of genes involved in DNA damage and repair and redox signaling. In vivo genetic deletion of D2 significantly reduces cancer progression and suggests that targeting THs may represent a general tool reducing invasiveness in p53-mutated neoplasms

Characterization of Neisseria meningitidis rifampicin resistant strains: highlight on molecular mechanisms underlying the phenotypic traits associated with the RpoB H553Y substitution

Rifampicin chemoprophylaxis against Neisseria meningitidis infections led to the onset of rifampicin resistance in clinical isolates harboring point mutations in the rpoB gene, coding for the RNA polymerase β chain. These resistant strains are rare in medical practice, suggesting their decreased fitness in the human host. In this study, we isolated rifampicin-resistant rpoB mutants from hypervirulent serogroup C strain 93/4286 and analyzed their different properties, including the ability to grow and survive in different culture media and in differentiated THP-1 human monocytes and to compete with the wild-type strain in vitro. Our results demonstrate that different rpoB mutations (H553Y, H553R and S549F) may have different effects, ranging from low- to high-cost, on bacterial fitness in vitro. Moreover, we found that the S549F mutation confers temperature sensitivity, possibly explaining why it is observed very rarely in clinical isolates. Comparative high-throughput RNA sequencing analysis of bacteria grown in chemically defined medium demonstrated that the low-cost H553Y substitution resulted in global transcriptional changes that functionally mimic the stringent response. Interestingly, many virulence-associated genes, including those coding for meningococcal type IV pili, porin A, adhesins/invasins, IgA protease, two-partner secretion system HrpA/HrpB, enzymes involved in resistance to oxidative injury, lipooligosaccharide sialylation, and capsular polysaccharide biosynthesis, were down regulated in the H553Y mutant compared to their level of regulation in the wild-type strain. These data might account for the reduced capacity of H553Y mutant to grow and survive in differentiated THP-1 cells and explain the rarity of this mutant in clinical isolates

Cardiovascular and Neuronal Consequences of Thyroid Hormones Alterations in the Ischemic Stroke

Ischemic stroke is one of the leading global causes of neurological morbidity and decease. Its etiology depends on multiple events such as cardiac embolism, brain capillaries occlusion and atherosclerosis, which ultimately culminate in blood flow interruption, incurring hypoxia and nutrient deprivation. Thyroid hormones (THs) are pleiotropic modulators of several metabolic pathways, and critically influence different aspects of tissues development. The brain is a key TH target tissue and both hypo- and hyperthyroidism, during embryonic and adult life, are associated with deranged neuronal formation and cognitive functions. Accordingly, increasing pieces of evidence are drawing attention on the consistent relationship between the THs status and the acute cerebral and cardiac diseases. However, the concrete contribution of THs systemic or local alteration to the pathology outcome still needs to be fully addressed. In this review, we aim to summarize the multiple influences that THs exert on the brain and heart patho-physiology, to deepen the reasons for the harmful effects of hypo- and hyperthyroidism on these organs and to provide insights on the intricate relationship between the THs variations and the pathological alterations that take place after the ischemic injury

Analisi del ruolo patogenetico del gene siaA di Neisseria meningitidis in un modello murino di meningite meningococcica.

Il polisaccaride capsulare di Neisseria meningitidis è uno dei principali fattori di virulenza che permette al batterio di eludere l’immunità dell’ospite consentendogli di sopravvivere nel circolo ematico. Tuttavia, la capsula meningococcica svolge ruoli contrastanti nella patogenesi: se da un lato, le proprietà anti-fagocitiche sono essenziali per la crescita batterica nel sangue, dall’altro, l’espressione del polisaccaride capsulare inibisce la colonizzazione e l’invasione della barriera nasofaringea. La presenza della capsula dipende dall’espressione dei geni coinvolti nella biosintesi, modificazione e trasporto della capsula, che si trovano nelle regioni A, B, e C del locus cps. I polisaccaridi capsulari dei sierogruppi B, C, W-135 e Y contengono acido sialico e la regione A del locus cps presenta un set di geni conservati: siaA, siaB, e siaC, necessari per la sintesi dell’acido sialico e il gene sierogruppo-specifico siaD, codificante per l’enzima polisialil-transferasi. Per valutare il ruolo della capsula nell’istaurarsi dell’infezione meningoccica in un modello murino d’infezione abbiamo utilizzato il ceppo 93/4286 di sierogruppo C, inattivato nel locus genico siaA (93/4286ΩsiaA), mediante inserimento di una cassetta circolare recante il gene per la resistenza all’eritromicina (ermC’). Dati di sopravvivenza, parametri clinici e microbiologici hanno consentito di dimostrare che il ceppo isogenico difettivo nella componente enzimatica SiaA mostra una ridotta capacità di stabilire meningite meningococcica nell’ospite murino. Il ceppo isogenico 93/4286ΩsiaA ha mostrato una LD50 di circa 108 CFU rispetto ad una LD50 di 104 CFU per il ceppo selvatico. La meningite indotta sperimentalmente è stata costantemente associata a disseminazione dei meningococchi dal cervello al sangue ed agli organi vitali, in particolare milza e fegato: a 48h dall’infezione il ceppo 93/4286ΩsiaA subisce una completa clearance in tutti gli organi interessati. Tali dati sperimentali hanno mostrato che il ceppo isogenico difettivo nel locus genico siaA esibisce una drastica attenuazione della virulenza, confermando un ruolo cruciale della capsula nella disseminazione dei meningococci nel modello murino d’infezione

Il trasportatore GltT di tipo ABC per l'L-glutammato come possibile target per lo sviluppo di vaccini.

E' noto che mutanti isogenici di N. meningitidis GltT-difettivi, alterati nella componente permeasica di un sistema di trasporto di tipo ABC per l'L-glutammato, mostrano una virulenza fortemente attenuata se confrontati con il ceppo selvatico in un modello murino di meningite meningococcica. Inoltre, i mutanti isogenici GltT-difettivi, esibiscono una ridotta capacità di crescita e sopravvivenza in un modello di infezione cellulare. In aggiunta, recenti dati di letteratura indicano che l'utilizzo dell'L-glutammato promuove la sopravvivenza meningococcica in vivo proteggendo il batterio dal burst ossidativo, confermando il ruolo centrale dell'amminoacido e del trasportatore GltT per l'istaurarsi dell'infezione sistemica. Partendo da tali presupposti abbiamo ipotizzato che le componenti del trasportatore GltT potrebbero rappresentare nuovi specifici target per lo sviluppo di vaccini contro la meningite meningococcica. In particolare, nel presente lavoro sono state selezionate le porzioni meno idrofobiche delle componenti del trasportatore GltT. I frammenti genici codificanti le suddette componenti proteiche sono stati clonati nel vettore di espressione pGEX2TK e le proteine sono state over-espresse in fusione con la Glutatione S-transferasi in cellule di E. coli BL21lambdaDE3, e purificate mediante cromatografia per affinità. Si è quindi proceduto alla produzione di anticorpi specifici diretti contro le proteine ricombinanti nell'ospite murino. I sieri immuni ottenuti sono stati testati mediante saggi ELISA ed è stata, quindi, saggiata la loro attività battericida in saggi di infezione in vitro, per correlarlo con il grado di protezione nell'uomo. L'attività battericida dei sieri ottenuti è stata valutata contro il ceppo di meningococco di sierogruppo C 93/4286, utilizzando come fonte di complemento un pool di siero di coniglio neonato (Sigma). Il titolo battericida del siero anti-GltT è riportato come il reciproco della diluizione del siero che determina la morte di oltre il 50% dei batteri

Thyroid hormone regulates glutamine metabolism and anaplerotic fluxes by inducing mitochondrial glutamate aminotransferase GPT2

: Thyroid hormones (THs) are key metabolic regulators coordinating short- and long-term energy needs. In skeletal muscle, THs modulate energy metabolism in pathophysiological conditions. Indeed, hypo- and hyperthyroidism are leading causes of muscle weakness and strength; however, the metabolic pathways underlying these effects are still poorly understood. Using molecular, biochemical, and isotope-tracing approaches combined with mass spectrometry and denervation experiments, we find that THs regulate glutamine metabolism and anaplerotic fluxes by up-regulating the glutamate pyruvate transaminase 2 (GPT2) gene. In humans, GPT2 autosomal recessive mutations cause a neurological syndrome characterized by intellectual disability, microcephaly, and progressive motor symptoms. Here, we demonstrate a role of the TH/GPT2 axis in skeletal muscle in which it regulates muscle weight and fiber diameter in resting and atrophic conditions and results in protection from muscle loss during atrophy. These results describe an anabolic route by which THs rewire glutamine metabolism toward the maintenance of muscle mass

GltT ABC-type transporter: potential candidate for development of vaccine against Neisseria meningitidis.

Introduction Neisseria meningitidis is a Gram-negative encapsulated diplococcus, the major cause of meningitis and sepsis. During the infectious cycle, N. meningitidis is able to utilize only few compounds as energy sources. In intracellular milieu, the main energy sources for meningococci are pyruvate, lactate, and certain aminoacid, such as L-glutamate. The meningococcus possesses a L-glutamate ABC-type transporter, GltT. Literature data demonstrated that the GltT transporter is required for bacterial survival within epithelial cells, and survival in human whole blood, and in a meningococcal meningitis murine model. In order to evaluate whether the GltT transporter may represent a potential candidate for vaccine development, protein components of the transporter have been induced in in vitro expression system, purified, and, after active immunization assay, specific antibodies were tested for their bactericidal activity. Materials and Methods The NMC1935 gene, encoding for the periplasmic protein of GltT transporter was cloned in expression vector. The periplasmic protein with Glutathione S-transferase (GST) tag was over-expressed in E. coli and then purified by affinity chromatography. Therefore we proceeded with specific antibodies production by active immunization assay in vivo. ELISA analysis determined the titer of immune anti-sera. To investigate the capacity for serum bactericidal killing of meningococci, serogroup C 93/4286 strain was grown with different dilution of the anti-GltT antiserum and rabbit complement. Results The anti-GltT antiserum analyzed showed a bactericidal activity of 93/4286 strain with all dilution tested compared to the control where bacteria were incubated alone or with rabbit complement. Discussion and Conclusion Although early diagnosis and antibiotic treatment enhance survival, prevention through vaccination would appear the best way to limit meningococcal disease. In this study we evaluated the GltT transporter as potential vaccinal candidate. The GltT transporter is a glutamate uptake system highly conserved between meningococcal strain, in vitro and in vivo data demonstrated that this component is essential to meningococcal infection and survival. The anti-GltT antiserum tested showed a promising bactericidal activity, passive immunization assay will be performed