Tissue transglutaminase activates integrin-linked kinase and ß-catenin in ovarian cancer.

Ovarian cancer (OC) is the most lethal gynecological cancer. OC cells have high proliferative capacity, are invasive, resist apoptosis, and tumors often display rearrangement of extracellular matrix (ECM) components, contributing to accelerated tumor progression. The multifunctional protein tissue transglutaminase (TG2) is known to be secreted in the tumor microenvironment, where it interacts with fibronectin (FN) and the cell surface receptor integrin ß1. However, the mechanistic role of TG2 in cancer cell proliferation is unknown. Here, we demonstrate that TG2 directly interacts with and facilitates the phosphorylation and activation of the integrin effector protein integrin-linked kinase (ILK) at Ser246. We show that TG2 and p-Ser246-ILK form a complex that is detectable in patient-derived OC primary cells grown on FN-coated slides. In addition, we show that coexpression of TGM2 and ILK correlates with poor clinical outcome. Mechanistically, we demonstrate that TG2-mediated ILK activation causes phosphorylation of glycogen synthase kinase-3α/ß, allowing ß-catenin nuclear translocation and transcriptional activity. Furthermore, inhibition of TG2 and ILK using small molecules, neutralizing antibodies, or shRNA-mediated knockdown blocks cell adhesion to the FN matrix, as well as the Wnt receptor response to the Wnt-3A ligand, and ultimately, cell adhesion, growth, and migration. In conclusion, we demonstrate that TG2 directly interacts with and activates ILK in OC cells and tumors and define a new mechanism that links ECM cues with ß-catenin signaling in OC. These results suggest a central role of TG2-FN-integrin clusters in ECM rearrangement and indicate that downstream effector ILK may represent a potential new therapeutic target in OC.

Tissue transglutaminase regulates ß-catenin signaling through a c-Src-dependent mechanism.

Tissue transglutaminase (TG2) is a multifunctional enzyme involved in protein cross-linking and cell adhesion to fibronectin (FN). In cancer, TG2 induces an epithelial to mesenchymal transition, contributing to metastasis. Because cadherins bind ß-catenin at cell-cell junctions, disruption of adherens junctions destabilizes cadherin-catenin complexes. The goal of the present study was to analyze whether and how TG2 interacts with and regulates ß-catenin signaling in ovarian cancer (OC) cells. We observed a significant correlation between TG2 and ß-catenin expression levels in OC cells and tumors. TG2 augmented Wnt/ß-catenin signaling, as evidenced by enhanced ß-catenin transcriptional activity, inducing transcription of target genes cyclin D1 and c-Myc. By promoting integrin-mediated cell adhesion to FN, TG2 physically associates with and recruits c-Src, which in turn phosphorylates ß-catenin at Tyr(654), releasing it from E-cadherin and rendering it available for transcriptional regulation. By interacting with FN and enhancing ß-catenin signaling, complexed TG2 stimulates OC cell proliferation. In summary, our data demonstrate that TG2 regulates ß-catenin expression and function in OC cells and define the c-Src-dependent mechanism through which this occurs.

Transglutaminase 2 and phospholipase A2 interactions in the inflammatory response in human Thp-1 monocytes.

Several experimental approaches have demonstrated that transglutaminase 2 (TG2) increased activity is involved in monocyte activation and inflammatory response. Preliminary results also demonstrate a TG-mediated post-translational modification of phospholipase A2 (PLA2), which catalyzes the release of arachidonic acid from its lipid storage sites. The control of PLA2-mediated production of eicosanoids has been found to be of great benefit for inflammatory disease treatment. However, the identification of the mechanisms of PLA2 activation is a very complex issue, because of the presence of multiple PLA2 forms. The aim of this study was to characterize the interactions between TG2 and sPLA2 in LPS-stimulated THP-1 cells, which were treated with TPA to induce early differentiated macrophage-type model. We demonstrated that increases in TG2 enzyme activity and protein expression may be considered an early event in monocyte/macrophage activation by LPS. Under these conditions, TG2 protein was co-immunoprecipitated with PLA2 by monoclonal antibody directed against the secretory form of the enzyme (sPLA2-V). Concomitantly, the PLA2 enzyme activity increased in TPA-treated cells exposed to LPS; these high levels of enzyme activity were significant reduced by R283, a site-specific inhibitor of TG2. Moreover, confocal laser scanning microscopy analysis of double-immunostained cytochemical specimens confirmed a co-localization of BAPA-labeled proteins and sPLA2-V in LPS-treated cells. These findings give evidence of a complex TG2/sPLA2-V, suggesting the possibility that sPLA2-V is a substrate for TG2. These results demonstrated that TG2 increases produced a sustained activation of PLA2 activity, suggesting a functional interaction between these enzymes in the regulation of inflammatory response.

Integrin-linked kinase-frizzled 7 interaction maintains cancer stem cells to drive platinum resistance in ovarian cancer.

Background: Platinum-based chemotherapy regimens are a mainstay in the management of ovarian cancer (OC), but emergence of chemoresistance poses a significant clinical challenge. The persistence of ovarian cancer stem cells (OCSCs) at the end of primary treatment contributes to disease recurrence. Here, we hypothesized that the extracellular matrix protects CSCs during chemotherapy and supports their tumorigenic functions by activating integrin-linked kinase (ILK), a key enzyme in drug resistance. Methods: TCGA datasets and OC models were investigated using an integrated proteomic and gene expression analysis and examined ILK for correlations with chemoresistance pathways and clinical outcomes. Canonical Wnt pathway components, pro-survival signaling, and stemness were examined using OC models. To investigate the role of ILK in the OCSC-phenotype, a novel pharmacological inhibitor of ILK in combination with carboplatin was utilized in vitro and in vivo OC models. Results: In response to increased fibronectin (FN) secretion and integrin ß1 clustering, aberrant ILK activation supported the OCSC phenotype, contributing to OC spheroid proliferation and reduced response to platinum treatment. Complexes formed by ILK with the Wnt receptor frizzled 7 (Fzd7) were detected in tumors and showed a strong correlation with metastatic progression. Moreover, TCGA datasets confirmed that combined expression of ILK and Fzd7 in high grade serous ovarian tumors is correlated with reduced response to chemotherapy and poor patient outcomes. Mechanistically, interaction of ILK with Fzd7 increased the response to Wnt ligands, thereby amplifying the stemness-associated Wnt/ß-catenin signaling. Notably, preclinical studies showed that the novel ILK inhibitor compound 22 (cpd-22) alone disrupted ILK interaction with Fzd7 and CSC proliferation as spheroids. Furthermore, when combined with carboplatin, this disruption led to sustained AKT inhibition, apoptotic damage in OCSCs and reduced tumorigenicity in mice. Conclusions: This "outside-in" signaling mechanism is potentially actionable, and combined targeting of ILK-Fzd7 may represent a new therapeutic strategy to eradicate OCSCs and improve patient outcomes.

Integrin-linked kinase-frizzled 7 interaction maintains cancer stem cells to drive platinum resistance in ovarian cancer.

BACKGROUND: Platinum-based chemotherapy regimens are a mainstay in the management of ovarian cancer (OC), but emergence of chemoresistance poses a significant clinical challenge. The persistence of ovarian cancer stem cells (OCSCs) at the end of primary treatment contributes to disease recurrence. Here, we hypothesized that the extracellular matrix protects CSCs during chemotherapy and supports their tumorigenic functions by activating integrin-linked kinase (ILK), a key enzyme in drug resistance. METHODS: TCGA datasets and OC models were investigated using an integrated proteomic and gene expression analysis and examined ILK for correlations with chemoresistance pathways and clinical outcomes. Canonical Wnt pathway components, pro-survival signaling, and stemness were examined using OC models. To investigate the role of ILK in the OCSC-phenotype, a novel pharmacological inhibitor of ILK in combination with carboplatin was utilized in vitro and in vivo OC models. RESULTS: In response to increased fibronectin secretion and integrin ß1 clustering, aberrant ILK activation supported the OCSC phenotype, contributing to OC spheroid proliferation and reduced response to platinum treatment. Complexes formed by ILK with the Wnt receptor frizzled 7 (Fzd7) were detected in tumors and correlated with metastatic progression. Moreover, TCGA datasets confirmed that combined expression of ILK and Fzd7 in high grade serous ovarian tumors is correlated with reduced response to chemotherapy and poor patient outcomes. Mechanistically, interaction of ILK with Fzd7 increased the response to Wnt ligands, thereby amplifying the stemness-associated Wnt/ß-catenin signaling. Notably, preclinical studies showed that the novel ILK inhibitor compound 22 (cpd-22) alone disrupted ILK interaction with Fzd7 and CSC proliferation as spheroids. Furthermore, when combined with carboplatin, this disruption led to sustained AKT inhibition, apoptotic damage in OCSCs and reduced tumorigenicity in mice. CONCLUSIONS: This "outside-in" signaling mechanism is potentially actionable, and combined targeting of ILK-Fzd7 may lead to new therapeutic approaches to eradicate OCSCs and improve patient outcomes.

Transglutaminase 2 interaction with small heat shock proteins mediate cell survival upon excitotoxic stress.

Transglutaminase 2 has been postulated to be involved in the pathogenesis of central nervous system neurodegenerative disorders. However, its role in neuronal cell death remains to be elucidated. Excitotoxicity is a common event underlying neurodegeneration. We aimed to evaluate the protein targets for transglutaminase 2 in cell response to NMDA-induced excitotoxic stress, using SH-SY5Y neuroblastoma cells which express high tranglutaminase 2 levels upon retinoic acid-driven differentiation toward neurons. NMDA-evoked calcium increase led to transglutaminase 2 activation that mediated cell survival, as at first suggested by the exacerbation of NMDA toxicity in the presence of R283, a synthetic competitive inhibitor of transglutaminase active site. Assays of R283-mediated transglutaminase inhibition showed the involvement of enzyme activity in NMDA-induced reduction in protein basal levels of pro-apoptotic caspase-3 and the stress protein Hsp20. However, this occurred in a way different from protein cross-linking, given that macromolecular assemblies were not observed in our experimental conditions for both proteins. Co-immunoprecipitation experiments provided evidence for the interaction, in basal conditions, between transglutaminase 2 and Hsp20, as well as between Hsp20 and Hsp27, a major anti-apoptotic protein promoting caspase-3 inactivation and degradation. NMDA treatment disrupted both these interactions that were restored upon transglutaminase 2 inhibition with R283. These results suggest that transglutaminase 2 might be protective against NMDA-evoked excitotoxic insult in neuronal-like SH-SY5Y cells in a way, independent from transamidation that likely involves its interaction with the complex Hsp20/Hsp27 playing a pro-survival role.

Protective effects of zonisamide against rotenone-induced neurotoxicity.

Zonisamide (ZNS), an antiepileptic drug having beneficial effects also against Parkinson's disease symptoms, has proven to display an antioxidant effects in different experimental models. In the present study, the effects of ZNS on rotenone-induced cell injury were investigated in human neuroblastoma SH-SY5Y cells differentiated towards a neuronal phenotype. Cell cultures were exposed for 24 h to 500 nM rotenone with or without pre-treatment with 10-100 µM ZNS. Then, the following parameters were analyzed: (a) cell viability; (b) intracellular reactive oxygen species production; (c) mitochondrial transmembrane potential; (d) cell necrosis and apoptosis; (e) caspase-3 activity. ZNS dose-dependently suppressed rotenone-induced cell damage through a decrease in intracellular ROS production, and restoring mitochondrial membrane potential. Similarly to ZNS effects, the treatment with N-acetyl-cysteine (100 µM) displayed significant protective effects against rotenone-induced ROS production and Δψm at 4 and 12 h respectively, reaching the maximal extent at 24 h. Additionally, ZNS displayed antiapoptotic effects, as demonstrated by flow cytometric analysis of annexin V/propidium iodide double staining, and significant attenuated rotenone-increased caspase 3 activity. On the whole, these findings suggest that ZNS preserves mitochondrial functions and counteracts apoptotic signalling mechanisms mainly by an antioxidant action. Thus, ZNS might have beneficial effect against neuronal cell degeneration in different experimental models involving mitochondrial dysfunction.

Effects of low intensity static magnetic field on FTIR spectra and ROS production in SH-SY5Y neuronal-like cells.

Biological effects of man-made electromagnetic fields (EMFs) have been studied so far by experimental approaches exposing animals and cell cultures to EMFs. However, the evidence for cell toxicity induced by static magnetic field (SMF) is still uncertain. We investigated the effects produced by the exposure of human SH-SY5Y neuronal-like cells to a uniform magnetic field at intensities of 2.2 mT, which is less than the recommended public exposure limits set by the International Commission on Non-Ionizing Radiation Protection (ICNIRP). A decrease of membrane mitochondrial potential up to 30% was measured after 24 h of exposure to SMF in SH-SY5Y cells, and this effect was associated with reactive oxygen species production increase. Fourier transform infrared spectroscopy (FTIR) analysis showed that exposure to a static magnetic intensity around 2.2 mT changed the secondary structure of cellular proteins and lipid components. The vibration bands relative to the methylene group increased significantly after 4 h of exposure, whereas further exposure up to 24 h produced evident shifts of amide I and II modes and a relative increase in ß-sheet contents with respect to α-helix components. Our study demonstrated that a moderate SMF causes alteration in cell homeostasis, as indicated by FTIR spectroscopy observations of changes in protein structures that are part of cell response to magnetic field exposure.

Characterizing the regulatory Fas (CD95) epitope critical for agonist antibody targeting and CAR-T bystander function in ovarian cancer.

Receptor clustering is the most critical step to activate extrinsic apoptosis by death receptors belonging to the TNF superfamily. Although clinically unsuccessful, using agonist antibodies, the death receptors-5 remains extensively studied from a cancer therapeutics perspective. However, despite its regulatory role and elevated function in ovarian and other solid tumors, another tumor-enriched death receptor called Fas (CD95) remained undervalued in cancer immunotherapy until recently, when its role in off-target tumor killing by CAR-T therapies was imperative. By comprehensively analyzing structure studies in the context of the binding epitope of FasL and various preclinical Fas agonist antibodies, we characterize a highly significant patch of positively charged residue epitope (PPCR) in its cysteine-rich domain 2 of Fas. PPCR engagement is indispensable for superior Fas agonist signaling and CAR-T bystander function in ovarian tumor models. A single-point mutation in FasL or Fas that interferes with the PPCR engagement inhibited apoptotic signaling in tumor cells and T cells. Furthermore, considering that clinical and immunological features of the autoimmune lymphoproliferative syndrome (ALPS) are directly attributed to homozygous mutations in FasL, we reveal differential mechanistic details of FasL/Fas clustering at the PPCR interface compared to described ALPS mutations. As Fas-mediated bystander killing remains vital to the success of CAR-T therapies in tumors, our findings highlight the therapeutic analytical design for potentially effective Fas-targeting strategies using death agonism to improve cancer immunotherapy in ovarian and other solid tumors.

Monitoring of transglutaminase 2 under different oxidative stress conditions.

Transglutaminase 2 (TG2) is a multifunctional calcium-dependent enzyme which catalyzes the post-translational protein crosslinking with formation of intra- or inter-molecular epsilon(gamma-glutamyl)lysine bonds or polyamine incorporation. The up-regulation and activation of TG2 have been reported in a variety of physiological events, including cell differentiation, signal transduction, apoptosis, and wound healing, as well as in cell response to stress evoked by different internal and external stimuli. Here we review TG2 role in cell response to redox state imbalance both under physiological and pathological conditions, such as neurodegenerative disorders, inflammation, autoimmune diseases and cataractogenesis, in which oxidative stress plays a pathogenetic role and also accelerates disease progression. The increase in TG activity together with mitochondrial impairment and collapse of antioxidant enzymatic cell defences have been reported to be the prominent biochemical alterations becoming evident prior to neurodegeneration. Moreover, oxidative stress-induced TG2 pathway is involved in autophagy inhibition and aggresome formation, and TG2 has been suggested to function as a link between oxidative stress and inflammation by driving the decision as to whether a protein should undergo SUMO-mediated regulation or proteasomal degradation. Literature data suggest a strong association between oxidative stress and TG2 up-regulation, which in turn may result in cell survival or apoptosis, depending on cell type, kind of stressor, duration of insult, as well as TG2 intracellular localization and activity state. In particular, it may be suggested that TG2 plays a pro-survival role when the alteration of cell redox state homeostasis is not associated with intracellular calcium increase triggering TG2 transamidation activity.

Protective effects of agmatine in rotenone-induced damage of human SH-SY5Y neuroblastoma cells: fourier transform infrared spectroscopy analysis in a model of Parkinson's disease.

Agmatine is a novel neuromodulator that plays a protective role in the CNS in several models of cellular damage. However, the mechanisms involved in these protective effects in neurodegenerative diseases are poorly understood. Fourier transform infrared (FTIR) spectroscopy analysis detects biomolecular changes in disordered cells and tissues. In this report, we utilize FTIR spectroscopy to characterize the changes in rotenone-induced damage in neuronal-like differentiated SH-SY5Y neuroblastoma cells in the presence or absence of agmatine. The analysis of the FTIR spectra demonstrates significant alterations in rotenone-treated cells, whereas the FTIR spectra obtained after pre-incubation with agmatine (250 nM) significantly reduces these redox alterations and more closely resembles those of the control cells. In particular, rotenone-damaged cells demonstrate spectral alterations related to amide I, which correspond to an increase in ß-sheet components, and decreases in the amide II absorption intensity, suggesting a loss of N-H bending and C-N stretching. These alterations were also evident by Fourier self-deconvolution analysis. Thus, rotenone-induced increases in the levels of stretching vibration band related to the protein carboxyl group would account for a significant amount of misfolded proteins in the cell. Agmatine effectively reduces these effects of rotenone on protein structure. In conclusion, antioxidant and scavenging properties of agmatine reduce rotenone-produced cellular damage at the level of protein structure. These, together with other previous observations, demonstrate the therapeutic potential of agmatine in the treatment of Parkinson's disease.

The Outside-In Journey of Tissue Transglutaminase in Cancer.

Tissue transglutaminase (TG2) is a member of the transglutaminase family that catalyzes Ca2+-dependent protein crosslinks and hydrolyzes guanosine 5'-triphosphate (GTP). The conformation and functions of TG2 are regulated by Ca2+ and GTP levels; the TG2 enzymatically active open conformation is modulated by high Ca2+ concentrations, while high intracellular GTP promotes the closed conformation, with inhibition of the TG-ase activity. TG2's unique characteristics and its ubiquitous distribution in the intracellular compartment, coupled with its secretion in the extracellular matrix, contribute to modulate the functions of the protein. Its aberrant expression has been observed in several cancer types where it was linked to metastatic progression, resistance to chemotherapy, stemness, and worse clinical outcomes. The N-terminal domain of TG2 binds to the 42 kDa gelatin-binding domain of fibronectin with high affinity, facilitating the formation of a complex with ß-integrins, essential for cellular adhesion to the matrix. This mechanism allows TG2 to interact with key matrix proteins and to regulate epithelial to mesenchymal transition and stemness. Here, we highlight the current knowledge on TG2 involvement in cancer, focusing on its roles translating extracellular cues into activation of oncogenic programs. Improved understanding of these mechanisms could lead to new therapeutic strategies targeting this multi-functional protein.

Agmatine effects on mitochondrial membrane potential and NF-κB activation protect against rotenone-induced cell damage in human neuronal-like SH-SY5Y cells.

Agmatine, an endogenous arginine metabolite, has been proposed as a novel neuromodulator that plays protective roles in the CNS in several models of cellular damage. However, the mechanisms involved in these protective effects in neurodegenerative diseases are poorly understood. The present study was undertaken to investigate the effects of agmatine on cell injury induced by rotenone, commonly used in establishing in vivo and in vitro models of Parkinson's disease, in human-derived dopaminergic neuroblastoma cell line (SH-SY5Y). We report that agmatine dose-dependently suppressed rotenone-induced cellular injury through a reduction of oxidative stress. Similar effects were obtained by spermine, suggesting a scavenging effect for these compounds. However, unlike spermine, agmatine also prevented rotenone-induced nuclear factor-κB nuclear translocation and mitochondrial membrane potential dissipation. Furthermore, rotenone-induced increase in apoptotic markers, such as caspase 3 activity, Bax expression and cytochrome c release, was significantly attenuated with agmatine treatment. These findings demonstrate mitochondrial preservation with agmatine in a rotenone model of apoptotic cell death, and that the neuroprotective action of agmatine appears because of suppressing apoptotic signalling mechanisms. Thus, agmatine may have therapeutic potential in the treatment of Parkinson's disease by protecting dopaminergic neurons.

Frizzled-7 Identifies Platinum-Tolerant Ovarian Cancer Cells Susceptible to Ferroptosis.

Defining traits of platinum-tolerant cancer cells could expose new treatment vulnerabilities. Here, new markers associated with platinum-tolerant cells and tumors were identified using in vitro and in vivo ovarian cancer models treated repetitively with carboplatin and validated in human specimens. Platinum-tolerant cells and tumors were enriched in ALDH+ cells, formed more spheroids, and expressed increased levels of stemness-related transcription factors compared with parental cells. Additionally, platinum-tolerant cells and tumors exhibited expression of the Wnt receptor Frizzled-7 (FZD7). Knockdown of FZD7 improved sensitivity to platinum, decreased spheroid formation, and delayed tumor initiation. The molecular signature distinguishing FZD7+ from FZD7- cells included epithelial-to-mesenchymal (EMT), stemness, and oxidative phosphorylation-enriched gene sets. Overexpression of FZD7 activated the oncogenic factor Tp63, driving upregulation of glutathione metabolism pathways, including glutathione peroxidase 4 (GPX4), which protected cells from chemotherapy-induced oxidative stress. FZD7+ platinum-tolerant ovarian cancer cells were more sensitive and underwent ferroptosis after treatment with GPX4 inhibitors. FZD7, Tp63, and glutathione metabolism gene sets were strongly correlated in the ovarian cancer Tumor Cancer Genome Atlas (TCGA) database and in residual human ovarian cancer specimens after chemotherapy. These results support the existence of a platinum-tolerant cell population with partial cancer stem cell features, characterized by FZD7 expression and dependent on the FZD7-ß-catenin-Tp63-GPX4 pathway for survival. The findings reveal a novel therapeutic vulnerability of platinum-tolerant cancer cells and provide new insight into a potential "persister cancer cell" phenotype. SIGNIFICANCE: Frizzled-7 marks platinum-tolerant cancer cells harboring stemness features and altered glutathione metabolism that depend on GPX4 for survival and are highly susceptible to ferroptosis.

Critical role of transglutaminase and other stress proteins during neurodegenerative processes.

Proteolytic stress, resulting from the intracellular accumulation of misfolded or aggregated proteins, which exceed the capacity of the ubiquitin-proteasome system to degrade them, plays a relevant role in neurodegenerative disorders, such as Alzheimer's disease, Parkinson's disease, and Huntington's chorea. Most of toxic protein aggregates are characterised by the presence of isopeptide bonds (cross-links) catalysed by transglutaminase activity; further, several disease-specific proteins-tau, amyloid-beta, alpha-synuclein, huntingtin-are in vitro and/or in vivo substrates of transglutaminase 2. These findings suggest an important role for transglutaminase 2-mediated cross-linking reactions in neurodegeneration. Therefore, the use of transglutaminase activity inhibitors could ameliorate neuronal cell death. New therapeutic perspectives also arise from the possibility to prevent or reduce protein aggregation by enhancing the activation of heat shock proteins, which have been shown to be potent suppressors of neurodegeneration in cell cultures/animal models. Interestingly, some heat shock proteins have been shown to be in vitro or in vivo cross-linked by transglutaminase 2. These observations seem to suggest that transglutaminase activity could be involved in the stabilization of intracellular protein aggregates by interfering with proteasomal degradation of misfolded proteins. Further studies are needed to validate leading hypotheses and to open new prospects for developing therapeutic tools.

Transglutaminase 2 silencing reduced the beta-amyloid-effects on the activation of human THP-1 cells.

The aberrant expression and activation of transglutaminase 2 (TG2), the ubiquitous enzyme which catalyzes calcium-dependent protein cross-linking reactions, has been reported in many inflammatory diseases. Chronic inflammation, mediated by prolonged activation of brain-resident immunocompetent cells, appears to be involved in the pathogenesis of several age-related diseases, such as Alzheimer's disease. Given that increased TG2 expression has been observed in AD brains, this study was aimed to characterize the role of TG2 in THP-1 monocytes stimulated with amyloid-beta (Aß). Aß1-42 treatment dose-dependently increased TG2 expression in THP-1 cells. In particular, a fivefold up-regulation of TG2, compared with control cells, was observed in the presence of 0.5 µM Aß1-42. At the same concentration, Aß1-42 was able to promote monocyte maturation as suggested by increased expression of the cell surface antigen CD14 as well as the adhesion-promoting factor fibronectin. The stimulation of THP-1 cells with Aß1-42 also led to a significant up-regulation of tumor necrosis factor α (TNF-α) and matrix metalloproteinase 9 (MMP-9). Interestingly, THP-1 cell transfection with small interfering RNA directed against TG2 was able to reduce Aß1-42 increased levels of all the examined markers of monocyte maturation (CD14, fibronectin), and activation (TNF-α, MMP-9). These results indicate that TG2 up-regulation is required for the functional THP-1 monocyte activation induced by Aß1-42. This work suggests that TG2 inhibition may represent a therapeutic target to ameliorate the inflammation and progression in Alzheimer's disease.

Spontaneous neurocognitive retrieval of patients with sub-acute ischemic stroke is associated with dietary protein intake.

OBJECTIVE: To document the relationship between neurocognitive recovery and macronutrient intake of patients suffering from ischemic strokes. DESIGN: Thirty day prospective study of 17 patients suffering from sub-acute stroke (> 14 days from the index event; 10 males, 7 females; mean age 75 +/- 8 years) admitted to our rehabilitation unit. RESULTS: At admission (ADM), mean energy intake was inadequate (< 24 kcal/kg) for bodily needs, whereas protein (> 0.8 g/kg) and lipid (> 0.7 g/kg) intake was appropriate. Patients were moderately deficient for neurological (NIHSS 10.3 +/- 3.5) and cognitive tests (MMSE 22.5 +/- 3.3). NIHSS correlated negatively with proteins (r = -0.47, P = 0.05 at ADM; r = -0.52, P = 0.03 at 30 days) and positively with carbohydrate/protein ratio (CHO/protein; r = +0.45, P = 0.06 at ADM; r = +0.48, P = 0.05 at 30 days). However, MMSE correlated positively with proteins (r = +0.77, P = 0.0003 at ADM; r = +0.55, P = 0.02 at 30 days) and negatively with (CHO/Prot; r = -0.57, P = 0.02 at ADM; not significant at 30 days). The relationship remained significant even when the data at ADM and at 30 days where pooled. CONCLUSIONS: In sub-acute strokes, patient neurological and cognitive retrieval could positively be associated with protein intake.

The relationship between the xenotransplantation limitations and Charles Darwin theory on human evolution.

The need for an alternative source of donor organs, together with the expansion of scientific data in this field, has focused attention on xenotransplantation as a possible alternative to allotransplantation in the treatment of patients with end-stage disease of vital organs. The fundamental property called 'evolution' was not discovered in the study of living matter by Darwin, but in the study of the foundations of the Logic of Nature, i.e. in first level Galilean Science. The work of Darwin was aiming at the discovery of the origin of the human species and the property of living matter called 'evolution' was intended to prove what the origin was of the human species. In this letter to editor we present our opinion about the relationship between the xenotransplantation limitations and Charles Darwin theory on Human evolution.

MutSignatures: an R package for extraction and analysis of cancer mutational signatures.

Cancer cells accumulate somatic mutations as result of DNA damage, inaccurate repair and other mechanisms. Different genetic instability processes result in characteristic non-random patterns of DNA mutations, also known as mutational signatures. We developed mutSignatures, an integrated R-based computational framework aimed at deciphering DNA mutational signatures. Our software provides advanced functions for importing DNA variants, computing mutation types, and extracting mutational signatures via non-negative matrix factorization. Specifically, mutSignatures accepts multiple types of input data, is compatible with non-human genomes, and supports the analysis of non-standard mutation types, such as tetra-nucleotide mutation types. We applied mutSignatures to analyze somatic mutations found in smoking-related cancer datasets. We characterized mutational signatures that were consistent with those reported before in independent investigations. Our work demonstrates that selected mutational signatures correlated with specific clinical and molecular features across different cancer types, and revealed complementarity of specific mutational patterns that has not previously been identified. In conclusion, we propose mutSignatures as a powerful open-source tool for detecting the molecular determinants of cancer and gathering insights into cancer biology and treatment.

Expression pattern of transglutaminases in the early differentiation stage of erupting rat incisor.

Several studies demonstrated that transglutaminases play a key role in extracellular matrix stabilization needed for cell differentiation. We evaluated transglutaminase expression and activity in the pre-secretory stage of differentiation of the continuously erupting rat incisor. We observed that transglutaminase-mediated incorporation of monodansylcadaverine into protein substrates was specifically located in the apical loop, and along the basement membrane joining mesenchyme and inner dental epithelium in the odontogenic organ. Enzyme activity was associated with mRNAs for transglutaminase 1 and 2. Notably, labelling cells for these isoenzymes were observed in both mesenchymal and epithelial compartments, but not in the basement membrane, in the ameloblast facing pulp anterior region, where ameloblast and odontoblast differentiation begins. These findings demonstrate that transglutaminase 1 and transglutaminase 2 are expressed at a major extent in the pre-secretory stage of regenerating rat incisor, where they probably play complementary roles in cell signalling between mesenchyme and epithelium and extracellular matrix.