Search

Ferroptosis is a pervasive non-apoptotic form of cell death highly relevant in various degenerative diseases and malignancies. The hallmark of ferroptosis is uncontrolled and overwhelming peroxidation of polyunsaturated fatty acids contained in membrane phospholipids, which eventually leads to rupture of the plasma membrane. Ferroptosis is unique in that it is essentially a spontaneous, uncatalyzed chemical process based on perturbed iron and redox homeostasis contributing to the cell death process, but that it is nonetheless modulated by many metabolic nodes that impinge on the cells’ susceptibility to ferroptosis. Among the various nodes affecting ferroptosis sensitivity, several have emerged as promising candidates for pharmacological intervention, rendering ferroptosis-related proteins attractive targets for the treatment of numerous currently incurable diseases. Herein, the current members of a Germany-wide research consortium focusing on ferroptosis research, as well as key external experts in ferroptosis who have made seminal contributions to this rapidly growing and exciting field of research, have gathered to provide a comprehensive, state-of-the-art review on ferroptosis. Specific topics include: basic mechanisms, in vivo relevance, specialized methodologies, chemical and pharmacological tools, and the potential contribution of ferroptosis to disease etiopathology and progression. We hope that this article will not only provide established scientists and newcomers to the field with an overview of the multiple facets of ferroptosis, but also encourage additional efforts to characterize further molecular pathways modulating ferroptosis, with the ultimate goal to develop novel pharmacotherapies to tackle the various diseases associated with – or caused by – ferroptosis.

According to the ICH S3A Q&A, microsampling is applicable to pharmaceutical drugs and toxicological analysis. Few studies have reported the effect of microsampling on the toxicity of immunotoxicological drugs. The aim of this multicenter study was to evaluate the toxicological effects of serial microsampling on rats treated with azathioprine as a model drug with immunotoxic effects. Fifty microliters of blood were collected from the jugular vein of Sprague-Dawley rats at six time points from day 1 to 2 and 7 time points from day 27 to 28. The study was performed at three organizations independently. The microsampling effect on clinical signs, body weights, food consumption, hematological parameters, biochemical parameters, urinary parameters, organ weights, and tissue pathology was evaluated. Azathioprine-induced changes were observed in certain hematological and biochemical parameters and thymus weight and pathology. Microsampling produced minimal or no effects on almost all parameters; however, at 2 organizations, azathioprine-induced changes were apparently masked for two leukocytic, one coagulation, and two biochemical parameters. In conclusion, azathioprine toxicity could be assessed appropriately as overall profiles even with blood microsampling. However, microsampling may influence azathioprine-induced changes in certain parameters, especially leukocytic parameters, and its usage should be carefully considered.

The coronavirus disease 2019 (COVID-19) pandemic has been associated with high mortality worldwide. Owing to its complicated pathophysiology, diagnostic and prognostic biomarkers for effective patient management remain scarce. We analyzed kynurenine, tryptophan, and serotonin levels in the serum of patients with COVID-19 via liquid chromatography/mass spectrometry analysis. Serum serotonin levels were decreased in patients with more severe COVID-19, along with increased kynurenine and decreased tryptophan concentrations. Patients with moderate disease who subsequently worsened showed significantly lower serotonin concentrations compared with those who did not experience severe disease. Serum serotonin levels may represent a valuable biomarker for COVID-19 severity and prognosis.

The prognosis of patients with severe cases of COVID-19 is poor; thus, biomarkers for earlier prediction of COVID-19 progression are vital. We measured levels of five lung injury-related biomarkers, SP-D, KL-6, presepsin, kallistatin and stratifin, in serum samples collected serially during hospitalization from 31 patients with mild/moderate or severe/critical COVID-19 pneumonia, and their predictive performances were compared. Like the previously reported presepsin, a new biomarker candidate, stratifin, was significantly elevated with the onset of severe or critical symptoms in COVID-19 patients and decreased with symptom improvement. Notably, changes in stratifin and presepsin levels were distinctly earlier than those in SP-D, KL-6 and even SpO2/FiO2 values. Furthermore, serum levels of these biomarkers were significantly higher at the pre-severe stage (before the start of oxygen support) of patients who eventually advanced to severe/critical stages than in the patients who remained at the mild/moderate stage. These results were confirmed in an independent cohort, including 71 mild/moderate and 14 severe/critical patients, for whom the performance of stratifin and presepsin in discriminating between mild/moderate and pre-severe conditions of COVID-19 patients was superior to that of the SpO2/FiO2 ratio. Therefore, we concluded that stratifin and presepsin could be used as prognostic biomarkers for severe COVID-19 progression.

Background: The cross-reactive antibody response against seasonal human coronaviruses (HCoVs) was evaluated according to disease severity in patients with COVID-19 in Japan.Methods: In total, 194 paired serum samples collected from 97 patients with COVID-19 (mild, 35; severe, 62) were analyzed on admission and during convalescence. IgG antibodies against the nucleocapsid (N) and spike (S) proteins of SARS-CoV-2 and four seasonal HCoVs (HCoV-NL63, -229E, -OC43, and -HKU1) were detected by enzyme-linked immunosorbent assays.Results: There was no difference in optical density (OD) values for seasonal HCoVs on admission between the severe and mild cases. In addition, a specific pattern of disease severity-associated OD values for HCoVs was not identified. Significant increases in OD values from admission to convalescence for HCoV-HKU1and -OC43 IgG-S, and for HCoV-NL63 and -229E IgG-N were observed in the severe cases. Significant differences were observed between the mild and severe cases for HCoV-HKU1 and -OC43 IgG-S OD values during convalescence. Correlations were found between the fold changes for HCoV-OC43 IgG-S OD values, and for SARS-CoV-2 IgG-S OD values, and C-reactive protein, lactate dehydrogenase, and lymphocyte levels.Conclusion: There was no association between the antibody titer for seasonal HCoVs in the early phase of COVID-19 and disease severity.

A case of squamous cell carcinoma (SCC) of the head and neck that developed thrombosis inside the flap during the surgery and subendocardial infarction after the surgery is presented. An 82-year-old man was referred to our hospital because of nonhealing of a tooth extraction socket. The patient's medical history was unremarkable, with no ischemic heart disease. The preoperative blood tests and the physiological and imaging findings were also unremarkable other than an elevated hemoglobin A1c (HbA1c; 6.6%). He was diagnosed with SCC of the lower gingiva on biopsy. After intensive insulin therapy for glycemic control, tumor resection including mandibular segmental resection and neck dissection, followed by reconstruction with a vascularized fibular flap, was performed. Although there was visible thrombosis inside the fibular veins of the flap during the surgery, microvascular anastomosis and reconstruction were done without any complications after irrigation with heparin saline. Subendocardial infarction developed 10 hours after the surgery in the intensive care unit, but circulatory dynamics recovered after percutaneous coronary intervention. Two years after the surgery, the patient has had no decrease in basic activities of daily living, no local tumor recurrence, and no distant metastasis.

Astrocytes have shown longstanding promise as therapeutic targets for various central nervous system diseases. To facilitate drug development targeting astrocytes, we have recently developed a new conditionally immortalized human astrocyte cell line, termed HASTR/ci35 cells. In this study, in order to further increase their chances to contribute to various astrocyte studies, we report on the development of a culture method that improves HASTR/ci35 cell differentiation status and provide several proofs related to their astrocyte characteristics. The culture method is based on the simultaneous elimination of serum effects and immortalization signals. The results of qPCR showed that the culture method significantly enhanced several astrocyte marker gene expression levels. Using the differentiated HASTR/ci35, we examined their response profiles to nucleotide treatment and inflammatory stimuli, along with their membrane fatty acid composition. Consequently, we found that they responded to ADP or UTP treatment with a transient increase of intracellular Ca2+ concentration, and that they could show reactive response to interleukin-1β treatments. Furthermore, the membrane phospholipids of the cells were enriched with polyunsaturated fatty acids. To summarize, as a unique human astrocyte model carrying the capability of a differentiation induction properties, HASTR/ci35 cells are expected to contribute substantially to astrocyte-oriented drug development studies. Keywords: Astrocytes, Drug development, Immortalized cells, In vitro model, Central nervous system

Craniofacial deformities with multifactorial etiologies, such as cleft palate and facial dysmorphism, represent some of the most frequent congenital birth defects seen in humans. Their pathogeneses are often related to cranial neural crest (CNC) cells. During CNC cell migration, changes in cell shape and formation, as well as maintenance of subcellular structures, such as filopodia and lamellipodia, are dependent on the complex functions of Rho family small GTPases, which are regulators of actin cytoskeletal organization. Cdc42, a member of the Rho family of small GTPases, is known to play critical roles in organogenesis of various tissues. To investigate the physiological functions of Cdc42 during craniofacial development, we generated CNC-derived cell-specific inactivated Cdc42 mutant mice (Cdc42fl/fl;P0-cre). Most of the Cdc42fl/fl;P0-cre neonates were viable at birth, though they appeared weaker and no milk was found in their stomachs, and all died within a few days. They had a short face and intracranial bleeding, and abnormal calcification of the cranium. Cdc42fl/fl;P0-cre neonates also demonstrated a cleft palate and there was no fusion of the secondary palate because of failure of palatal shelf elongation for the process of palate closure. Cdc42 is crucial for facial and palatal formation during craniofacial development. Keywords: Cdc42, Cleft palate, Conditional knockout mice, P0-cre transgenic mice

Selenocysteine (Sec) insertion sequence-binding protein 2 (SBP2) is essential for the biosynthesis of Sec-containing proteins, termed selenoproteins. Subjects with mutations in the SBP2 gene have decreased levels of several selenoproteins, resulting in a complex phenotype. Selenoproteins play a significant role in antioxidative defense, and deficiencies in these proteins can lead to increased oxidative stress. However, lipid peroxidation and the effects of antioxidants in subjects with SBP2 gene mutations have not been studied. In the present study, we evaluated the lipid peroxidation products in the blood of a subject (the proband) with mutations in the SBP2 gene. We found that the proband had higher levels of free radical-mediated lipid peroxidation products, such as 7β-hydroxycholesterol, than the control subjects. Treatment of the proband with vitamin E (α-tocopherol acetate, 100 mg/day), a lipid-soluble antioxidant, for 2 years reduced lipid peroxidation product levels to those of control subjects. Withdrawal of vitamin E treatment for 7 months resulted in an increase in lipid peroxidation products. Collectively, these results clearly indicate that free radical-mediated oxidative stress is increased in the subject with SBP2 gene mutations and that vitamin E treatment effectively inhibits the generation of lipid peroxidation products.

Aging, also called senescence, is thought to be a physiological phenomenon that commonly occurs in various organs and tissues (Enoki et al., 2007 [1]). Many older adults experience dysfunction in their salivary glands, for example xerostomia, which is defined as dry mouth resulting from reduced or absent saliva flow (Nagler et al., 2004 [2]). In the present study, we investigated gene expression in submandibular glands of young (8 weeks old) and adult (50 weeks old) mice to analyze association of aging with gene expression profiling in mouse submandibular glands. Whole-genome gene expression profiles were analyzed using an Illumina Sentrix system with Mouse-WG-6 v.2 Expression BeadChips (Illumina). Of the genes screened, 284 showed detection values at a significance level of P < 0.01. Among those, the expression of 94 genes (33%) showed a greater decrease in adult mice as compared to young mice. On the other hand, that of 190 genes (77%) was increased in the adults more than in young mice. The data obtained in this study are publicly available in the Gene Expression Omnibus (GEO) database (accession number GSE66857).

Lipid peroxidation products have been known to induce cellular adaptive responses and enhance tolerance against subsequent oxidative stress through up-regulation of antioxidant compounds and enzymes. 24S-hydroxycholesterol (24SOHC) which is endogenously produced oxysterol in the brain plays an important role in maintaining brain cholesterol homeostasis. In this study, we evaluated adaptive responses induced by brain-specific oxysterol 24SOHC in human neuroblastoma SH-SY5Y cells. Cells treated with 24SOHC at sub-lethal concentrations showed significant reduction in cell death induced by subsequent treatment with 7-ketocholesterol (7KC) in both undifferentiated and retinoic acid-differentiated SH-SY5Y cells. These adaptive responses were also induced by other oxysterols such as 25-hydroxycholesterol and 27-hydroxycholesterol which are known to be ligands of liver X receptor (LXR). Co-treatment of 24SOHC with 9-cis retinoic acid, a retinoid X receptor ligand, enhanced the adaptive responses. Knockdown of LXRβ by siRNA diminished the adaptive responses induced by 24SOHC almost completely. The treatment with 24SOHC induced the expression of LXR target genes, such as ATP-binding cassette transporter A1 (ABCA1) and G1 (ABCG1). The 24SOHC-induced adaptive responses were significantly attenuated by siRNA for ABCG1 but not by siRNA for ABCA1. Taken together, these results strongly suggest that 24SOHC at sub-lethal concentrations induces adaptive responses via transcriptional activation of LXR signaling pathway, thereby protecting neuronal cells from subsequent 7KC-induced cytotoxicity.