Molecular analysis of the human cytoglobin mRNA isoforms.

Multiple functions have been proposed for the ubiquitously expressed vertebrate globin cytoglobin (Cygb), including nitric oxide (NO) metabolism, lipid peroxidation/signalling, superoxide dismutase activity, reactive oxygen/nitrogen species (RONS) scavenging, regulation of blood pressure, antifibrosis, and both tumour suppressor and oncogenic effects. Since alternative splicing can expand the biological roles of a gene, we investigated whether this mechanism contributes to the functional diversity of Cygb. By mining of cDNA data and molecular analysis, we identified five alternative mRNA isoforms for the human CYGB gene (V-1 to V-5). Comprehensive RNA-seq analyses of public datasets from human tissues and cells confirmed that the canonical CYGB V-1 isoform is the primary CYGB transcript in the majority of analysed datasets. Interestingly, we revealed that isoform V-3 represented the predominant CYGB variant in hepatoblastoma (HB) cell lines and in the majority of analysed normal and HB liver tissues. CYGB V-3 mRNA is transcribed from an alternate upstream promoter and hypothetically encodes a N-terminally truncated CYGB protein, which is not recognized by some antibodies used in published studies. Little to no transcriptional evidence was found for the other CYGB isoforms. Comparative transcriptomics and flow cytometry on CYGB+/+ and gene-edited CYGB-/- HepG2 HB cells did not unveil a knockout phenotype and, thus, a potential function for CYGB V-3. Our study reveals that the CYGB gene is transcriptionally more complex than previously described as it expresses alternative mRNA isoforms of unknown function. Additional experimental data are needed to clarify the biological meaning of those alternative CYGB transcripts.

Biallelic MAD2L1BP (p31comet) mutation is associated with mosaic aneuploidy and juvenile granulosa cell tumors.

MAD2L1BP-encoded p31comet mediates Trip13-dependent disassembly of Mad2- and Rev7-containing complexes and, through this antagonism, promotes timely spindle assembly checkpoint (SAC) silencing, faithful chromosome segregation, insulin signaling, and homology-directed repair (HDR) of DNA double-strand breaks. We identified a homozygous MAD2L1BP nonsense variant, R253*, in 2 siblings with microcephaly, epileptic encephalopathy, and juvenile granulosa cell tumors of ovary and testis. Patient-derived cells exhibited high-grade mosaic variegated aneuploidy, slowed-down proliferation, and instability of truncated p31comet mRNA and protein. Corresponding recombinant p31comet was defective in Trip13, Mad2, and Rev7 binding and unable to support SAC silencing or HDR. Furthermore, C-terminal truncation abrogated an identified interaction of p31comet with tp53. Another homozygous truncation, R227*, detected in an early-deceased patient with low-level aneuploidy, severe epileptic encephalopathy, and frequent blood glucose elevations, likely corresponds to complete loss of function, as in Mad2l1bp-/- mice. Thus, human mutations of p31comet are linked to aneuploidy and tumor predisposition.

NOXA Accentuates Apoptosis Induction by a Novel Histone Deacetylase Inhibitor.

Epigenetic modifiers of the histone deacetylase (HDAC) family are often dysregulated in cancer cells. Experiments with small molecule HDAC inhibitors (HDACi) have proven that HDACs are a vulnerability of transformed cells. We evaluated a novel hydroxamic acid-based HDACi (KH16; termed yanostat) in human pancreatic ductal adenocarcinoma (PDAC) cells, short- and long-term cultured colorectal cancer (CRC) cells, and retinal pigment epithelial cells. We show that KH16 induces cell cycle arrest and apoptosis, both time and dose dependently in PDAC and CRC cells. This is associated with altered expression of BCL2 family members controlling intrinsic apoptosis. Recent data illustrate that PDAC cells frequently have an altered expression of the pro-apoptotic BH3-only protein NOXA and that HDACi induce an accumulation of NOXA. Using PDAC cells with a deletion of NOXA by CRISPR-Cas9, we found that a lack of NOXA delayed apoptosis induction by KH16. These results suggest that KH16 is a new chemotype of hydroxamic acid HDACi with superior activity against solid tumor-derived cells. Thus, KH16 is a scaffold for future research on compounds with nanomolar activity against HDACs.

Rapid translocation of intracellular toll-like receptors depends on endosomal NADPH oxidase.

Endosomal toll-like receptors (TLRs) must be translocated from the endoplasmic reticulum (ER) to the endosome and proteolytically cleaved within the endosome before they can induce cellular signals. As ligands for these TLRs are also liberated from apoptotic or necrotic cells, this process is controlled by several mechanisms which shall ensure that there is no inadvertent activation. We have shown previously that antiphospholipid antibodies induce endosomal NADPH-oxidase (NOX) followed by the translocation of TLR7/8 to the endosome. We show now that endosomal NOX is required for the rapid translocation of TLR3, TLR7/8, and TLR9. Deficiency of gp91phox, the catalytic subunit of NOX2, or inhibition of endosomal NOX by the chloride channel blocker niflumic acid both prevent immediate (i.e., within 30 min) translocation of these TLRs as shown by confocal laser scanning microscopy. Under these conditions, the induction of mRNA synthesis for TNF-α and secretion of TNF-α is delayed by approx. 6-9 h. However, maximal expression of TNF-α mRNA or secretion of TNF-α is not significantly reduced. In conclusion, these data add NOX2 as another component involved in the orchestration of cellular responses to ligands of endosomal TLRs.

Rural Differences in Victim Vulnerability and Revictimization of Intimate Partner Violence.

The aim of this study was to examine the association between the victim vulnerability factors included in the intimate partner violence (IPV) risk assessment tool used by the Swedish police (Brief Spousal Assault Form for the Evaluation of Risk [B-SAFER]) and rates of IPV revictimization among female victims living in rural towns, countryside, or remote areas. This study also aimed to examine the interaction between rurality and IPV revictimization in relation to victim vulnerability. The sample consisted of 695 cases of male-to-female perpetrated IPV, which had been reported to the Swedish police and subjected to a B-SAFER assessment. Rates of revictimization were examined in police registers. The results demonstrated that several vulnerability factors could discriminate between IPV revictimization across rurality. There was also an interaction effect between rurality and IPV revictimization in relation to the number of victim vulnerability factors present, where revictimization was more common for victims with many vulnerability factors living in more sparsely populated areas.

A complex epigenome-splicing crosstalk governs epithelial-to-mesenchymal transition in metastasis and brain development.

Epithelial-to-mesenchymal transition (EMT) renders epithelial cells migratory properties. While epigenetic and splicing changes have been implicated in EMT, the mechanisms governing their crosstalk remain poorly understood. Here we discovered that a C2H2 zinc finger protein, ZNF827, is strongly induced during various contexts of EMT, including in brain development and breast cancer metastasis, and is required for the molecular and phenotypic changes underlying EMT in these processes. Mechanistically, ZNF827 mediated these responses by orchestrating a large-scale remodelling of the splicing landscape by recruiting HDAC1 for epigenetic modulation of distinct genomic loci, thereby slowing RNA polymerase II progression and altering the splicing of genes encoding key EMT regulators in cis. Our findings reveal an unprecedented complexity of crosstalk between epigenetic landscape and splicing programme in governing EMT and identify ZNF827 as a master regulator coupling these processes during EMT in brain development and breast cancer metastasis.

Increased stability of the TM helix oligomer abrogates the apoptotic activity of the human Fas receptor.

Human death receptors control apoptotic events during cell differentiation, cell homeostasis and the elimination of damaged or infected cells. Receptor activation involves ligand-induced structural reorganizations of preformed receptor trimers. Here we show that the death receptor transmembrane domains only have a weak intrinsic tendency to homo-oligomerize within a membrane, and thus these domains potentially do not significantly contribute to receptor trimerization. However, mutation of Pro183 in the human CD95/Fas receptor transmembrane helix results in a dramatically increased interaction propensity, as shown by genetic assays. The increased interaction of the transmembrane domain is coupled with a decreased ligand-sensitivity of cells expressing the Fas receptor, and thus in a decreased number of apoptotic events. Mutation of Pro183 likely results in a substantial rearrangement of the self-associated Fas receptor transmembrane trimer, which likely abolishes further signaling of the apoptotic signal but may activate other signaling pathways. Our study shows that formation of a stable Fas receptor transmembrane helix oligomer does not per se result in receptor activation.

Neuroscience20 (BRAIN20, SPINE20, and MENTAL20) Health Initiative: A Global Consortium Addressing the Human and Economic Burden of Brain, Spine, and Mental Disorders Through Neurotech Innovations and Policies.

Neurological disorders significantly impact the world's economy due to their often chronic and life-threatening nature afflicting individuals which, in turn, creates a global disease burden. The Group of Twenty (G20) member nations, which represent the largest economies globally, should come together to formulate a plan on how to overcome this burden. The Neuroscience-20 (N20) initiative of the Society for Brain Mapping and Therapeutics (SBMT) is at the vanguard of this global collaboration to comprehensively raise awareness about brain, spine, and mental disorders worldwide. This paper aims to provide a comprehensive review of the various brain initiatives worldwide and highlight the need for cooperation and recommend ways to bring down costs associated with the discovery and treatment of neurological disorders. Our systematic search revealed that the cost of neurological and psychiatric disorders to the world economy by 2030 is roughly $16T. The cost to the economy of the United States is $1.5T annually and growing given the impact of COVID-19. We also discovered there is a shortfall of effective collaboration between nations and a lack of resources in developing countries. Current statistical analyses on the cost of neurological disorders to the world economy strongly suggest that there is a great need for investment in neurotechnology and innovation or fast-tracking therapeutics and diagnostics to curb these costs. During the current COVID-19 pandemic, SBMT, through this paper, intends to showcase the importance of worldwide collaborations to reduce the population's economic and health burden, specifically regarding neurological/brain, spine, and mental disorders.

mTOR Driven Gene Transcription Is Required for Cholesterol Production in Neurons of the Developing Cerebral Cortex.

Dysregulated mammalian target of rapamycin (mTOR) activity is associated with various neurodevelopmental disorders ranging from idiopathic autism spectrum disorders (ASD) to syndromes caused by single gene defects. This suggests that maintaining mTOR activity levels in a physiological range is essential for brain development and functioning. Upon activation, mTOR regulates a variety of cellular processes such as cell growth, autophagy, and metabolism. On a molecular level, however, the consequences of mTOR activation in the brain are not well understood. Low levels of cholesterol are associated with a wide variety of neurodevelopmental disorders. We here describe numerous genes of the sterol/cholesterol biosynthesis pathway to be transcriptionally regulated by mTOR complex 1 (mTORC1) signaling in vitro in primary neurons and in vivo in the developing cerebral cortex of the mouse. We find that these genes are shared targets of the transcription factors SREBP, SP1, and NF-Y. Prenatal as well as postnatal mTORC1 inhibition downregulated expression of these genes which directly translated into reduced cholesterol levels, pointing towards a substantial metabolic function of the mTORC1 signaling cascade. Altogether, our results indicate that mTORC1 is an essential transcriptional regulator of the expression of sterol/cholesterol biosynthesis genes in the developing brain. Altered expression of these genes may be an important factor contributing to the pathogenesis of neurodevelopmental disorders associated with dysregulated mTOR signaling.

Lipid presentation by the protein C receptor links coagulation with autoimmunity.

Antiphospholipid antibodies (aPLs) cause severe autoimmune disease characterized by vascular pathologies and pregnancy complications. Here, we identify endosomal lysobisphosphatidic acid (LBPA) presented by the CD1d-like endothelial protein C receptor (EPCR) as a pathogenic cell surface antigen recognized by aPLs for induction of thrombosis and endosomal inflammatory signaling. The engagement of aPLs with EPCR-LBPA expressed on innate immune cells sustains interferon- and toll-like receptor 7-dependent B1a cell expansion and autoantibody production. Specific pharmacological interruption of EPCR-LBPA signaling attenuates major aPL-elicited pathologies and the development of autoimmunity in a mouse model of systemic lupus erythematosus. Thus, aPLs recognize a single cell surface lipid-protein receptor complex to perpetuate a self-amplifying autoimmune signaling loop dependent on the cooperation with the innate immune complement and coagulation pathways.

COVID-19: Review of a 21st Century Pandemic from Etiology to Neuro-psychiatric Implications.

COVID-19 is a severe infectious disease that has claimed >150,000 lives and infected millions in the United States thus far, especially the elderly population. Emerging evidence has shown the virus to cause hemorrhagic and immunologic responses, which impact all organs, including lungs, kidneys, and the brain, as well as extremities. SARS-CoV-2 also affects patients', families', and society's mental health at large. There is growing evidence of re-infection in some patients. The goal of this paper is to provide a comprehensive review of SARS-CoV-2-induced disease, its mechanism of infection, diagnostics, therapeutics, and treatment strategies, while also focusing on less attended aspects by previous studies, including nutritional support, psychological, and rehabilitation of the pandemic and its management. We performed a systematic review of >1,000 articles and included 425 references from online databases, including, PubMed, Google Scholar, and California Baptist University's library. COVID-19 patients go through acute respiratory distress syndrome, cytokine storm, acute hypercoagulable state, and autonomic dysfunction, which must be managed by a multidisciplinary team including nursing, nutrition, and rehabilitation. The elderly population and those who are suffering from Alzheimer's disease and dementia related illnesses seem to be at the higher risk. There are 28 vaccines under development, and new treatment strategies/protocols are being investigated. The future management for COVID-19 should include B-cell and T-cell immunotherapy in combination with emerging prophylaxis. The mental health and illness aspect of COVID-19 are among the most important side effects of this pandemic which requires a national plan for prevention, diagnosis and treatment.

Rbfox1 Is Expressed in the Mouse Brain in the Form of Multiple Transcript Variants and Contains Functional E Boxes in Its Alternative Promoters.

The RNA-binding protein RBFOX1 is an important regulator of neuron development and neuronal excitability. Rbfox1 is a dosage-sensitive gene and in both mice and humans, decreased expression of Rbfox1 has been linked to neurodevelopmental disorders. Alternative promoters drive expression of Rbfox1 transcript isoforms that encode an identical protein. The tissue- and developmental stage-specific expression of these isoforms, as well as the underlying regulatory mechanisms, are, however, unclear. Here, we set out to capture all of the Rbfox1 transcript isoforms and identify transcriptional mechanisms that regulate brain-specific Rbfox1 expression. Isoform sequencing identified multiple alternative Rbfox1 transcript variants in the mouse cerebral cortex, including transcripts with novel first exons, alternatively spliced exons and 3'-truncations. Quantitative RT-PCR determined the expression of the alternative first exons in the developing cerebral cortex and different subregions of the juvenile brain. Alternative first exons were found to be highly stage- and subregion specific in their expression patterns suggesting that they fulfill specific functions during cortex development and in different brain regions. Using reporter assays we found that the promoter regions of the two first exons E1B and E1C/E1C.1 contain several functional E-boxes. Together, we provide an extensive picture of Rbfox1 isoform expression. We further identified important regulatory mechanisms that drive neuron-specific Rbfox1 expression. Thus, our study forms the basis for further research into the mechanisms that ensure physiological Rbfox1 expression in the brain. It also helps to understand why, in patients with neurodevelopmental disorders deletion of individual RBFOX1 transcript isoforms could affect brain function.

Inhibition of histone deacetylation rescues phenotype in a mouse model of Birk-Barel intellectual disability syndrome.

Mutations in the actively expressed, maternal allele of the imprinted KCNK9 gene cause Birk-Barel intellectual disability syndrome (BBIDS). Using a BBIDS mouse model, we identify here a partial rescue of the BBIDS-like behavioral and neuronal phenotypes mediated via residual expression from the paternal Kcnk9 (Kcnk9pat) allele. We further demonstrate that the second-generation HDAC inhibitor CI-994 induces enhanced expression from the paternally silenced Kcnk9 allele and leads to a full rescue of the behavioral phenotype suggesting CI-994 as a promising molecule for BBIDS therapy. Thus, these findings suggest a potential approach to improve cognitive dysfunction in a mouse model of an imprinting disorder.

Family-Only Perpetrators of Intimate Partner Violence: A Systematic Review.

This article presents the first systematic review of family-only intimate partner violence (IPV) perpetrators (as originally proposed by Holtzworth-Munroe & Stuart). The aims of the present review were to summarize and describe the prevalence of the family-only perpetrator subtype, as well as to investigate what characteristics were associated with perpetrators within this subtype. Electronic literature searches in several databases (e.g., PsychINFO, Web of Science, and PubMed) were carried out. Of the 3,434 studies identified, 30 studies met the inclusion criteria as well as the methodological quality criteria. Thematic analyses were conducted, where several themes and subthemes were identified. The proportion of family-only perpetrators, averaged across sample types, was 47.5%. Drawing on the thematic analyses of the reviewed studies, family-only perpetrators presented as a less violent subtype, displaying several pro-social personality traits, as well as a lower degree of psychopathology. The findings were in line with Holtzworth-Munroe and Stuart's predictions. The findings also demonstrated the utility of a 2-fold typology, consisting of a family-only and a generally violent (GV) subtype, as well as the need to reconsider the one-size-fits-all approach to IPV treatment. We also included a discussion of the terminology of the subtypes and propose an adoption of the terms "partner only violent" and "generally violent" subtypes.

Dietary wheat amylase trypsin inhibitors promote features of murine non-alcoholic fatty liver disease.

We previously demonstrated that a common dietary protein component, wheat amylase trypsin inhibitors (ATI), stimulate intestinal macrophages and dendritic cells via toll like receptor 4. Activation of these intestinal myeloid cells elicits an inflammatory signal that is propagated to mesenteric lymph nodes, and that can facilitate extraintestinal inflammation. Mice were fed a well-defined high fat diet, with (HFD/ATI) or without (HFD) nutritionally irrelevant amounts of ATI. Mice on HFD/ATI developed only mild signs of intestinal inflammation and myeloid cell activation but displayed significantly higher serum triglycerides and transaminases compared to mice on HFD alone. Moreover, they showed increased visceral and liver fat, and a higher insulin resistance. ATI feeding promoted liver and adipose tissue inflammation, with M1-type macrophage polarization and infiltration, and enhanced liver fibrogenesis. Gluten, the major protein component of wheat, did not induce these pathologies. Therefore, wheat ATI ingestion in minute quantities comparable to human daily wheat consumption exacerbated features of the metabolic syndrome and non-alcoholic steatohepatitis, despite its irrelevant caloric value.

Tissue factor pathway inhibitor primes monocytes for antiphospholipid antibody-induced thrombosis.

Antiphospholipid antibodies (aPLs) with complex lipid and/or protein reactivities cause complement-dependent thrombosis and pregnancy complications. Although cross-reactivities with coagulation regulatory proteins contribute to the risk for developing thrombosis in patients with antiphospholipid syndrome, the majority of pathogenic aPLs retain reactivity with membrane lipid components and rapidly induce reactive oxygen species-dependent proinflammatory signaling and tissue factor (TF) procoagulant activation. Here, we show that lipid-reactive aPLs activate a common species-conserved TF signaling pathway. aPLs dissociate an inhibited TF coagulation initiation complex on the cell surface of monocytes, thereby liberating factor Xa for thrombin generation and protease activated receptor 1/2 heterodimer signaling. In addition to proteolytic signaling, aPLs promote complement- and protein disulfide isomerase-dependent TF-integrin ß1 trafficking that translocates aPLs and NADPH oxidase to the endosome. Cell surface TF pathway inhibitor (TFPI) synthesized by monocytes is required for TF inhibition, and disabling TFPI prevents aPL signaling, indicating a paradoxical prothrombotic role for TFPI. Myeloid cell-specific TFPI inactivation has no effect on models of arterial or venous thrombus development, but remarkably prevents experimental aPL-induced thrombosis in mice. Thus, the physiological control of TF primes monocytes for rapid aPL pathogenic signaling and thrombosis amplification in an unexpected crosstalk between complement activation and coagulation signaling.

Assessing risk for inpatient physical violence in a female forensic psychiatric sample - comparing HCR-20v2 with the female additional manual to the HCR-20v2.

Purpose and aim: Out from the sparse literature on risk assessment for violence committed by women the Female Additional Manual (FAM) was developed to be a complement to the HCR-20v2. The aim of this study was to investigate and compare the psychometrics of the HCR-20v2 with and without the FAM on risk for inpatient physical violence for female forensic psychiatric patients. Methods: The participants were 100 female patients admitted to forensic psychiatric care in a high-security clinic, assessed by clinicians with the HCR-20v2 during their admission. Researchers performed the FAM, both retrospectively and prospectively. The follow-up period was 12 months before being discharged. Results: Four main results were found; first, many risk factors were present although the summary risk ratings were mainly low to moderate. Secondly, the reliability was in general good, where the HCR-20v2 mainly showed higher reliability without than with the FAM, indicating that FAM risk factors did equal or did not contribute to a higher reliability. Third, the internal validity was higher for the HCR-20v2 than for the FAM. Risk factors correlated stronger with the summary risk ratings for the HCR-20v2 than for the FAM. Fourth, the validity for inpatient physical violence was high for the total score of both the HCR-20v2 and the FAM, but contradictory to previous finding the validity for summary risk ratings was not significant. Conclusions: The results support the use of HCR-20v2 when assessing risk for inpatient violence for female forensic psychiatric patients, but with only some support for adding or changing risk factors according to the FAM.

Intimate Partner Violence in Urban, Rural, and Remote Areas: An Investigation of Offense Severity and Risk Factors.

This study compared the severity of intimate partner violence (IPV) and the relationship between risk factors for IPV and overall risk judgments of future IPV in urban, rural, and remote areas. IPV risk assessments conducted by the Swedish police between 2010 and 2014 in urban ( n = 564), rural ( n = 456), and remote ( n = 196) areas were examined. Rurality was associated with the severity of IPV reported, as well as the presence of risk factors and their relationship to overall risk judgments. Cases in remote areas included more severe IPV as well as more risk factors.

Application of patient-derived liver cancer cells for phenotypic characterization and therapeutic target identification.

Primary liver cancer (PLC) ranks among the most lethal solid cancers worldwide due to lack of effective biomarkers for early detection and limited treatment options in advanced stages. Development of primary culture models that closely recapitulate phenotypic and molecular diversities of PLC is urgently needed to improve the patient outcome. Long-term cultures of 7 primary liver cancer cell lines of hepatocellular and cholangiocellular origin were established using defined culture conditions. Morphological and histological characteristics of obtained cell lines and xenograft tumors were analyzed and compared to original tumors. Time course analyses of transcriptomic and genomic changes were performed using next-generation sequencing (NGS). Key oncogenic alterations were identified by targeted NGS and cell lines carrying potentially actionable mutations were treated with corresponding specific inhibitors. PDCL fully resembled morphological features of the primary cancers in vitro and in vivo over extended period in culture. Genomic alterations as well as transcriptome profiles showed high similarity with primary tumors and remained stable during long-term culturing. Targeted-NGS confirmed that key oncogenic mutations such as TP53, KRAS, CTNNB1 as well as actionable mutations (e.g. MET, cKIT, KDR) were highly conserved in PDCL and amenable for individualized therapeutic approaches. Integrative genomic and transcriptomic approaches further demonstrated that PDCL more closely resemble molecular and prognostic features of PLC than established cell lines and are valuable tool for direct target evaluation. Our integrative analysis demonstrates that PDCL represents refined model for discovery of relevant molecular subgroups and exploration of precision medicine approaches for the treatment of this deadly disease.