Central Administration of Hydrogen Sulfide Donor NaHS Reduces Iba1-Positive Cells in the PVN and Attenuates Rodent Angiotensin II Hypertension

Hydrogen sulfide (H2S) is a gaseous signaling molecule with neuromodulatory, anti-inflammatory, and anti-hypertensive effects. Here, we investigate whether chronic intracerebroventricular (ICV) infusion of sodium hydrosulfide (NaHS), an H2S donor, can alleviate angiotensin II (Ang II)–induced hypertension (HTN), improve autonomic function, and impact microglia in the paraventricular nucleus (PVN) of the hypothalamus, a brain region associated with autonomic control of blood pressure (BP) and neuroinflammation in HTN. Chronic delivery of Ang II (200 ng/kg/min, subcutaneous) for 4 weeks produced a typical increase in BP and sympathetic drive and elevated the number of ionized calcium binding adaptor molecule 1–positive (Iba1+) cells in the PVN of male, Sprague–Dawley rats. ICV co-infusion of NaHS (at 30 and/or 60 nmol/h) significantly attenuated these effects of Ang II. Ang II also increased the abundance of cecal Deltaproteobacteria and Desulfovibrionales, among others, which was prevented by ICV NaHS co-infusion at 30 and 60 nmol/h. We observed no differences in circulating H2S between the groups. Our results suggest that central H2S may alleviate rodent HTN independently from circulating H2S via effects on autonomic nervous system and PVN microglia.

Innate and adaptive immunity to transfused allogenic red blood cells in mice requires MyD88

Red blood cell (RBC) transfusion therapy is essential for the survival of patients with hematological disorders such as sickle cell anemia. A potentially fatal complication of transfusion is development of non-ABO alloantibodies to polymorphic RBC antigens, yet mechanisms of alloantibody formation remain unclear. Human and mouse RBCs acquire a “storage lesion” prior to transfusion, which in mice contributes to immunogenicity. We previously reported that mouse splenic dendritic cells (DCs) are required for RBC alloimmunization and are activated by sterile and leukoreduced mouse RBCs after storage. Yet how syngeneic RBCs activate innate immune pathways to induce DC activation is unknown. We now show that DC activation to transfused RBCs occurs regardless of alloantigen presence, suggesting that RBC damage induced during storage triggers innate immune receptors. We discovered an unexpected dependence of RBC alloimmunization on the Toll-like receptor (TLR) signaling adaptor molecule MyD88. TLRs are a class of pattern recognition receptors (PRRs) that regulate DC activation and signal through two adaptor molecules, MyD88 and TRIF. We show that the inflammatory cytokine response, DC activation, and the subsequent alloantibody response to transfused syngeneic RBCs require MyD88 but not TRIF, suggesting a restricted set of PRRs are responsible for sensing RBCs and triggering alloimmunization.

Evidence of Reelin Signaling in GBM and Its Derived Cancer Stem Cells

Glioblastoma (GBM) is the most aggressive and malignant form of primary brain cancer, characterized by an overall survival time ranging from 12 to 18 months. Despite the progress in the clinical treatment and the growing number of experimental data aimed at investigating the molecular bases of GBM development, the disease remains characterized by a poor prognosis. Recent studies have proposed the existence of a population of GBM cancer stem cells (CSCs) endowed with self-renewal capability and a high tumorigenic potential that are believed to be responsible for the resistance against common chemotherapy and radiotherapy treatments. Reelin is a large secreted extracellular matrix glycoprotein, which contributes to positioning, migration, and laminar organization of several central nervous system structures during brain development. Mutations of the reelin gene have been linked to disorganization of brain structures during development and behavioral anomalies. In this study, we explored the expression of reelin in GBM and its related peritumoral tissue and performed the same analysis in CSCs isolated from both GBM (GCSCs) and peritumoral tissue (PCSCs) of human patients. Our findings reveal (i) the higher expression of reelin in GBM compared to the peritumoral tissue by immunohistochemical analysis, (ii) the mRNA expression of both reelin and its adaptor molecule Dab1 in either CSC subtypes, although at a different extent; and (iii) the contribution of CSCs-derived reelin in the migration of human primary GBM cell line U87MG. Taken together, our data indicate that the expression of reelin in GBM may represent a potential contribution to the regulation of GBM cancer stem cells behavior, further stimulating the interest on the reelin pathway as a potential target for GBM treatment.

USP18 positively regulates innate antiviral immunity by promoting K63-linked polyubiquitination of MAVS

Activation of MAVS, an adaptor molecule in Rig-I-like receptor (RLR) signaling, is indispensable for antiviral immunity, yet the molecular mechanisms modulating MAVS activation are not completely understood. Ubiquitination has a central function in regulating the activity of MAVS. Here, we demonstrate that a mitochondria-localized deubiquitinase USP18 specifically interacts with MAVS, promotes K63-linked polyubiquitination and subsequent aggregation of MAVS. USP18 upregulates the expression and production of type I interferon following infection with Sendai virus (SeV) or Encephalomyocarditis virus (EMCV). Mice with a deficiency of USP18 are more susceptible to RNA virus infection. USP18 functions as a scaffold protein to facilitate the re-localization of TRIM31 and enhances the interaction between TRIM31 and MAVS in mitochondria. Our results indicate that USP18 functions as a post-translational modulator of MAVS-mediated antiviral signaling.

Suppressive Role of Bam32/DAPP1 in Chemokine-Induced Neutrophil Recruitment

Bam32 (B cell adaptor molecule of 32 kDa) functions in the immune responses of various leukocytes. However, the role of neutrophil Bam32 in inflammation is entirely unknown. Here, we determined the role of Bam32 in chemokine CXCL2-induced neutrophil chemotaxis in three mouse models of neutrophil recruitment. By using intravital microscopy in the mouse cremaster muscle, we found that transmigrated neutrophil number, neutrophil chemotaxis velocity, and total neutrophil chemotaxis distance were increased in Bam32−/− mice when compared with wild-type (WT) mice. In CXCL2-induced mouse peritonitis, the total emigrated neutrophils were increased in Bam32−/− mice at 2 but not 4 h. The CXCL2-induced chemotaxis distance and migration velocity of isolated Bam32−/− neutrophils in vitro were increased. We examined the activation of small GTPases Rac1, Rac2, and Rap1; the levels of phospho-Akt2 and total Akt2; and their crosstalk with Bam32 in neutrophils. The deficiency of Bam32 suppressed Rap1 activation without changing the activation of Rac1 and Rac2. The pharmacological inhibition of Rap1 by geranylgeranyltransferase I inhibitor (GGTI298) increased WT neutrophil chemotaxis. In addition, the deficiency of Bam32, as well as the inhibition of Rap1 activation, increased the levels of CXCL2-induced Akt1/2 phosphorylation at Thr308/309 in neutrophils. The inhibition of Akt by SH-5 attenuated CXCL2-induced adhesion and emigration in Bam32−/− mice. Together, our results reveal that Bam32 has a suppressive role in chemokine-induced neutrophil chemotaxis by regulating Rap1 activation and that this role of Bam32 in chemokine-induced neutrophil recruitment relies on the activation of PI3K effector Akt.

Protrudin functions from the endoplasmic reticulum to support axon regeneration in the adult CNS

Adult mammalian central nervous system axons have intrinsically poor regenerative capacity, so axonal injury has permanent consequences. One approach to enhancing regeneration is to increase the axonal supply of growth molecules and organelles. We achieved this by expressing the adaptor molecule Protrudin which is normally found at low levels in non-regenerative neurons. Elevated Protrudin expression enabled robust central nervous system regeneration both in vitro in primary cortical neurons and in vivo in the injured adult optic nerve. Protrudin overexpression facilitated the accumulation of endoplasmic reticulum, integrins and Rab11 endosomes in the distal axon, whilst removing Protrudin’s endoplasmic reticulum localization, kinesin-binding or phosphoinositide-binding properties abrogated the regenerative effects. These results demonstrate that Protrudin promotes regeneration by functioning as a scaffold to link axonal organelles, motors and membranes, establishing important roles for these cellular components in mediating regeneration in the adult central nervous system.

A comparative study of the phenotype with kainic acid-induced seizure in DBA/2 mice from three different sources

The kainic acid-induced seizure mouse model is widely used in epilepsy research. In this study, we applied kainic acid to the subcutaneous injections of three different sources of DBA/2 mice to compare and evaluate the seizure response. The three mouse sources consisted of DBA/2Kor1 (Korea FDA source), DBA/2A (USA source), and DBA/2 (Japan source), and were purchased from different vendors. To compare the responses of DBA/2 mice to kainic acid injections, we examined the survival rate, seizure phenotype scoring, and behavioral changes. We also evaluated brain lesions using histopathological analysis. Following the administration of kainic acid, almost half of the cohort survived, and the seizure phenotype displayed a moderate level of sensitivity (2 ~ 4 out of 6). In the histopathologic analysis, there was no change in morphological features, and levels of glial fibrillary acidic protein (GFAP) and ionized calcium binding adaptor molecule 1 (Iba-1) increased in the kainic acid-treated groups. However, there was no difference in the neuronal nuclei (NeuN) expression level. All the data showed that the responses in the kainic acid-treated group were similar across the three strains. In conclusion, our results suggest that the three sources of DBA/2 mice (DBA/2Kor1, DBA/2A, and DBA/2B) have similar pathological responses to kainic acid-induced seizures.

Distinct integrin activation pathways for effector and regulatory T cell trafficking and function

Integrin activation mediates lymphocyte trafficking and immune functions. Conventional T cell (Tconv cell) integrin activation requires Rap1-interacting adaptor molecule (RIAM). Here, we report that Apbb1ip−/− (RIAM-null) mice are protected from spontaneous colitis due to IL-10 deficiency, a model of inflammatory bowel disease (IBD). Protection is ascribable to reduced accumulation and homing of Tconv cells in gut-associated lymphoid tissue (GALT). Surprisingly, there are abundant RIAM-null regulatory T cells (T reg cells) in the GALT. RIAM-null T reg cells exhibit normal homing to GALT and lymph nodes due to preserved activation of integrins αLβ2, α4β1, and α4β7. Similar to Tconv cells, T reg cell integrin activation and immune function require Rap1; however, lamellipodin (Raph1), a RIAM paralogue, compensates for RIAM deficiency. Thus, in contrast to Tconv cells, RIAM is dispensable for T reg cell integrin activation and suppressive function. In consequence, inhibition of RIAM can inhibit spontaneous Tconv cell–mediated autoimmune colitis while preserving T reg cell trafficking and function.