One-Year Outcomes of Postintensive Care Syndrome in Critically Ill Coronavirus Disease 2019 Patients: A Single Institutional Study.

IMPORTANCE: Postintensive care syndrome has a strong impact on coronavirus disease 2019 survivors. OBJECTIVES: Assess the 1-year prevalence of postintensive care syndrome after coronavirus disease 2019. DESIGN SETTING AND PARTICIPANTS: This was a single-center prospective cohort using questionnaires and telephone calls from 4 months to 1 year after ICU discharge. Patients who were treated for coronavirus disease 2019-related acute respiratory distress between March 19, 2020, and April 30, 2020, participated. MAIN OUTCOMES AND MEASURES: Postintensive care syndrome was evaluated according to physical, mental, and cognitive domains. We surveyed the 8-item standardized Short Form questionnaire for assessing physical postintensive care syndrome; the Impact of Event Scale-Revised and the Hospital Anxiety and Depression Scale for assessing mental postintensive care syndrome; and Short-Memory Questionnaire for assessing cognitive postintensive care syndrome. The primary outcome was postintensive care syndrome occurrence of any domain at 1 year. Furthermore, the co-occurrence of the three postintensive care syndrome domains was assessed. RESULTS: Eighteen patients consented to the study and completed the survey. The median age was 57.5 years, and 78% of the patients were male. Median Acute Physiology and Chronic Health Evaluation-II score was 18. During ICU stay, 78% received invasive mechanical ventilation, and 83% received systemic steroid administration. Early mobilization was implemented in 61%. Delirium occurred in 44%. The median days of ICU and hospital stay were 6 and 23.5, respectively. Overall postintensive care syndrome occurrence was 67%. Physical, mental, and cognitive postintensive care syndrome occurred in 56%, 50%, and 33% of patients, respectively. The co-occurrence of all three domains of postintensive care syndrome was 28%. Age and Acute Physiology and Chronic Health Evaluation-II scores were higher, and systemic steroids were more commonly used in the postintensive care syndrome groups compared with the nonpostintensive care syndrome groups. Chronic symptoms were more common in the postintensive care syndrome groups than the nonpostintensive care syndrome groups. CONCLUSIONS AND RELEVANCE: Patients who suffered critical illness from coronavirus disease 2019 had a high frequency of postintensive care syndrome after 1 year. Long-term follow-up and care should be continuously offered.

The histone H4 lysine 20 monomethyl mark, set by PR-Set7 and stabilized by L(3)mbt, is necessary for proper interphase chromatin organization.

Drosophila PR-Set7 or SET8 is a histone methyltransferase that specifically monomethylates histone H4 lysine 20 (H4K20). L(3)MBT has been identified as a reader of methylated H4K20. It contains several conserved domains including three MBT repeats binding mono- and dimethylated H4K20 peptides. We find that the depletion of PR-Set7 blocks de novo H4K20me1 resulting in the immediate activation of the DNA damage checkpoint, an increase in the size of interphase nuclei, and drastic reduction of cell viability. L(3)mbt on the other hand stabilizes the monomethyl mark, as L(3)mbt-depleted S2 cells show a reduction of more than 60% of bulk monomethylated H4K20 (H4K20me1) while viability is barely affected. Ploidy and basic chromatin structure show only small changes in PR-Set7-depleted cells, but higher order interphase chromatin organization is significantly affected presumably resulting in the activation of the DNA damage checkpoint. In the absence of any other known functions of PR-Set7, the setting of the de novo monomethyl mark appears essential for cell viability in the presence or absence of the DNA damage checkpoint, but once newly assembled chromatin is established the monomethyl mark, protected by L(3)mbt, is dispensable.

Functional characterization of the Drosophila Hmt4-20/Suv4-20 histone methyltransferase.

Di- and trimethylation of histone H4 lysine20 (H4K20) are thought to play an important role in controlling gene expression in vertebrates and in Drosophila. By inducing a null mutation in Drosophila Suv4-20, we show that it encodes the histone H4 lysine20 di- and trimethyltransferase. In Suv4-20 mutants, the H4K20 di- and trimethyl marks are strongly reduced or absent, and the monomethyl mark is significantly increased. We find that even with this biochemical function, Suv4-20 is not required for survival and does not control position-effect variegation (PEV).

Aberrant monomethylation of histone H4 lysine 20 activates the DNA damage checkpoint in Drosophila melanogaster.

PR-Set7 is a histone methyltransferase that specifically monomethylates histone H4 lysine 20 (K20) and is essential for cell proliferation. Our results show that in PR-Set7 mutants, the DNA damage checkpoint is activated. This phenotype is manifested by reduction in both the mitotic and the S phase indexes, a delay in the progression through early mitosis, and strong reduction of cyclin B. Furthermore, in a double mutant of PR-Set7 and mei-41 (the fly ATR orthologue), the abnormalities of mitotic progression and the cyclin B protein level were rescued. PR-Set7 also showed a defect in chromosome condensation that was enhanced in the double mutant. We therefore propose that monomethylated H4K20 is involved in the maintenance of proper higher order structure of DNA and is consequently essential for chromosome condensation.

Stress-induced transcription of the endoplasmic reticulum oxidoreductin gene ERO1 in the yeast Saccharomyces cerevisiae.

Gene transcription changes dramatically in response to various stresses. This event is an obligatory step for adaptation of cells to certain environments. Endoplasmic reticulum (ER) oxidoreductin encoded by the ERO1 gene of the yeast Saccharomyces cerevisiae is essential for the formation of protein disulfide bonds in the ER and for cell viability. We show that transcription of ERO1 is regulated by two transcriptional activators in response to different stresses. In the unfolded protein response induced by the reductant dithiothreitol, transcription factor Hac1 activates ERO1 transcription through a sequence that diverges from the consensus Hac1-binding sequence. Heat shock transcription factor Hsf1 activates ERO1 in response to heat, ethanol, and oxidative stresses. Using cells containing mutations in the Hac1- and Hsf1-binding sequences of the chromosomal ERO1 promoter, we demonstrate that Hac1-regulated transcription of ERO1 confers resistance to dithiothreitol. Although mutations in the Hsf1-binding sequences do not affect the sensitivity of cells to heat, ethanol, or oxidative stresses, both the Hac1- and Hsf1-regulated pathways are critical for normal growth under complex stress conditions.

Functional compatibility between isoform alpha and beta of type II DNA topoisomerase.

DNA topoisomerase II (topo II) plays a crucial role in controlling the conformation of both DNA and whole chromosomes. This activity is essential for several cellular events such as DNA replication, transcription, chromosome condensation and segregation. In mammals, two genes code for isoforms of topo II, termed alpha and beta. They are similar in primary structure and have almost identical catalytic properties in vitro. We transfected HeLa cells with small interfering RNAs (siRNAs) targeted against either topo IIalpha or IIbeta, and succeeded in knocking down the expression of the corresponding protein. Chromosomes were condensed and aligned at metaphase in topo IIalpha-knockdown cells. Although some lagging chromosomes were observed, they were still segregated at anaphase despite the absence of topo IIalpha. When both topo IIalpha and topo IIbeta were removed, the segregation of chromosomes was severely arrested, suggesting that topo IIbeta could partially substitute for topo IIalpha. Double-knockdown experiments also revealed that topo II was required for shortening of the chromosome axis.

Induction of apoptosis by depletion of DNA topoisomerase IIalpha in mammalian cells.

Inactivation of topoisomerase (topo) IIalpha arrests murine embryonic development. In topo IIalpha-depleted embryos, nuclei were partitioned to daughter cells without complete separation and formed an interconnecting droplet-like structure. The present study examined the fates of topo IIalpha-depleted cells with the droplet-like nuclear structure. When the embryos with abnormal nuclei were further incubated, apoptosis was induced along with the formation of fragmented and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling positive nuclei. ICRF-193 treatment of embryos activated caspases. Apoptosis induced by ICRF-193 was suppressed by z-VAD-fmk, a caspase inhibitor, and pifithrin-alpha, a p53 inhibitor. Moreover, when mitosis was blocked by nocodazole, ICRF-193-induced nuclear abnormalities and apoptosis were abolished. These data suggest that cycling through the M-phase is essential for ICRF-193-induced apoptosis. Nuclear abnormalities similar to those of topo IIalpha-depleted embryos were induced in HeLa cells in which topo IIalpha was knocked down by transfection with short interfering RNA (siRNA) against topo IIalpha, followed by induction of apoptosis. Our results suggest that topo IIalpha-depleted cells with the droplet-like nuclear structure induce apoptosis, which is dependent on caspase and p53 activity during the G1 phase in mammalian cells.

Epitope distribution of randomly established monoclonal antibodies against human type II DNA topoisomerases.

The epitopes of about 100 monoclonal antibodies against human type II DNA topoisomerase were mapped along the enzyme molecules. Although they were randomly and independently established, epitope sites were unevenly distributed the toward N-terminal or C-terminal region. We suggest that the central catalytic domain is hidden inside the molecule and inaccessible to the antigen recognition sites. Using antibodies, we demonstrate the distinct localization of isoforms of Topo II in cultured cells. Some particularly useful antibodies are listed.