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Abstract Bone homeostasis is maintained by an intricate balance between osteoclasts and osteoblasts, which becomes disturbed in osteoporosis. Metallothioneins (MTs) are major contributors in cellular zinc regulation. However, the role of MTs in bone cell regulation has remained unexplored. Single-cell RNA sequencing analysis discovered that, unlike the expression of other MT members, the expression of MT3 was unique to osteoclasts among various macrophage populations and was highly upregulated during osteoclast differentiation. This unique MT3 upregulation was validated experimentally and supported by ATAC sequencing data analyses. Downregulation of MT3 by gene knockdown or knockout resulted in excessive osteoclastogenesis and exacerbated bone loss in ovariectomy-induced osteoporosis. Transcriptome sequencing of MT3 knockdown osteoclasts and gene set enrichment analysis indicated that the oxidative stress and redox pathways were enriched, which was verified by MT3-dependent regulation of reactive oxygen species (ROS). In addition, MT3 deficiency increased the transcriptional activity of SP1 in a manner dependent on intracellular zinc levels. This MT3-zinc-SP1 axis was crucial for the control of osteoclasts, as zinc chelation and SP1 knockdown abrogated the promotion of SP1 activity and osteoclastogenesis by MT3 deletion. Moreover, SP1 bound to the NFATc1 promoter, and overexpression of an inactive SP1 mutant negated the effects of MT3 deletion on NFATc1 and osteoclastogenesis. In conclusion, MT3 plays a pivotal role in controlling osteoclastogenesis and bone metabolism via dual axes involving ROS and SP1. The present study demonstrated that MT3 elevation is a potential therapeutic strategy for osteolytic bone disorders, and it established for the first time that MT3 is a crucial bone mass regulator.

Abstract Osimertinib, a selective third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), effectively targets the EGFR T790M mutant in non-small cell lung cancer (NSCLC). However, the newly identified EGFR C797S mutation confers resistance to osimertinib. In this study, we explored the role of pyruvate dehydrogenase kinase 1 (PDK1) in osimertinib resistance. Patients exhibiting osimertinib resistance initially displayed elevated PDK1 expression. Osimertinib-resistant cell lines with the EGFR C797S mutation were established using A549, NCI-H292, PC-9, and NCI-H1975 NSCLC cells for both in vitro and in vivo investigations. These EGFR C797S mutant cells exhibited heightened phosphorylation of EGFR, leading to the activation of downstream oncogenic pathways. The EGFR C797S mutation appeared to increase PDK1-driven glycolysis through the EGFR/AKT/HIF-1α axis. Combining osimertinib with the PDK1 inhibitor leelamine helped successfully overcome osimertinib resistance in allograft models. CRISPR-mediated PDK1 knockout effectively inhibited tumor formation in xenograft models. Our study established a clear link between the EGFR C797S mutation and elevated PDK1 expression, opening new avenues for the discovery of targeted therapies and improving our understanding of the roles of EGFR mutations in cancer progression.

Abstract The clinical progression of neurodegenerative diseases correlates with the spread of proteinopathy in the brain. The current understanding of the mechanism of proteinopathy spread is far from complete. Here, we propose that inflammation is fundamental to proteinopathy spread. A sequence variant of α-synuclein (V40G) was much less capable of fibril formation than wild-type α-synuclein (WT-syn) and, when mixed with WT-syn, interfered with its fibrillation. However, when V40G was injected intracerebrally into mice, it induced aggregate spreading even more effectively than WT-syn. Aggregate spreading was preceded by sustained microgliosis and inflammatory responses, which were more robust with V40G than with WT-syn. Oral administration of an anti-inflammatory agent suppressed aggregate spreading, inflammation, and behavioral deficits in mice. Furthermore, exposure of cells to inflammatory cytokines increased the cell-to-cell propagation of α-synuclein. These results suggest that the inflammatory microenvironment is the major driver of the spread of synucleinopathy in the brain.

Abstract The International Myeloma Working Group has recently revised the diagnostic criteria for primary plasma cell leukemia (PCL) to circulating plasma cells (CPCs) ≥ 5% in a peripheral blood smear. The present study validated new criteria in patients with multiple myeloma or PCL diagnosed using the previous diagnostic criteria, who were administered immunomodulatory drugs or proteasome inhibitors as induction therapy. We analyzed the medical records of 1357 patients from eight hospitals in South Korea. The median age of the all patients was 64 years, and 187 (13.8%) had CPCs at diagnosis. Only 79 (5.8%) of the patients had ≥ 5% CPCs. The median overall survival (OS) of patients with CPCs ≥ 5% and ≥ 20% was similar, but had significantly inferior median progression-free survival (PFS) and median OS than those with CPCs < 5% (13.1 vs. 21.5 months, P

Abstract Cell-to-cell propagation of α-synuclein is thought to be the underlying mechanism of Parkinson’s disease progression. Recent evidence suggests that inflammation plays an important role in the propagation of protein aggregates. However, the mechanism by which inflammation regulates the propagation of aggregates remains unknown. Here, using in vitro cultures, we found that soluble factors secreted from activated microglia promote cell-to-cell propagation of α-synuclein and further showed that among these soluble factors, TNF-α had the most robust stimulatory activity. Treatment of neurons with TNF-α triggered cellular senescence, as shown by transcriptomic analyses demonstrating induction of senescence-associated genes and immunoanalysis of senescence phenotype marker proteins. Interestingly, secretion of α-synuclein was increased in senescent neurons, reflecting acquisition of a senescence-associated secretory phenotype (SASP). Using vacuolin-1, an inhibitor of lysosomal exocytosis, and RNAi against rab27a, we demonstrated that the SASP was mediated by lysosomal exocytosis. Correlative light and electron microscopy and immunoelectron microscopy confirmed that propagating α-synuclein aggregates were present in electron-dense lysosome-like compartments. TNF-α promoted the SASP through stimulation of lysosomal exocytosis, thereby increasing the secretion of α-synuclein. Collectively, these results suggest that TNF-α is the major inflammatory factor that drives cell-to-cell propagation of α-synuclein by promoting the SASP and subsequent secretion of α-synuclein.

Bone remodeling: Regulatory protein identified A critical mechanism for maintaining bone health uncovered by scientists in South Korea could provide insights into bone disease development. Bone remodeling is a lifetime process of bone generation that ensures bones remain healthy. Osteoclasts (OC), cells that break down bone, differentiate from white blood cell populations. Disruption to OC formation and function plays a critical role in bone diseases, yet the regulatory mechanisms in OC generation are unclear. Hong-Hee Kim at Seoul National University and co-workers investigated the role of a protein called DCTN1, which is involved in skeletal assembly processes. The team found that inhibiting DCTN1 suppressed the expression of key proteins needed for OC formation in cell cultures and mouse models. Overexpressing DCTN1 was equally damaging, suggesting the protein plays a key regulatory role.

Kidney disease: stem cells to the rescue Stem cells can halt the progression of kidney damage owing to diabetes by reducing inflammation and improving energy production in kidney cells. Eun Hee Koh at the University of Ulsan College of Medicine in Seoul, South Korea, and colleagues found that adult stem cells, known as mesenchymal stem cells (MSCs), derived from human umbilical cord blood had a protective effect on the kidneys of diabetic mice. Repeated administration of MSCs prevented the recruitment of pro-inflammatory cells into the kidney and increased the levels of arginase-1, a marker of cells with anti-inflammatory activity. Experiments in cells showed that MSCs stimulated the production of arginase-1 in that, in turn, were able to increase the production and activity of mitochondria in kidney cells. This study confirms an important role for MSCs in organ repair.

Atopic dermatitis (AD) presents significant therapeutic challenges due to its poorly understood etiology. Eosinophilia, a hallmark of allergic inflammation, is implicated in AD pathogenesis. Interleukin-10 (IL-10)-producing regulatory B (Breg) cells exhibit potent anti-inflammatory effects. However, their role in controlling AD-related eosinophilia is not well understood. To investigate the impact of eosinophils on AD, we employed IL-5Rα-deficient (Il5ra -/- ) mice, which lack functional eosinophils. Induction of AD in these mice resulted in attenuated disease symptoms, underscoring the critical role of eosinophils in AD development. Additionally, the adoptive transfer of purified Breg cells into mice with AD significantly alleviated disease severity. Mechanistic studies revealed that IL-10 produced by Breg cells directly inhibits eosinophil activation and infiltration into the skin. In vitro experiments further confirmed that Breg cells inhibited eosinophil peroxidase secretion in an IL-10-dependent manner. Our collective findings demonstrate that IL-10 from Breg cells alleviates AD by suppressing eosinophil activation and tissue infiltration. This study elucidates a novel regulatory mechanism of Breg cells, providing a foundation for future Breg-mediated therapeutic strategies for AD., (© 2024. The Author(s).)

Innate lymphoid cells (ILCs) play an important role in maintaining tissue homeostasis and various inflammatory responses. ILCs are typically classified into three subsets, as is the case for T-cells. Recent studies have reported that IL-10-producing type 2 ILCs (ILC2 10 s) have an immunoregulatory function dependent on IL-10. However, the surface markers of ILC2 10 s and the role of ILC2 10 s in contact hypersensitivity (CHS) are largely unknown. Our study revealed that splenic ILC2 10 s are extensively included in PD-L1 high Sca-1 + ILCs and that IL-27 amplifies the development of PD-L1 high Sca-1 + ILCs and ILC2 10 s. Adoptive transfer of PD-L1 high Sca-1 + ILCs suppressed oxazolone-induced CHS in an IL-10-dependent manner Taken together, our results demonstrate that ILC2 10 s are critical for the control of CHS and suggest that ILC2 10 s can be used as target cells for the treatment of CHS., (© 2024. The Author(s).)

Huntington's disease is caused by CAG trinucleotide expansions in the gene encoding huntingtin. N- terminal fragments of huntingtin with polyglutamine produce aggregates in the endosome-lysosomal system, where proteolytic fragments of huntingtin is generated. Heat shock protein 70 (HSP70) prevents the formation of protein aggregates, but the effect of HSP70 on the huntingtin in the endosome-lysosomal system is unknown. This study was to determine whether HSP70 alters the distribution of huntingtin in endosome-lysosomal system. HSP70 expressing stable cells (NIH/3T3 or cerebral hybrid cell line A1) were generated, and mutant [(CAG)(100)] huntingtin was transiently overexpressed. Analysis of subcellular distribution by immunocytochemistry or proteolysis cleavage by Western blotting was performed. 18 CAG repeat wild type [WT; (CAG)(18)] huntingtin was used as a control. Cells with huntingtin showed patterns of endosome-lysosomal accumulation, from a "dispersed vacuole (DV)" type into a coalescent "perinuclear vacuole (PV)" type over time. In WT huntingtin, HSP70 increased the cells with the PV types that enhanced the proteolytic cleavage of huntingtin. However, HSP70 reduced cells of the DV and PV types expressing mutant huntingtin, that result in less proteolysis than that of control. In addition, intranuclear inclusions were formed only in mutant cells, which was not affected by HSP70. These results suggest that HSP70 alters the distribution of huntingtin in the endosome- lysosomal system, and that this contributes to huntingtin proteolysis.