ABSTRACT
Oxygen minimum zones (OMZs) are expanding due to increased sea surface temperatures, subsequent increased oxygen demand through respiration, reduced oxygen solubility, and thermal stratification driven in part by anthropogenic climate change. Devil's Hole, Bermuda is a model ecosystem to study OMZ microbial biogeochemistry because the formation and subsequent overturn of the suboxic zone occur annually. During thermally driven stratification, suboxic conditions develop, with organic matter and nutrients accumulating at depth. In this study, the bioavailability of the accumulated dissolved organic carbon (DOC) and the microbial community response to reoxygenation of suboxic waters was assessed using a simulated overturn experiment. The surface inoculated prokaryotic community responded to the deep (formerly suboxic) 0.2 µm filtrate with cell densities increasing 2.5-fold over 6 days while removing 5 µmol L-1 of DOC. After 12 days, the surface community began to shift, and DOC quality became less diagenetically altered along with an increase in SAR202, a Chloroflexi that can degrade recalcitrant dissolved organic matter (DOM). Labile DOC production after 12 days coincided with an increase of Nitrosopumilales, a chemoautotrophic ammonia oxidizing archaea (AOA) that converts ammonia to nitrite based on the ammonia monooxygenase (amoA) gene copy number and nutrient data. In comparison, the inoculation of the deep anaerobic prokaryotic community into surface 0.2 µm filtrate demonstrated a die-off of 25.5% of the initial inoculum community followed by a 1.5-fold increase in cell densities over 6 days. Within 2 days, the prokaryotic community shifted from a Chlorobiales dominated assemblage to a surface-like heterotrophic community devoid of Chlorobiales. The DOM quality changed to less diagenetically altered material and coincided with an increase in the ribulose-1,5-bisphosphate carboxylase/oxygenase form I (cbbL) gene number followed by an influx of labile DOM. Upon reoxygenation, the deep DOM that accumulated under suboxic conditions is bioavailable to surface prokaryotes that utilize the accumulated DOC initially before switching to a community that can both produce labile DOM via chemoautotrophy and degrade the more recalcitrant DOM.
ABSTRACT
Aniridia is a rare congenital syndrome that is associated with reduced visual acuity and progressive loss of vision. Aniridia patients may also develop systemic health issues associated with defects in the pancreas, digestive, and central nervous systems. The spectrum of symptoms associated with aniridia is due to haploinsufficiency of the paired box 6 gene (PAX6) and its role in the development and maintenance of the affected tissues. Here, we isolated pancreatic islets from mice heterozygous for Pax6 to test whether a Pax6-specific miRNA suppression (target protector) strategy can restore PAX6 protein levels. We show that miR-7 and miR-375 target specific sites within the Pax6 3' UTR in a mouse pancreatic ß-insulinoma cell line. Tough decoys (Tuds) against miR-7 and miR-375 increase expression of a mouse Pax6 3' UTR luciferase reporter and increase PAX6 protein levels in these cells. Finally, we demonstrate that the shielding of the miR-7 binding site with a target protector restores PAX6 protein levels in the Pax6 heterozygous islets. The data presented here represent a proof of concept for RNA-based therapy for the progressive defects associated with aniridia and suggest the target protector approach may be a useful therapeutic strategy for other haploinsufficiency diseases.
ABSTRACT
Microcirculatory damage is a common complication for those with vascular risk factors, such as diabetes. To resolve vascular insults, the brain's immune cells (microglia) must rapidly envelop the site of injury. Currently, it is unknown whether Type 1 diabetes, a condition associated with chronic immune system dysfunction, alters microglial responses to damage and what mechanisms are responsible. Using in vivo two-photon microscopy in adult male mice, we show that microglial envelopment of laser-induced cerebral microbleeds is diminished in a hyperglycemic mouse model of Type 1 diabetes, which could not be fully rescued with chronic insulin treatment. Microglia were important for vessel repair because reduced microglial accumulation in diabetic mice or near-complete depletion in healthy controls was associated with greater secondary leakage of the damaged vessel. Broadly suppressing inflammation with dexamethasone in diabetic mice but not healthy controls, significantly enhanced microglial responses to microbleeds and attenuated secondary vessel leakage. These enhancements were associated with changes in IFN-γ signaling because dexamethasone suppressed abnormally high levels of IFN-γ protein levels in brain and blood serum of diabetic mice. Further, blocking IFN-γ in diabetic mice with neutralizing antibodies restored normal microglial chemotaxic responses and purinoceptor P2ry12 gene expression, as well as mitigated secondary leakage. These results suggest that abnormal IFN-γ signaling disrupts microglial function in the diabetic brain, and that immunotherapies targeting IFN-γ can stimulate microglial repair of damaged vessels.SIGNIFICANCE STATEMENT Although Type 1 diabetes is an established risk factor for vascular complications, such as microbleeds, and is known to hinder wound healing in the body, no study has examined how diabetes impacts the brain's innate immune reparative response (involving cells called microglia) to vascular injury. Here we show that microglial responses to brain microbleeds were diminished in diabetic animals, which also exacerbated secondary leakage from damaged vessels. These impairments were related to abnormally high levels of the proinflammatory cytokine IFN-γ because reducing IFN-γ with immunosuppressant drugs or blocking antibodies helped restore normal microglial responses and repair of damaged vessels. These data highlight the use of IFN-γ modulating therapeutics to enhance vascular repair in at-risk populations.
