CYP19A1 mediates severe SARS-CoV-2 disease outcome in males.

Male sex represents one of the major risk factors for severe COVID-19 outcome. However, underlying mechanisms that mediate sex-dependent disease outcome are as yet unknown. Here, we identify the CYP19A1 gene encoding for the testosterone-to-estradiol metabolizing enzyme CYP19A1 (also known as aromatase) as a host factor that contributes to worsened disease outcome in SARS-CoV-2-infected males. We analyzed exome sequencing data obtained from a human COVID-19 cohort (n = 2,866) using a machine-learning approach and identify a CYP19A1-activity-increasing mutation to be associated with the development of severe disease in men but not women. We further analyzed human autopsy-derived lungs (n = 86) and detect increased pulmonary CYP19A1 expression at the time point of death in men compared with women. In the golden hamster model, we show that SARS-CoV-2 infection causes increased CYP19A1 expression in the lung that is associated with dysregulated plasma sex hormone levels and reduced long-term pulmonary function in males but not females. Treatment of SARS-CoV-2-infected hamsters with a clinically approved CYP19A1 inhibitor (letrozole) improves impaired lung function and supports recovery of imbalanced sex hormones specifically in males. Our study identifies CYP19A1 as a contributor to sex-specific SARS-CoV-2 disease outcome in males. Furthermore, inhibition of CYP19A1 by the clinically approved drug letrozole may furnish a new therapeutic strategy for individualized patient management and treatment.

Region-Specific Effects of Fractionated Low-Dose Versus Single-Dose Radiation on Hippocampal Neurogenesis and Neuroinflammation.

BACKGROUND: Despite technical advances in hippocampus-sparing radiotherapy, radiation-induced injury to neural stem cell compartments may affect neurocognitive functions. In pre-clinical mouse models with fractionated low-dose radiation (FLDR) and single-dose radiation (SDR), the accurate response to radiation-induced injury was analyzed in different hippocampal subregions. METHODS: Adult and juvenile C57BL/6NCrl mice were exposed to FLDR (20 × 0.1 Gy, daily exposure from Monday to Friday for 4 weeks) or SDR (1 × 2 Gy). In addition, 72 h after the last exposure, neuroglia (astrocytes and microglia) and neuroprogenitor cells were characterized and quantified in the hippocampal cornu ammonis (CA) and dentate gyrus (DG) by immunofluorescence studies. RESULTS: After analyzing different hippocampal subregions, it was observed that radiation responses varied between non-neurogenic CA, with no detectable inflammatory alterations, and neurogenic DG, characterized by impaired neurogenesis and subsequent neuroinflammation. Age-dependent differences in radiosensitivity appeared to depend on the varying proliferative potential of neural stem cell niches. Using the same overall dose for FLDR and SDR (2 Gy), both the cumulative dose over time and also the single dose fraction have decisive impacts on hippocampal damage. CONCLUSION: Region-specific effects of radiation-induced hippocampal injury relies primarily on cell deaths of proliferating neuroprogenitors. Dose per fraction defines the extent of neuronal injury, and subsequently activated microglia and reactive astrocytes modulate dynamic processes of neuroinflammation. Thus, limiting both cumulative doses and dose fractions to hippocampal DG is an important issue of clinical radiotherapy to preserve neurocognitive functions.

Cultured Human Foreskin as a Model System for Evaluating Ionizing Radiation-Induced Skin Injury.

PURPOSE: Precise molecular and cellular mechanisms of radiation-induced dermatitis are incompletely understood. Histone variant H2A.J is associated with cellular senescence and modulates senescence-associated secretory phenotype (SASP) after DNA-damaging insults, such as ionizing radiation (IR). Using ex vivo irradiated cultured foreskin, H2A.J was analyzed as a biomarker of radiation-induced senescence, potentially initiating the inflammatory cascade of radiation-induced skin injury. METHODS: Human foreskin explants were collected from young donors, irradiated ex vivo with 10 Gy, and cultured in air-liquid interphase for up to 72 h. At different time-points after ex vivo IR exposure, the foreskin epidermis was analyzed for proliferation and senescence markers by immunofluorescence and immunohistochemical staining of sectioned tissue. Secretion of cytokines was measured in supernatants by ELISA. Using our mouse model with fractionated in vivo irradiation, H2A.J expression was analyzed in epidermal stem/progenitor cell populations localized in different regions of murine hair follicles (HF). RESULTS: Non-vascularized foreskin explants preserved their tissue homeostasis up to 72 h (even after IR exposure), but already non-irradiated foreskin epithelium expressed high levels of H2A.J in all epidermal layers and secreted high amounts of cytokines. Unexpectedly, no further increase in H2A.J expression and no obvious upregulation of cytokine secretion was observed in the foreskin epidermis after ex vivo IR. Undifferentiated keratinocytes in murine HF regions, by contrast, revealed low H2A.J expression in non-irradiated skin and significant radiation-induced H2A.J upregulations at different time-points after IR exposure. Based on its staining characteristics, we presume that H2A.J may have previously underestimated the importance of the epigenetic regulation of keratinocyte maturation. CONCLUSIONS: Cultured foreskin characterized by highly keratinized epithelium and specific immunological features is not an appropriate model for studying H2A.J-associated tissue reactions during radiation-induced dermatitis.

Histone Variant H2A.J Is Enriched in Luminal Epithelial Gland Cells.

H2A.J is a poorly studied mammalian-specific variant of histone H2A. We used immunohistochemistry to study its localization in various human and mouse tissues. H2A.J showed cell-type specific expression with a striking enrichment in luminal epithelial cells of multiple glands including those of breast, prostate, pancreas, thyroid, stomach, and salivary glands. H2A.J was also highly expressed in many carcinoma cell lines and in particular, those derived from luminal breast and prostate cancer. H2A.J thus appears to be a novel marker for luminal epithelial cancers. Knocking-out the H2AFJ gene in T47D luminal breast cancer cells reduced the expression of several estrogen-responsive genes which may explain its putative tumorigenic role in luminal-B breast cancer.

Fractionated Low-Dose Radiation Induces Long-Lasting Inflammatory Responses in the Hippocampal Stem Cell Niche.

PURPOSE: Despite major technical advances in hippocampus-sparing radiation therapy, radiation-induced injury to the neural stem cell compartment may affect neurocognitive functions. In the brain, glial cells modulate neuronal functions and are major mediators of neuroinflammation. In a preclinical mouse model with fractionated low-dose radiation (LDR), the complex response to radiation-induced injury was analyzed in the hippocampal stem cell compartment over a period of 6 months. METHODS AND MATERIALS: Adult and juvenile C57BL/6NCrl mice were exposed to low doses of ionizing radiation (IR; 20 fractions of 0.1 Gy, for up to 4 weeks) daily. At 72 hours and 1, 3, and 6 months after fractionated LDR, magnetic resonance imaging (9.4 T) was conducted to detect structural and functional abnormalities in the hippocampal region. Using immunofluorescence and histologic studies, neuroglia cells (astrocytes, microglia, oligodendrocytes) were quantified and neuroinflammatory responses were characterized in the hippocampal dentate gyrus. Using in vivo bromodeoxyuridine labeling, the cell fate of newly generated progenitor cells was tracked in the subgranular zone of the dentate gyrus during fractionated LDR. RESULTS: Low doses of IR induced long-lasting inflammatory responses with local increases of activated microglia and reactive astrocytes, which were most pronounced in the juvenile hippocampus within the first months after LDR. Glial activation with the consequent release of proinflammatory mediators increased local blood flow and vascular permeability in the hippocampal region. Cell fate mapping of progenitors located in the subgranular zone revealed a transient shift from neurogenesis to gliogenesis. CONCLUSIONS: Glial cell activation and transient neuroinflammation may reflect radiation-induced neuronal damage in the hippocampal stem cell niche. The increased proliferative capacity of the developing brain may explain the enhanced hippocampal radiosensitivity, with stronger inflammatory reactions in the juvenile hippocampus. Thus, limiting the radiation dose to the hippocampal region is an important issue of clinical radiation therapy at all ages to preserve neurocognitive functions.

Human skin aging is associated with increased expression of the histone variant H2A.J in the epidermis.

Cellular senescence is an irreversible growth arrest that occurs as a result of damaging stimuli, including DNA damage and/or telomere shortening. Here, we investigate histone variant H2A.J as a new biomarker to detect senescent cells during human skin aging. Skin biopsies from healthy volunteers of different ages (18-90 years) were analyzed for H2A.J expression and other parameters involved in triggering and/or maintaining cellular senescence. In the epidermis, the proportions of H2A.J-expressing keratinocytes increased from ≈20% in young to ≈60% in aged skin. Inverse correlations between Ki67- and H2A.J staining in germinative layers may reflect that H2A.J-expressing cells having lost their capacity to divide. As cellular senescence is triggered by DNA-damage signals, persistent 53BP1-foci, telomere lengths, and telomere-associated damage foci were analyzed in epidermal keratinocytes. Only slight age-related telomere attrition and few persistent nuclear 53BP1-foci, occasionally colocalizing with telomeres, suggest that unprotected telomeres are not a significant cause of senescence during skin aging. Quantification of integrin-α6+ basal cells suggests that the number and function of stem/progenitor cells decreased during aging and their altered proliferation capacities resulted in diminished tissue renewal with epidermal thinning. Collectively, our findings suggest that H2A.J is a sensitive marker of epidermal aging in human skin.

DNA damage accumulation during fractionated low-dose radiation compromises hippocampal neurogenesis.

BACKGROUND AND PURPOSE: High-precision radiotherapy is an effective treatment modality for tumors. Intensity-modulated radiotherapy techniques permit close shaping of high doses to tumors, however healthy organs outside the target volume are repeatedly exposed to low-dose radiation (LDR). The inherent vulnerability of hippocampal neurogenesis is likely the determining factor in radiation-induced neurocognitive dysfunctions. Using preclinical in-vivo models with daily LDR we attempted to precisely define the pathophysiology of radiation-induced neurotoxicity. MATERIAL AND METHODS: Genetically defined mouse strains with varying DNA repair capacities were exposed to fractionated LDR (5×/10×/15×/20×0.1 Gy) and dentate gyri from juvenile and adult mice were analyzed 72 h after last exposure and 1, 3, 6 months after 20 × 0.1 Gy. To examine the impact of LDR on neurogenesis, persistent DNA damage was assessed by quantifying 53BP1-foci within hippocampal neurons. Moreover, subpopulations of neuronal stem/progenitor cells were quantified and dendritic arborization of developing neurons were assessed. To unravel molecular mechanisms involved in radiation-induced neurotoxicity, hippocampi were analyzed using mass spectrometry-based proteomics and affected signaling networks were validated by immunoblotting. RESULTS: Radiation-induced DNA damage accumulation leads to progressive decline of hippocampal neurogenesis with decreased numbers of stem/progenitor cells and reduced complexities of dendritic architectures, clearly more pronounced in repair-deficient mice. Proteome analysis revealed substantial changes in neurotrophic signaling, with strong suppression directly after LDR and compensatory upregulation later on to promote functional recovery. CONCLUSION: Hippocampal neurogenesis is highly sensitive to repetitive LDR. Even low doses affect signaling networks within the neurogenic niche and interrupt the dynamic process of generation and maturation of neuronal stem/progenitor cells.