Highlights d cGAS-STING activation and mitochondrial damage in tubules mediate acute kidney injury d cGAS-STING activation induces tubular inflammation and progression of AKI d Mitochondrial DNA leakage into the cytosol increased in AKIinduced tubular damage d Cytosolic mitochondrial DNA activates cGAS-STING signaling in tubular inflammation

Reverse genetics is a powerful tool to study single-stranded RNA viruses. Despite tremendous efforts having been made to improve the methodology for constructing flavivirus cDNAs, the cause of toxicity of flavivirus cDNAs in bacteria remains unknown. Here we performed mutational analysis studies to identify Escherichia coli promoter (ECP) sequences within nucleotides (nt) 1 to 3000 of the dengue virus type 2 (DENV2) and Japanese encephalitis virus (JEV) genomes. Eight and four active ECPs were demonstrated within nt 1 to 3000 of the DENV2 and JEV genomes, respectively, using fusion constructs containing DENV2 or JEV segments and empty vector reporter gene Renilla luciferase. The Flavivirus genus consists of more than 70 members that are categorized into several antigenic groups (46). Most flaviviruses are transmitted by mosquito or tick vectors and cause serious human and animal diseases (46). They include dengue virus (DENV), Japanese encephalitis virus (JEV), West Nile virus (WNV), yellow fever virus (YFV), and tick-borne encephalitis virus (TBEV). DENV and JEV cause some of the most serious arthropod-borne viral illnesses. There are four different serotypes of dengue virus, DENV1, DENV2, DENV3, and DENV4. Dengue cases have been reported in over 100 countries, and an estimated 2.5 billion people live in areas in which dengue is epidemic (26, 27, 49). DENV infection often leads to dengue fever, dengue hemorrhagic fever, and dengue shock syndrome (24,28,48). JEV transmission has been observed in the Southern Hemisphere and has the potential to become a worldwide public health threat. JEV can cause permanent neuropsychiatric sequelae and is sometimes fatal in children (56,60,61).Flaviviruses are enveloped RNA viruses that consist of single-stranded, positive-sense, 10.5-to 11-kb genomic RNA. The genome is associated with multiple copies of capsid proteins that are translated as a single polyprotein. After entering a host cell, the translated polyprotein is then cleaved into three structural proteins (C, prM, and E) and seven nonstructural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) by host proteases and a single virus-encoded protease to initiate viral replication (12,18). Introduction of flavivirus genomic RNA into susceptible cell lines can result in the production of infectious virus particles (1). This phenomenon has prompted the study of flavivirus virology via introduction of flavivirus genomic RNA that has been transcribed in vitro from fulllength flavivirus infectious cDNA.Reverse genetics is a powerful method for studying the viral replication of positive-strand RNA viruses (8). Unfortunately, the instability of full-length flavivirus cDNA in Escherichia coli has been a major hurdle in a attempt to construct flavivirus cDNAs (reviewed in references 4, 54, and 63). Several strategies have been developed to avoid or overcome the instability of infectious flavivirus cDNA. The instability of plasmids containing full-length YFV was avoided using an in vitro ligation approach involving two plasmids (5...

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The prevalence of chronic kidney disease (CKD), characterized by progressive renal dysfunction with tubulointerstitial fibrosis, is increasing because of societal aging. Uremic toxins, accumulated during renal dysfunction, cause kidney damage, leading to renal deterioration. A recent metabolomic analysis revealed that plasma D-serine accumulation is associated with faster progression of renal dysfunction in CKD patients. However, the causal relationship and the underlying mechanisms remain unclear. Herein, we demonstrated that D-serine markedly induced cellular senescence and apoptosis in a human proximal tubular cell line, HK-2, and primary culture of human renal tubular cells. The former was accompanied by G2/M cell cycle arrest and senescence-associated secretory phenotype, including pro-fibrotic and pro-inflammatory factors, contributing to tubulointerstitial fibrosis. Integrated stress response mediated by the general control nonderepressible 2 played an important role in D-serine-induced tubular cell toxicity and pro-fibrotic phenotypes, accelerating CKD progression and kidney aging. D-serine upregulated the L-serine synthesis pathway. Furthermore, D-serine-induced suppression of tubular cell proliferation was ameliorated by L-serine administration, indicating that D-serine exposure induced an L-serine-deprived state in tubular cells, compensated by L-serine synthesis. Thus, this study unveils molecular mechanisms underlying D-serine-induced tubular damage and pro-fibrotic phenotypes, suggesting that D-serine is a uremic toxin involved in CKD pathogenesis.

Dietary contents and their metabolites are closely related to chronic kidney disease (CKD) progression. Advanced glycated end products (AGEs) are a type of uremic toxin produced by glycation. AGE accumulation is not only the result of elevated glucose levels or reduced renal clearance capacity, but it also promotes CKD progression. Indoxyl sulfate, another uremic toxin derived from amino acid metabolism, accumulates as CKD progresses and induces tubulointerstitial fibrosis and glomerular sclerosis. Specific types of amino acids (d-serine) or fatty acids (palmitate) are reported to be closely associated with CKD progression. Promising therapeutic targets associated with nutrition include uremic toxin absorbents and inhibitors of AGEs or the receptor for AGEs (RAGE). Probiotics and prebiotics maintain gut flora balance and also prevent CKD progression by enhancing gut barriers and reducing uremic toxin formation. Nrf2 signaling not only ameliorates oxidative stress but also reduces elevated AGE levels. Bardoxolone methyl, an Nrf2 activator and NF-κB suppressor, has been tested as a therapeutic agent, but the phase 3 clinical trial was terminated owing to the high rate of cardiovascular events. However, a phase 2 trial has been initiated in Japan, and the preliminary analysis reveals promising results without an increase in cardiovascular events.