Whole Genome Expression Analyses of miRNAs and mRNAs Suggest the Involvement of miR-320a and miR-155-3p and their Targeted Genes in Lithium Response in Bipolar Disorder.

Lithium is the mainstay in the maintenance of bipolar disorder (BD) and the most efficacious pharmacological treatment in suicide prevention. Nevertheless, its use is hampered by a high interindividual variability and important side effects. Genetic and epigenetic factors have been suggested to modulate lithium response, but findings so far have not allowed identifying molecular targets with predictive value. In this study we used next generation sequencing to measure genome-wide miRNA expression in lymphoblastoid cell lines from BD patients excellent responders (ER, n = 12) and non-responders (NR, n = 12) to lithium. These data were integrated with microarray genome-wide expression data to identify pairs of miRNA/mRNA inversely and significantly correlated. Significant pairs were prioritized based on strength of association and in-silico miRNA target prediction analyses to select candidates for validation with qRT-PCR. Thirty-one miRNAs were differentially expressed in ER vs. NR and inversely correlated with 418 genes differentially expressed between the two groups. A total of 331 of these correlations were also predicted by in-silico algorithms. miR-320a and miR-155-3p, as well as three of their targeted genes (CAPNS1 (Calpain Small Subunit 1) and RGS16 (Regulator of G Protein Signaling 16) for miR-320, SP4 (Sp4 Transcription Factor) for miR-155-3p) were validated. These miRNAs and mRNAs were previously implicated in psychiatric disorders (miR-320a and SP4), key processes of the central nervous system (CAPNS1, RGS16, SP4) or pathways involved in mental illnesses (miR-155-3p). Using an integrated approach, we identified miRNAs and their targeted genes potentially involved in lithium response in BD.

Interactions between killer immunoglobulin-like receptors and their human leucocyte antigen Class I ligands influence the outcome of unrelated haematopoietic stem cell transplantation for thalassaemia: a novel predictive algorithm.

In a study conducted on 114 patients undergoing unrelated donor haematopoietic stem cell transplantation (HSCT) for thalassaemia, we observed that the lack of activating killer immunoglobulin-like receptors (KIRs) on donor natural killer (NK) cells significantly increased the risk of graft-versus-host disease (GvHD) [hazard risk (HR) 4.2, 95% confidence interval (CI) 1.7-10.1, P = 0.002] and transplantation-related mortality (HR 4.7, 95% CI 1.6-14.2, P = 0.01). The risk of GvHD furthermore increased when recipients heterozygous for HLA-C KIR ligand groups (C1/C2) were transplanted from donors completely lacking activating KIRs (HR 6.1, 95% CI 1.9-19.2, P = 0.002). We also found that the risk of rejection was highest when the recipient was homozygous for the C2 HLA-KIR ligand group and the donor carried two or more activating KIRs (HR 6.8, 95% CI 1.9-24.4, P = 0.005). By interpolating the number of donor activating KIRs with recipient HLA-C KIR ligands, we created an algorithm capable of stratifying patients according to the immunogenetic risk of complications following unrelated HSCT. In clinical practice, this predictive tool could serve as an important supplement to clinical judgement and decision-making.

VAPB ER-Aggregates, A Possible New Biomarker in ALS Pathology.

A point mutation (P56S) in the gene-encoding vesicle-associated membrane-protein-associated protein B (VAPB) leads to an autosomal-dominant form of amyotrophic lateral sclerosis (ALS), classified as ALS-8. The mutant VAPB is characterized by ER-associated aggregates that lead to a complete reorganization of ER structures. Growing evidences suggest VAPB involvement in ALS pathomechanisms. In fact, numerous studies demonstrated VAPB alteration also in sporadic ALS (sALS) and showed the presence of its aggregates when others ALS-related gene are mutant. Recently, the identification of new biomarkers in peripheral blood mononuclear cells (PBMCs) has been proposed as a good noninvasive option for studying ALS. Here, we evaluated VAPB as a possible ALS pathologic marker analyzing PBMCs of sALS patients. Immunofluorescence analysis (IFA) showed a peculiar pattern of VAPB aggregates in sALS, not evident in healthy control (HC) subjects and in Parkinson's disease (PD) PBMCs. This specific pattern led us to suppose that VAPB could be misfolded in sALS. The data indirectly confirmed by flow cytometry assay (FCA) showed a reduction of VAPB fluorescent signals in sALS. However, our observations were not associated with the presence of a genetic mutation or altered gene expression of VAPB. Our study brings further evidences of the VAPB role in ALS as a diagnostic biomarker.

Mitochondria morphology and DNA content upon sublethal exposure to beta-amyloid(1-42) peptide.

Brains affected by Alzheimer's disease (AD) show a large spectrum of mitochondrial alterations at both morphological and genetic level. The causal link between amyloid beta peptides (AP) and mitochondrial dysfunction has been established in cellular models of AD using Abeta concentrations capable of triggering massive neuronal death. However, mitochondrial changes related to sublethal exposure to Abeta are less known. Here we show that subtoxic, 1 microM Abeta(1-42) exposure does not change the mitochondrial shape of living cells, as visualized upon the uptake of the non-potentiometric fluorescent probe Mitotracker Green and enhanced yellow fluorescent protein (EYFP)-tagged cytochrome c oxidase expression. Immunolocalization of oxidative adducts 8-hydroxy-2'-deoxyguanosine, 8-hydroxyguanine and 8-hydroxyguanosine demonstrates that one-micromolar concentration of Abeta(1-42) is also not sufficient to elicit dramatic qualitative changes in the RNA/DNA oxidative products. However, in comparison with controls, semi-quantitative analysis of the overall mitochondrial mass by integrated fluorescence intensity reveals an ongoing down-regulation in mitochondrial biosynthesis or, conversely, an enhanced autophagic demise of Abeta treated cells. Furthermore, a significant increase of the full-length mitochondrial DNA (mtDNA) from Abeta-treated versus control cells is found, as measured by long range polymerase chain reaction (PCR). Such up-regulation is accompanied by extensive fragmentation of the unamplified mtDNA, probably due to the detrimental effect of Abeta. We interpret these results as a sequence of compensatory responses induced by mtDNA damage, which are devoted to repression of oxidative burst. In conclusion, our findings suggest that early therapeutic interventions aimed at prevention of mitochondrial oxidative damage may delay AD progression and help in treating AD patients.

Homozygosity for killer immunoglobin-like receptor haplotype A predicts complete molecular response to treatment with tyrosine kinase inhibitors in chronic myeloid leukemia patients.

Several recent reports suggest a possible role for killer immunoglobulin-like receptors (KIR) in the onset of chronic myeloid leukemia (CML) and response to therapy with tyrosine kinase inhibitors (TKIs). To explore this hypothesis, we studied KIRs and their human leukocyte antigen class I ligands in 59 consecutive patients with chronic-phase CML (mean age, 53 years; range, 23-81 years) and a group of 121 healthy control participants belonging to the same ethnic group as the patients. The 2-year cumulative incidence of complete molecular response, obtained after a median of 27 months (range, 4-52 months), was 51.2%. An increased frequency of the activating receptor KIR2DS1 (pm = 0.05) and a reduced frequency of the KIR-ligand combination KIR2DS2/2DL2 absent/C1 present (pm = 0.001) were significantly associated with CML. Moreover, KIR repertoires in patients appeared to influence response to TKI therapy. Homozygosity for KIR haplotype A (pm = 0.01), a decreased frequency of the inhibitory KIR gene KIR2DL2 (pm = 0.02), and low numbers of inhibitory KIR genes (pm = 0.05) were all significantly associated with achievement of complete molecular remission. These data suggest that a decrease in properly stimulated and activated NK cells might contribute to the occurrence of CML and indicate homozygosity for KIR haplotype A as a promising immunogenetic marker of complete molecular response that could help clinicians decide whether to withdraw treatment in patients with CML.

Absence of activating killer immunoglobulin-like receptor genes combined with hepatitis C viral genotype is predictive of hepatocellular carcinoma.

Killer immunoglobulin-like receptors and their human leukocyte antigen class I ligands have a critical role in natural killer cell response to viral pathogens and tumors. To investigate whether killer immunoglobulin-like receptor genes could influence the chronic course of hepatitis C virus infection and/or progression to hepatocellular carcinoma we retrospectively analyzed a cohort of 228 patients transplanted for hepatitis C virus-induced cirrhotic end stage liver disease, combined or not with hepatocellular carcinoma. We found that patients completely lacking activating killer immunoglobulin-like receptor genes had a high risk of developing hepatocellular carcinoma. Hepatitis C viral genotype and viral load are other risk factors that can influence the course of chronic hepatitis C virus infection. In our study, the risk conferred by hepatitis C viral genotypes was enhanced in patients lacking activating killer immunoglobulin-like receptors. These results point to an important role for activating killer immunoglobulin-like receptors in the control of hepatitis C virus infection and progression to hepatocellular carcinoma. In clinical practice, assessment of killer immunoglobulin-like receptor and hepatitis C viral genotype combinations should allow for more accurate monitoring of patients with chronic hepatitis C virus infection.

Recipient CTLA-4*CT60-AA genotype is a prognostic factor for acute graft-versus-host disease in hematopoietic stem cell transplantation for thalassemia.

Polymorphisms of the cytotoxic T-lymphocyte antigen-4 gene (CTLA-4) have been associated with autoimmune diseases and it has recently been reported that donor genotypes correlate with the outcome of allogeneic hematopoietic stem cell transplantation in leukemia patients. With the aim of confirming this finding in thalassemia patients, we investigated the influence of genotype distribution of 3 CTLA-4 gene polymorphisms in 72 thalassemia patients and their unrelated donors. A significant association was observed for recipient CT60-AA genotype and onset of grade II-IV (63.2% vs 24.5%; p = 0.001) and grade III-IV (36.4% vs 7.6%; p = 0.005) acute graft-versus-host disease (aGVHD). The same association was observed for the 88-base-pair allele of the CTLA-4 (AT)n polymorphism, which was determined to be in complete linkage disequilibrium with the CT60 A allele. Multinomial Cox regression demonstrated that this association was independent of CT60 donor genotypes or other risk factors (p = 0.016; hazard ratio = 2.8). Our data confirm that the genetic variability in CTLA-4 is an important prognostic factor for aGVHD and suggest that some of the risk factors for this complication are generated by recipient cells that persist after the myeloablative conditioning regimen.

Status of donor-recipient HLA class I ligands and not the KIR genotype is predictive for the outcome of unrelated hematopoietic stem cell transplantation in beta-thalassemia patients.

Several studies have investigated the role played by killer immunoglobulin-like receptors (KIRs) and their ligands on the outcome of hematopoietic stem cell transplantation (HSCT) in patients affected by oncohematologic diseases. However, the interpretation of the results of these studies is considerably hampered by the heterogeneity of the diseases, disease status at transplantation, and the different protocols employed for both conditioning and graft-versus-host disease (GVHD) prophylaxis. To better define the role of KIRs in HSCT, we studied KIR genotypes and HLA class I ligands in a homogeneous group of 45 thalassemia patients transplanted with bone marrow cells from an HLA-identical, unrelated donor. Patients that were heterozygotes for HLA-Cw groups 1 (HLA-Cw(Asn80)) and 2 (HLA-Cw(Lys80)) had a higher risk of developing acute GVHD than C1/C1 or C2/C2 homozygotes (relative risk [RR] = 8.75; 95% confidence interval [CI]: 1.63-46.76; P = .007). Vice versa, all patients who experienced primary/secondary graft failure were C1/C1 or C2/C2 homozygotes (RR = 20.45; 95% CI = 1.08-384.24; P = .009). Moreover, the presence of the HLA-A11 antigen conferred protection against GVHD (0% versus 35%, P = .02). Our results suggest that C1/C2 heterozygosity, may favor the development of donor alloreactivity and thereby increase the risk of GVHD. Conversely, C1/C1 and C2/C2 homozygosity seems to reduce the risk of GVHD but may increase the incidence of graft rejection. These data may be helpful in tailoring the intensity of GVHD prophylaxis and conditioning regimens in thalassemia patients receiving HSCT from an HLA-identical volunteer donor.