Zinc Overload Induces Damage to H9c2 Cardiomyocyte Through Mitochondrial Dysfunction and ROS-Mediated Mitophagy.

Zinc homeostasis is essential for maintaining redox balance, cell proliferation, and apoptosis. However, excessive zinc exposure is toxic and leads to mitochondrial dysfunction. In this study, we established a zinc overload model by treating rat cardiomyocyte H9c2 cells with Zn2+ at different concentrations. Our results showed that zinc overload increased LDH and reactive oxygen species (ROS) levels, leading to cell death, mitochondrial membrane potential decrease and impaired mitochondrial function and dynamics. Furthermore, zinc overload activated the PINK1/Parkin signaling pathway and induced mitochondrial autophagy via ROS, while NAC inhibited mitophagy and weakened the activation of PINK1/Parkin pathway, thereby preserving mitochondrial biogenesis. In addition, our data also showed that Mfn2 deletion increased ROS production and exacerbated cytotoxicity induced by zinc overload. Our results therefore suggest that Zn2+-induced ROS generation causes mitochondrial autophagy and mitochondrial dysfunction, damaging H9c2 cardiomyocytes. Additionally, Mfn2 may play a key role in zinc ion-mediated endoplasmic reticulum and mitochondrial interactions. Our results provide a new perspective on zinc-induced toxicology.

Factors Associated with Follow-Up Adherence After Teleophthalmology for Diabetic Eye Screening Before and During the COVID-19 Pandemic.

Abstract Background: Follow-up adherence with in-person care is critical for achieving improved clinical outcomes in telemedicine screening programs. We sought to quantify the impact of the COVID-19 pandemic upon follow-up adherence and factors associated with follow-up adherence after teleophthalmology for diabetic eye screening. Methods: We retrospectively reviewed medical records of adults screened in a clinical teleophthalmology program at urban and rural primary care clinics between May 2015 and December 2020. We defined follow-up adherence as medical record documentation of an in-person eye exam within 1 year among patients referred for further care. Regression models were used to identify factors associated with follow-up adherence. Results: Among 948 patients, 925 (97.6%) had health insurance and 170 (17.9%) were referred for follow-up. Follow-up adherence declined from 62.7% (n = 52) prepandemic to 46.0% (n = 40) during the pandemic (p = 0.04). There was a significant decline in follow-up adherence among patients from rural (p < 0.001), but not urban (p = 0.72) primary care clinics. Higher median household income (odds ratio [OR] 1.68, 95% confidence interval [CI]: 1.19-2.36) and obtaining care from an urban clinic (OR 5.29, 95% CI: 2.09-13.43) were associated with greater likelihood of follow-up during the pandemic. Discussion: Follow-up adherence remains limited after teleophthalmology screening even in a highly insured patient population, with a further decline observed during the COVID-19 pandemic. Our results suggest that rural patients and those with lower socioeconomic status experienced greater barriers to follow-up eye care during the COVID-19 pandemic. Conclusions: Addressing barriers to in-person follow-up care is needed to effectively improve clinical outcomes after teleophthalmology screening.

Extramacular Drusen and Progression of Age-Related Macular Degeneration: Age Related Eye Disease Study 2 Report 30.

PURPOSE: To identify the prevalence of extramacular drusen and their role in the progression of age-related macular degeneration (AMD). DESIGN: Retrospective analysis of a prospective cohort study. PARTICIPANTS: The study was conducted in 4168 eyes (2998 participants) with intermediate AMD in one or both eyes enrolled in the Age-Related Eye Disease Study 2 (AREDS2), a 5-year multicenter study of nutritional supplements. METHODS: Baseline 3-field 30-degree color photographs were evaluated for drusen characteristics outside the macular grid, including size, area, and location. The characteristics of extramacular drusen were compared with those of drusen within the macula. MAIN OUTCOME MEASURES: Progression rates to late AMD. RESULTS: Although extramacular drusen were observed in 3624 (86.9%) eyes, they represented a small area (< 0.5 mm2) in 50.3% of eyes, with only 17.5% exhibiting an area of > 1 disc area. Eyes with extramacular drusen exhibited larger macular drusen size and area than eyes without extramacular drusen (P < 0.001). Extramacular drusen were not associated with progression to late AMD. The hazard ratio adjusted for baseline age, sex, smoking, AMD severity level, and reticular pseudodrusen for 4043 eyes at risk of developing late AMD over 5 years was 1.17 (95% confidence interval [CI], 0.88-1.54; P = 0.27) for geographic atrophy and 0.96 (95% CI, 0.76-1.2; P = 0.7) for neovascular AMD. CONCLUSIONS: Extramacular drusen are commonly observed in eyes with AMD and are more frequent with an increasing drusen burden within the macula. In eyes with intermediate AMD, extramacular drusen do not confer additional risk to previously identified risk factors in progression to late AMD.

Hippocampal epigenetic and insulin-like growth factor alterations in noninvasive versus invasive mechanical ventilation in preterm lambs.

BACKGROUND: The brain of chronically ventilated preterm human infants is vulnerable to collateral damage during invasive mechanical ventilation (IMV). Damage is manifest, in part, by learning and memory impairments, which are hippocampal functions. A molecular regulator of hippocampal development is insulin-like growth factor 1 (IGF1). A gentler ventilation strategy is noninvasive respiratory support (NRS). We tested the hypotheses that NRS leads to greater levels of IGF1 messenger RNA (mRNA) variants and distinct epigenetic profile along the IGF1 gene locus in the hippocampus compared to IMV. METHODS: Preterm lambs were managed by NRS or IMV for 3 or 21 days. Isolated hippocampi were analyzed for IGF1 mRNA levels and splice variants for promoter 1 (P1), P2, and IGF1A and 1B, DNA methylation in P1 region, and histone covalent modifications along the gene locus. RESULTS: NRS had significantly greater levels of IGF1 P1 (predominant transcript), and 1A and 1B mRNA variants compared to IMV at 3 or 21 days. NRS also led to more DNA methylation and greater occupancy of activating mark H3K4 trimethylation (H3K4me3), repressive mark H3K27me3, and elongation mark H3K36me3 compared to IMV. CONCLUSIONS: NRS leads to distinct IGF1 mRNA variant levels and epigenetic profile in the hippocampus compared to IMV. IMPACT: Our study shows that 3 or 21 days of NRS of preterm lambs leads to distinct IGF1 mRNA variant levels and epigenetic profile in the hippocampus compared to IMV. Preterm infant studies suggest that NRS leads to better neurodevelopmental outcomes later in life versus IMV. Also, duration of IMV is directly related to hippocampal damage; however, molecular players remain unknown. NRS, as a gentler mode of respiratory management of preterm neonates, may reduce damage to the immature hippocampus through an epigenetic mechanism.

The Effect of Preoperative Opioid Dosage on Postoperative Outcomes in Patients Undergoing Knee Surgery.

BACKGROUND: Opioid prescription before knee replacement surgery is associated with longer hospital stays, more postsurgical pain, and a higher rate of complications. Despite the growing evidence against opioids, they remain popular preoperative pain management prescriptions. OBJECTIVES: The purpose of this study was to examine the effects of dosage of preoperative opioid use on orthopedic knee surgery pain control and postoperative outcomes and complications. STUDY DESIGN: Observational, retrospective evaluation. SETTING: University of Wisconsin Madison hospitals. METHODS: The patients underwent orthopedic knee surgery between May 1, 2014 and April 30, 2015. We randomly selected 197 patients and divided them into 2 groups that had preoperative opioid dosages of either low dose <= 120 mEq morphine (MME) or high dose >120 MME. Of 197 patients, 100 were in the low dose morphine group, whereas 97 were high dose. The cutoff at 120 MME was calculated to be the median dosage across all patients. The primary outcomes were compared, differences in postoperative pain control, and range of motion (ROM). Secondary outcomes included anesthetic complications, length of hospital stay, postoperative opioid dose, and postoperative complications. RESULTS: There were no statistically significant differences between the groups with regard to postoperative pain control, ROM, and immediate postoperative complications. Both groups showed similar length of hospitalization (2.199 to 2.304 days; P = 0.374), rate of postoperative infection, and joint intervention. The high dose group was more likely to have postoperative hemarthrosis and emergency department (ED) visits. However, the low dose group was more likely to have hypertension concurrently. LIMITATIONS: Because the study length was restricted to one year, the lack of data on longer term prognosis may limit extrapolation of data. Subjectivity of pain is difficult to measure and compare objectively. This study was not randomized prospectively, which may bias certain results due to unobserved differences. CONCLUSIONS: Preoperative opioid dose did not affect postoperative pain control or ROM in patients who received knee surgeries. Higher preoperative opioid doses were associated with more hemarthrosis and ED visits. Further exploration into quality of life indices and surgical complications such as need for revision may be a fruitful avenue. KEY WORDS: Opioids, analgesic, knee pain, total knee replacement, knee surgery, pre-operative opioids, knee outcomes.

Distributed representations of temporal stimulus associations across regular-firing and fast-spiking neurons in rat medial prefrontal cortex.

The prefrontal cortex has been implicated in various cognitive processes, including working memory, executive control, decision making, and relational learning. One core computational requirement underlying all these processes is the integration of information across time. When rodents and rabbits associate two temporally discontiguous stimuli, some neurons in the medial prefrontal cortex (mPFC) change firing rates in response to the preceding stimulus and sustain the firing rate during the subsequent temporal interval. These firing patterns are thought to serve as a mechanism to buffer the previously presented stimuli and signal the upcoming stimuli; however, how these critical properties are distributed across different neuron types remains unknown. We investigated the firing selectivity of regular-firing, burst-firing, and fast-spiking neurons in the prelimbic region of the mPFC while rats associated two neutral conditioned stimuli (CS) with one aversive stimulus (US). Analyses of firing patterns of individual neurons and neuron ensembles revealed that regular-firing neurons maintained rich information about CS identity and CS-US contingency during intervals separating the CS and US. Moreover, they further strengthened the latter selectivity with repeated conditioning sessions over a month. The selectivity of burst-firing neurons for both stimulus features was weaker than that of regular-firing neurons, indicating the difference in task engagement between two subpopulations of putative excitatory neurons. In contrast, putative inhibitory, fast-spiking neurons showed a stronger selectivity for CS identity than for CS-US contingency, suggesting their potential role in sensory discrimination. These results reveal a fine-scaled functional organization in the prefrontal network supporting the formation of temporal stimulus associations.NEW & NOTEWORTHY To associate stimuli that occurred separately in time, the brain needs to bridge the temporal gap by maintaining what was presented and predicting what would follow. We show that in rat medial prefrontal cortex, the former function is associated with a subpopulation of putative inhibitory neurons, whereas the latter is supported by a subpopulation of putative excitatory neurons. Our results reveal a distinct contribution of these microcircuit components to neural representations of temporal stimulus associations.

IUGR disrupts the PPARγ-Setd8-H4K20me(1) and Wnt signaling pathways in the juvenile rat hippocampus.

Intrauterine growth restriction (IUGR) programs neurodevelopmental impairment and long-term neurological morbidities. Neurological morbidities in IUGR infants are correlated with changes hippocampal volume. We previously demonstrated that IUGR alters hippocampal cellular composition in both neonatal and juvenile rat pups in association with altered hippocampal gene expression and epigenetic determinants. PPARγ signaling is important for neurodevelopment as well as epigenetic integrity in the brain via the PPARγ-Setd8-H4K20me(1) axis and Wnt signaling. We hypothesized that IUGR would decrease expression of PPARγ, Setd8, and H4K20me(1) in juvenile rat hippocampus. We further hypothesized that reduced PPARγ-Setd8-H4K20me(1) would be associated with reduced Wnt signaling genes Wnt3a and ß-catenin, and wnt target gene Axin2. To test our hypothesis we used a rat model of uteroplacental insufficiency-induced IUGR. We demonstrated that PPARγ localizes to oligodendrocytes, neurons and astrocytes within the juvenile rat hippocampus. We also demonstrated that IUGR reduces levels of PPARγ, Setd8 and H4K20me(1) in male and female juvenile rat hippocampus in conjunction with reduced Wnt signaling components in only male rats. We speculate that reduced PPARγ and Wnt signaling may contribute to altered hippocampal cellular composition which, in turn, may contribute to impaired neurodevelopment and subsequent neurocognitive impairment in IUGR offspring.

Traumatic brain injury increased IGF-1B mRNA and altered IGF-1 exon 5 and promoter region epigenetic characteristics in the rat pup hippocampus.

Traumatic brain injury (TBI) is a major cause of acquired cognitive disability in childhood. Such disability may be blunted by enhancing the brain's endogenous neuroprotective response. An important endogenous neuroprotective response is the insulin-like growth factor-1 (IGF-1) mRNA variant, IGF-1B. IGF-1B mRNA, characterized by exon 5 inclusion, encodes the IGF-1 and Eb peptides. IGF-1A mRNA excludes exon 5 and encodes the IGF-1 and Ea peptides. A region in the human IGF-1B homologue acts as an exon-splicing enhancer (ESE) to increase IGF-1B mRNA. It is not known if TBI is associated with increased brain IGF-1B mRNA. Epigenetic modifications may underlie altered gene expression in the brain after TBI. We hypothesized that TBI would increase hippocampal IGF-1B mRNA in 17-day-old rats, associated with DNA methylation and/or histone modifications at the promoter site 1 (P1) or exon 5/ESE region. Hippocampi from rat pups after controlled cortical impact (CCI) were used to measure IGF-1B mRNA, DNA methylation, and histone modifications at the P1, P2, and exon5/ESE regions. In CCI hippocampi, IGF-1B mRNA peaked at post-injury day (PID) 2 (1700±320% sham), but normalized by PID 14. IGF-1A peaked at PID 3 (280±52% sham), and remained elevated at PID 14. Increased IGF-1B mRNA was associated with increased methylation at P1, and increased histone modifications associated with gene activation at P2 and exon5/ESE, together with differential methylation in the exon 5/ESE regions. We report for the first time that hippocampal IGF-1B mRNA increased after developmental TBI. We speculate that epigenetic modifications at the P2 and exon 5/ESE regions are important in the regulation of IGF-1B mRNA expression. The exon 5/ESE region may present a means for future therapies to target IGF-1B transcription after TBI.