Association between COVID-19 and sensorineural hearing loss: Evidence from a Mendelian randomization study in European and East Asian population.

BACKGROUND: Long coronavirus disease (COVID), characterized by persistent and sometimes debilitating symptoms following a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, has garnered increasing attention as a potential public health crisis. Emerging evidence indicates a higher incidence of hearing loss in individuals who have had COVID 2019 (COVID-19) compared to the general population. However, the conclusions were inconsistent, and the causal relationship between COVID-19 and sensorineural hearing loss remains unknown. METHODS: To addresses this outstanding issue, we performed Mendelian randomization analysis to detect the causal association between COVID-19 and hearing loss using the largest genome-wide association study data to date in the European population and confirmed the results in the East Asian population. Comprehensively sensitive analyses were followed, including Cochran's Q test, Mendelian randomization (MR)-Egger intercept test, MR-pleiotropy residual sum and outlier, and leave-one-out analysis, to validate the robustness of our results. RESULTS: Our results suggested that there is no causal association between COVID-19 and the risk of hearing loss in the European population. Neither the susceptibility, hospitalization, and severity of COVID-19 on hearing loss (inverse variance weighted method: odds ratio (OR) = 1.046, 95% confidence interval (CI) = 0.907-1.205, p = .537; OR = 0.995, 95% CI = 0.956-1.036, p = .823; OR = 0.995, 95% CI = 0.967-1.025, p = .76). Replicated analyses in the East Asian population yielded consistent results. No pleiotropy and heterogeneity were found in our results. CONCLUSION: In conclusion, our MR results do not support a genetically predicted causal relationship between COVID-19 and sensorineural hearing loss. Thus, the associations observed in prior observational studies may have been influenced by confounding factors rather than a direct cause-and-effect relationship. More clinical and mechanism research are needed to further understand this association in the future.

Association between gut microbiota and sensorineural hearing loss: a Mendelian randomization study.

Background: Several recent studies speculated that the gut microbiota is associated with sensorineural hearing loss (SNHL) and proposed the concept of the gut-inner ear axis. However, the causal effect of gut microbiota on SNHL is still unknown. In this study, we performed a two-sample Mendelian randomization (MR) analysis to estimate the causal effect of gut microbiota on SNHL. Methods: Gut microbiota data were obtained from the largest available genome-wide association study (n = 18,340) conducted by the MiBioGen consortium. The summary statistics of SNHL were obtained from the FinnGen consortium R8 release data (28,310 cases and 302,750 controls). The causal effects were estimated with inverse-variance weighted, MR-Egger, and weighted median. Reverse Mendelian randomization analysis was performed on the bacteria that were found to be associated with SNHL in forward Mendelian randomization analysis. We then performed sensitivity analyses, including Cochran's Q-test, MR-Egger intercept test, MR-PRESSO, cML-MA-BIC, and leave-one-out analysis, to detect heterogeneity and pleiotropy. Results: The inverse-variance weighted results suggested that Lachnospiraceae (UCG001) had a significant protective effect against SNHL (odds ratio = 0.85, 95% confidence interval: 0.78-0.93, P = 6.99 × 10-4). In addition, Intestinimonas (odds ratio = 0.89, 95% confidence interval: 0.82-0.97, P = 8.53 × 10-3) presented a suggestively protective effect on SNHL. Rikenellaceae (RC9gutgroup) (odds ratio = 1.08, 95% confidence interval: 1.02-1.15, P = 0.01) and Eubacterium (hallii group) (odds ratio = 1.12, 95% confidence interval: 1.00-1.24, P = 0.048) suggestively increase the risk of SNHL. The results of the reverse MR analysis showed that there is no significant causal effect of SNHL on the gut microbiota. No significant heterogeneity of instrumental variables or pleiotropy was detected. Conclusion: The evidence that the four genera mentioned above are associated with SNHL supports the hypothesis of a gut-inner ear axis. Our study provides microbial markers for the prevention and treatment of SNHL, and further studies are needed to explore the mechanisms of the gut microbiome-inner ear axis in health and diseases.

Progress on lysosomal PPT1-mediated regulation of cellular homeostasis and pathogenesis.

Palmitoyl protein thioesterase 1(PPT1) is a lysosomal enzyme that catalyzes the protein depalmitoylation. It is considered to play a crucial role in regulating lysosomes, mitochondria and lipid metabolism. PPT1 has been reported to play an important role in the occurrence and progression of diseases, such as neurological diseases and cancers. However, the regulatory mechanisms remain unknown. In this review, we summarize the progress of PPT1 function and mechanisms in neurological disorders and cancers, which will provide as reference and guidance for exploring the regulatory mechanisms of PPT1 and developing new drugs for treating related diseases in the future.

A feedback loop engaging propionate catabolism intermediates controls mitochondrial morphology.

D-2-Hydroxyglutarate (D-2HG) is an α-ketoglutarate-derived mitochondrial metabolite that causes D-2-hydroxyglutaric aciduria, a devastating developmental disorder. How D-2HG adversely affects mitochondria is largely unknown. Here, we report that in Caenorhabditis elegans, loss of the D-2HG dehydrogenase DHGD-1 causes D-2HG accumulation and mitochondrial damage. The excess D-2HG leads to a build-up of 3-hydroxypropionate (3-HP), a toxic metabolite in mitochondrial propionate oxidation, by inhibiting the 3-HP dehydrogenase HPHD-1. We demonstrate that 3-HP binds the MICOS subunit MIC60 (encoded by immt-1) and inhibits its membrane-binding and membrane-shaping activities. We further reveal that dietary and gut bacteria affect mitochondrial health by modulating the host production of 3-HP. These findings identify a feedback loop that links the toxic effects of D-2HG and 3-HP on mitochondria, thus providing important mechanistic insights into human diseases related to D-2HG and 3-HP.

A pair of transporters controls mitochondrial Zn2+ levels to maintain mitochondrial homeostasis.

Zn2+ is required for the activity of many mitochondrial proteins, which regulate mitochondrial dynamics, apoptosis and mitophagy. However, it is not understood how the proper mitochondrial Zn2+ level is achieved to maintain mitochondrial homeostasis. Using Caenorhabditis elegans, we reveal here that a pair of mitochondrion-localized transporters controls the mitochondrial level of Zn2+. We demonstrate that SLC-30A9/ZnT9 is a mitochondrial Zn2+ exporter. Loss of SLC-30A9 leads to mitochondrial Zn2+ accumulation, which damages mitochondria, impairs animal development and shortens the life span. We further identify SLC-25A25/SCaMC-2 as an important regulator of mitochondrial Zn2+ import. Loss of SLC-25A25 suppresses the abnormal mitochondrial Zn2+ accumulation and defective mitochondrial structure and functions caused by loss of SLC-30A9. Moreover, we reveal that the endoplasmic reticulum contains the Zn2+ pool from which mitochondrial Zn2+ is imported. These findings establish the molecular basis for controlling the correct mitochondrial Zn2+ levels for normal mitochondrial structure and functions.

The Rab7 effector WDR91 promotes autophagy-lysosome degradation in neurons by regulating lysosome fusion.

The effectors of the Rab7 small GTPase play multiple roles in Rab7-dependent endosome-lysosome and autophagy-lysosome pathways. However, it is largely unknown how distinct Rab7 effectors coordinate to maintain the homeostasis of late endosomes and lysosomes to ensure appropriate endolysosomal and autolysosomal degradation. Here we report that WDR91, a Rab7 effector required for early-to-late endosome conversion, is essential for lysosome function and homeostasis. Mice lacking Wdr91 specifically in the central nervous system exhibited behavioral defects and marked neuronal loss in the cerebral and cerebellar cortices. At the cellular level, WDR91 deficiency causes PtdIns3P-independent enlargement and dysfunction of lysosomes, leading to accumulation of autophagic cargoes in mouse neurons. WDR91 competes with the VPS41 subunit of the HOPS complex, another Rab7 effector, for binding to Rab7, thereby facilitating Rab7-dependent lysosome fusion in a controlled manner. WDR91 thus maintains an appropriate level of lysosome fusion to guard the normal function and survival of neurons.

CDK4/6 regulate lysosome biogenesis through TFEB/TFE3.

Lysosomes are degradation and signaling organelles that adapt their biogenesis to meet many different cellular demands; however, it is unknown how lysosomes change their numbers for cell division. Here, we report that the cyclin-dependent kinases CDK4/6 regulate lysosome biogenesis during the cell cycle. Chemical or genetic inactivation of CDK4/6 increases lysosomal numbers by activating the lysosome and autophagy transcription factors TFEB and TFE3. CDK4/6 interact with and phosphorylate TFEB/TFE3 in the nucleus, thereby inactivating them by promoting their shuttling to the cytoplasm. During the cell cycle, lysosome numbers increase in S and G2/M phases when cyclin D turnover diminishes CDK4/6 activity. These findings not only uncover the molecular events that direct the nuclear export of TFEB/TFE3, but also suggest a mechanism that controls lysosome biogenesis in the cell cycle. CDK4/6 inhibitors promote autophagy and lysosome-dependent degradation, which has important implications for the therapy of cancer and lysosome-related disorders.

The amino acid transporter SLC-36.1 cooperates with PtdIns3P 5-kinase to control phagocytic lysosome reformation.

Phagocytic removal of apoptotic cells involves formation, maturation, and digestion of cell corpse-containing phagosomes. The retrieval of lysosomal components following phagolysosomal digestion of cell corpses remains poorly understood. Here we reveal that the amino acid transporter SLC-36.1 is essential for lysosome reformation during cell corpse clearance in Caenorhabditis elegans embryos. Loss of slc-36.1 leads to formation of phagolysosomal vacuoles arising from cell corpse-containing phagosomes. In the absence of slc-36.1, phagosome maturation is not affected, but the retrieval of lysosomal components is inhibited. Moreover, loss of PPK-3, the C. elegans homologue of the PtdIns3P 5-kinase PIKfyve, similarly causes accumulation of phagolysosomal vacuoles that are defective in phagocytic lysosome reformation. SLC-36.1 and PPK-3 function in the same genetic pathway, and they directly interact with one another. In addition, loss of slc-36.1 and ppk-3 causes strong defects in autophagic lysosome reformation in adult animals. Our findings thus suggest that the PPK-3-SLC-36.1 axis plays a central role in both phagocytic and autophagic lysosome formation.

Maintenance of neural activities in torpid Rhinolophus ferrumequinum bats revealed by 2D gel-based proteome analysis.

Bats are the only mammals capable of self-powered flying. Many bat species hibernate in winter. A reversible control of cerebral activities is critical for bats to accommodate a repeated torpor-arousal cycle during hibernation. Little is known about the molecular mechanisms that regulate neuronal activities in torpid bats. In this study, Rhinolophus ferrumequinum bat brain proteins were fractionated, and their abundance in active and torpid states was compared. Results of 2D gel-based proteomics showed that 38% of identified proteins with a significant change in abundance are involved in synaptic vesicle recycling and cytoskeletal integrity. Changes in the abundance of proteins related to RNA splicing, proteostasis, redox homeostasis, mitochondrial function, and energy metabolism were also detected. In addition, the levels of GNAO1 (guanine nucleotide-binding protein Gαo subunit), an important modulator of neuronal transmembrane signaling, were significantly increased in the insoluble protein fraction of torpid bats; this may be due to GNAO1 palmitoylation making it insoluble. Our data provide molecular evidence for the maintenance of neuronal activities in torpid bats and suggest that a reversible palmitoylation of the G protein plays a role in the regulation of neuronal activities during bat hibernation.

Antioxidant Defenses in the Brains of Bats during Hibernation.

Hibernation is a strategy used by some mammals to survive a cold winter. Small hibernating mammals, such as squirrels and hamsters, use species- and tissue-specific antioxidant defenses to cope with oxidative insults during hibernation. Little is known about antioxidant responses and their regulatory mechanisms in hibernating bats. We found that the total level of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the brain of each of the two distantly related hibernating bats M. ricketti and R. ferrumequinum at arousal was lower than that at torpid or active state. We also found that the levels of malondialdehyde (product of lipid peroxidation) of the two hibernating species of bats were significantly lower than those of non-hibernating bats R. leschenaultia and C. sphinx. This observation suggests that bats maintain a basal level of ROS/RNS that does no harm to the brain during hibernation. Results of Western blotting showed that hibernating bats expressed higher amounts of antioxidant proteins than non-hibernating bats and that M. ricketti bats upregulated the expression of some enzymes to overcome oxidative stresses, such as superoxide dismutase, glutathione reductase, and catalase. In contrast, R. ferrumequinum bats maintained a relatively high level of superoxide dismutase 2, glutathione reductase, and thioredoxin-2 throughout the three different states of hibernation cycles. The levels of glutathione (GSH) were higher in M. ricketti bats than in R. ferrumequinum bats and were significantly elevated in R. ferrumequinum bats after torpor. These data suggest that M. ricketti bats use mainly antioxidant enzymes and R. ferrumequinum bats rely on both enzymes and low molecular weight antioxidants (e.g., glutathione) to avoid oxidative stresses during arousal. Furthermore, Nrf2 and FOXOs play major roles in the regulation of antioxidant defenses in the brains of bats during hibernation. Our study revealed strategies used by bats against oxidative insults during hibernation.

Molecular Evolution of the Nuclear Factor (Erythroid-Derived 2)-Like 2 Gene Nrf2 in Old World Fruit Bats (Chiroptera: Pteropodidae).

Mammals developed antioxidant systems to defend against oxidative damage in their daily life. Enzymatic antioxidants and low molecular weight antioxidants (LMWAs) constitute major parts of the antioxidant systems. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2, encoded by the Nrf2 gene) is a central transcriptional regulator, regulating transcription, of many antioxidant enzymes. Frugivorous bats eat large amounts of fruits that contain high levels of LMWAs such as vitamin C, thus, a reliance on LMWAs might greatly reduce the need for antioxidant enzymes in comparison to insectivorous bats. Therefore, it is possible that frugivorous bats have a reduced need for Nrf2 function due to their substantial intake of diet-antioxidants. To test whether the Nrf2 gene has undergone relaxed evolution in fruit-eating bats, we obtained Nrf2 sequences from 16 species of bats, including four Old World fruit bats (Pteropodidae) and one New World fruit bat (Phyllostomidae). Our molecular evolutionary analyses revealed changes in the selection pressure acting on Nrf2 gene and identified seven specific amino acid substitutions that occurred on the ancestral lineage leading to Old World fruit bats. Biochemical experiments were conducted to examine Nrf2 in Old World fruit bats and showed that the amount of catalase, which is regulated by Nrf2, was significantly lower in the brain, heart and liver of Old World fruit bats despite higher levels of Nrf2 protein in Old World fruit bats. Computational predictions suggest that three of these seven amino acid replacements might be deleterious to Nrf2 function. Therefore, the results suggest that Nrf2 gene might have experienced relaxed constraint in Old World fruit bats, however, we cannot rule out the possibility of positive selection. Our study provides the first data on the molecular adaptation of Nrf2 gene in frugivorous bats in compensation to the increased levels of LWMAs from their fruit-diet.

Phosphoenolpyruvate carboxykinase 1 gene (Pck1) displays parallel evolution between Old World and New World fruit bats.

Bats are an ideal mammalian group for exploring adaptations to fasting due to their large variety of diets and because fasting is a regular part of their life cycle. Mammals fed on a carbohydrate-rich diet experience a rapid decrease in blood glucose levels during a fast, thus, the development of mechanisms to resist the consequences of regular fasts, experienced on a daily basis, must have been crucial in the evolution of frugivorous bats. Phosphoenolpyruvate carboxykinase 1 (PEPCK1, encoded by the Pck1 gene) is the rate-limiting enzyme in gluconeogenesis and is largely responsible for the maintenance of glucose homeostasis during fasting in fruit-eating bats. To test whether Pck1 has experienced adaptive evolution in frugivorous bats, we obtained Pck1 coding sequence from 20 species of bats, including five Old World fruit bats (OWFBs) (Pteropodidae) and two New World fruit bats (NWFBs) (Phyllostomidae). Our molecular evolutionary analyses of these sequences revealed that Pck1 was under purifying selection in both Old World and New World fruit bats with no evidence of positive selection detected in either ancestral branch leading to fruit bats. Interestingly, however, six specific amino acid substitutions were detected on the ancestral lineage of OWFBs. In addition, we found considerable evidence for parallel evolution, at the amino acid level, between the PEPCK1 sequences of Old World fruit bats and New World fruit bats. Test for parallel evolution showed that four parallel substitutions (Q276R, R503H, I558V and Q593R) were driven by natural selection. Our study provides evidence that Pck1 underwent parallel evolution between Old World and New World fruit bats, two lineages of mammals that feed on a carbohydrate-rich diet and experience regular periods of fasting as part of their life cycle.

Critical roles of mitochondria in brain activities of torpid Myotis ricketti bats revealed by a proteomic approach.

Bats are the only mammals that fly and hibernate. Little is known about their overall metabolism in the brain during hibernation. In this study, brain proteins of torpid and active Myotis ricketti bats were fractionated and compared using a proteomic approach. Results showed that 21% (23 proteins) of identified proteins with significant expression changes were associated with amino acid metabolism and proteostasis. The expression levels of proteins involved in energy metabolism (15 proteins), cytoskeletal structure (18 proteins), and stress response (13 proteins) were also significantly altered in torpid bats. Over 30% (34 proteins) of differentially expressed proteins were associated with mitochondrial functions. Various post-translational modifications (PTMs) on PDHB, DLD, and ARG1 were detected, suggesting that bats use PTMs to regulate protein functions during torpor. Antioxidation and stress responses in torpid bats were similar to those of hibernated squirrels, suggesting a common strategy adopted by small hibernators against brain dysfunction. Since many amino acids that metabolize in mitochondria modulate neuronal transmissions, results of this study reveal pivotal roles of mitochondria in neural communication, metabolic regulation, and brain cell survival during bat hibernation. This article is part of a Special Issue entitled: Proteomics of non-model organisms. BIOLOGICAL SIGNIFICANCE: This study reveals the mechanisms used by bats to regulate brain activities during torpor. These mechanisms include post-translational modifications and differential expression of proteins involved in mitochondrial electron transport, anaerobic glycolysis, TCA cycle efflux, cytoskeletal plasticity, amino acid metabolism, vesicle structure, antioxidation defense, stress response, and proteostasis. Our study provides insights in metabolic regulation of flying mammals during torpor and common strategies used by small hibernators in response to hibernation. This article is part of a Special Issue entitled: Proteomics of non-model organisms.

Ancient invasion of an extinct gammaretrovirus in cetaceans.

Endogenous gammaretroviruses (EGVs) have been widely studied in terrestrial mammals but seldom so in marine species. A genomic mining of the bottlenose dolphin (Tursiops truncatus) genome revealed a new EGV, termed Tursiops truncatus endogenous retrovirus (TTEV), which is divergent from extant mammalian EGVs. Molecular clock dating estimated the invasion time of TTEV into the host genome to be approximately 10-19 million years ago (MYA), while a previously identified killer whale endogenous gammaretrovirus (KWERV) was estimated to have invaded the host genome approximately 3-5 MYA. Using a PCR-based technique, we then verified that similar endogenous viruses exist in nine cetacean genomes. Phylogenetic analysis revealed that these cetacean EGVs are highly divergent from their counterparts in other mammals, including KWERV from the killer whale. In sum, we conclude that there have been at least two invasion episodes of EGVs into cetaceans during their evolutionary history.