Preliminary Study on Clinical Characteristics and Pathogenesis of IQSEC2 Mutations Patients.

Background: The IQ motif and Sec7 domain ArfGEF 2 (IQSEC2), an X-linked gene that encodes the BRAG1 protein, is a guanine nucleotide exchange factor for the ADP ribosylation factor (ARF) protein family in the small guanosine triphosphate (GTP) binding protein. Mutations in this gene result in disorders such as intellectual disability (ID) and epilepsy. In this study, we analyze the clinical features of two patients with IQSEC2-mutation-related disease and discuss their possible pathogenesis. Methods: The two patients were diagnosed with ID and epilepsy. Genetic testing was performed using whole-exome sequencing, and the three-dimensional protein structure was analyzed. UCSC Genome Browser was used to analyze the conservation of IQSEC2 in different species. We compared IQSEC2 expression in the proband families with that in a control group, as well as the expression of the postsynaptic identity protein 95 (PSD-95), synapse-associated protein 97 (SAP97), ADP ribosylation factor 6 (ARF-6), and insulin receptor substrate 53kDa (IRSP53) genes interacting with IQSEC2. Results: We identified two semi-zygote mutations located in conserved positions in different species: an unreported de novo mutation, C.3576C>A (p. Tyr1192*), and a known mutation, c.2983C>T (p. Arg995Trp). IQSEC2 mutations resulted in significant changes in the predicted three-dimensional protein structure, while its expression in the two probands was significantly lower than that in the age-matched control group, and IQSEC2 expression in proband 1 was lower than that in his family members. The expression levels of PSD-95, ARF-6, and SAP97, IRSP 53, which interact with IQSEC2, were also significantly different from those in the family members and age-matched healthy children. Conclusion: The clinical phenotype resulting from IQSEC2 mutations can be explained by the significant decrease in its expression, loss of function of the mutant protein, and change in the expression of related genes. Our results provide novel insights into the molecular phenotype conferred by the IQSEC2 variants.

Mechanism of uncoupling protein 2­mediated myocardial injury in hypothermic preserved rat hearts.

In the present study, the alterations in uncoupling protein 2 (UCP2) expression following hypothermic preservation in rat hearts were investigated. Isolated rat hearts were preserved in Celsior solution for 3­12 h followed by 60 min of reperfusion. The cardiac function was evaluated using the Langendorff perfusion system. UCP2 and silent mating type information regulation 2 homolog 1 (SIRT1) proteins were detected by western blot analysis. The ATP production and mitochondrial reactive oxygen species (ROS) levels were assessed. Subsequent to preservation in ice­cold Celsior solution for 3­12 h, the UCP2 protein expression in rat hearts was observed to increase in a time­dependent manner. The UCP2 inhibitor genipin inhibited the hypothermic preservation­induced cardiac dysfunction, prevented a decline in ATP production induced by 9 h of preservation, however had no effect on the hypothermic preservation­induced increase in mitochondrial ROS levels. Compared with the control group, the SIRT1 protein expression in rat hearts reduced following hypothermic preservation. Compared with the 9­h preservation group, Celsior solution supplemented with the SIRT1 activator resveratrol (20 or 40 µmol/l) inhibited UCP2 protein overexpression, prevented the decline in ATP production and resulted in an improvement cardiac function. The SIRT1 inhibitor EX­527 abolished the resveratrol­induced inhibition of UCP2 overexpression and cardiac protection in the hypothermic preserved rat heart. These observations suggest that downregulation of UCP2 expression in the hypothermic preserved rat heart in part initiated the protective mechanism via the SIRT1 pathway.

Mitochondrial aldehyde dehydrogenase activity protects against lipopolysaccharide­induced cardiac dysfunction in rats.

Myocardial dysfunction in sepsis is associated with an increased risk of mortality. The mitochondrial aldehyde dehydrogenase (ALDH2) enzyme is crucial for protecting the heart from ischemic injury. The aim of the present study was to determine the role of ALDH2 in cardiac dysfunction induced by lipopolysaccharide (LPS). Male rats were treated intraperitoneally with LPS, and their stroke volume and cardiac output were evaluated using an M­mode echocardiograph. The expression levels and activity of ALDH2, nitric oxide content and inducible nitric oxide synthase (iNOS) activity, and the opening of the mitochondrial permeability transition pore (MPTP) were also evaluated. Treatment with LPS (5, 10, or 20 mg/kg) resulted in a steady decrease in cardiac output and stroke volume. The ALDH2 activity was decreased in a dose­dependent manner; however, the ALDH2 protein expression levels remained unchanged. Alda­1, a specific activator of ALDH2, increased the activity of ALDH2 and lessened LPS­induced cardiac dysfunction. A marked opening of the MPTP was observed 12 h following treatment with LPS, which was prevented by Alda­1. The improvement in cardiac function in response to treatment with Alda­1, was partially prevented by treatment with the MPTP inhibitor atractyloside. LPS treatment induced an increase in iNOS activation and inhibition of ALDH2 activity. The iNOS selective inhibitor, aminoguanidine, partially reversed the LPS­induced ALDH2 activity decrease and MPTP opening. These results indicate that ALDH2 activity may have a role in protecting against LPS­induced cardiac dysfunction. The potential mechanism may involve inhibition of MPTP opening and iNOS expression.