The authors concluded that Ccl2(-/-)/Cx3cr1(-/-) mice develop a broad spectrum of AMD abnormalities with early onset and high penetrance. These observations implicate certain chemokines and endoplasmic reticulum proteins in AMD pathogenesis. Similar to the mechanism of neurodegeneration caused by dysfunction of endoplasmic reticulum proteins, decreased chaperoning may cause misfolded protein accumulation, leading to drusen formation and retinal degeneration.

There is a significant genetic component associated with the risk of developing age-related macular degeneration (AMD), a leading cause of blindness among older people in Western countries. Based on the recognized role of CX3CR1, a chemokine receptor, in other age-related complex diseases and the possible involvement of the immune system in AMD pathogenesis, we tested for an association between CX3CR1 sequence variation/expression and AMD by screening two CX3CR1 single nucleotide polymorphisms (SNPs), V249I and T280M, among AMD patients and controls, and determining the level of CX3CR1 expression in AMD and normal eye tissues. PRINCIPAL FINDINGS1. An association between CX3CR1 1249/M280 alleles and AMD A significant increased prevalence of M280 and 1249 carriers or alleles was found among AMD cases vs. controls. Distribution of CX3CR1V249I and T280M did not deviate from Hardy-Weinberg equilibrium in any group (Table 1). A quantitative correlation between CX3CR1 1249/M280 and the prevalence of AMD phenotypeAs indicated in Table 1, the odds ratios (OR) of clinically diagnosed AMD compared with controls were 1.86 and 1.94 in terms of carrying 1249 and M280, respectively; however, the OR of pathologically diagnosed AMD reached 3.57 in terms of carrying M280. A higher OR was found for the clinically diagnosed AMD group compared with the aged controls whereas a lower OR was observed for the AMD group compared with the blood donor population of a much younger age. Allele frequency analysis yielded similar results (see full text version).3. Lower expression of CX3CR1 in the AMD maculae from individual bearing CX3CR1 T/M280 compared with individual bearing CX3CR1 T/T280

Background/Aims: Senescent Ccl2^–/– mice develop cardinal features of human age-related macular degeneration (AMD). Loss-of-function single-nucleotide polymorphisms within CX3CR1 are associated with AMD. Methods: We generated Ccl2^–/–/Cx3cr1^–/– [double-knockout (DKO)] mice and evaluated the eyes using fundoscopy routine histology, immunochemistry, biochemistry and proteomics. Results: At 6 weeks old, all DKO mice developed AMD-like retinal lesions such as abnormal retinal pigment epithelium cells, drusen, photoreceptor atrophy and choroidal neovascularization, which progressed with age and reversed with high omega-3 long-chain polyunsaturated fatty acid diet. N-retinylidene-N-retinylethanolamine (A2E), a major lipofuscin fluorophore, illustrated by an emission peak at ∼600 nm, was significantly higher in DKO retinal pigment epithelium. Decreased ERp29 was found in the retina of DKO mice. Conclusion: A broad spectrum of AMD pathologies with early onset and high penetrance in these mice implicate certain chemokines, A2E and endoplasmic reticulum proteins in AMD pathogenesis.

everse-transcriptase quantitative polymerase chain reaction (RT-qPCR) analysis of respiratory samples is the gold standard for coronavirus disease 2019 (COVID-19) diagnosis 1 , but it has limitations. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 RNA) levels in the upper respiratory tract rapidly decrease after infection 2 while lower respiratory tract levels remain high 3 . RT-qPCR assays performed after early SARS-CoV-2 infection may thus yield false negatives, but infection events are often unclear, complicating interpretation. Nasopharyngeal tissue highly expresses angiotensin-converting enzyme 2 (ref. 4 ), the primary receptor for SARS-CoV-2 (ref. 5 ), but this protein is also expressed in other tissues (for example, cardiac and small intestine 6 ) reported to develop SARS-CoV-2 infections and related pathology 7 . Nasopharyngeal RT-qPCR assay results thus may not accurately reflect the status of lower respiratory tract or extrapulmonary infections.Circulating SARS-CoV-2 RNA detectable in mild to severe COVID-19 correlates with and predicts disease severity 8-10 , and appears responsible for extrapulmonary infections 7 . However, RT-qPCR exhibits poor overall sensitivity for SARS-CoV-2 RNA in plasma (≤41%) 11 . More sensitive and robust blood-based SARS-CoV-2 RNA assays compatible with routine clinical tests could thus improve COVID-19 diagnosis and prognostic evaluation. However, only two studies employing droplet digital RT-PCR, which is not suited for clinical use, have used ultrasensitive approaches to detect SARS-CoV-2 RNA in serum or plasma.Infected cells can secrete extracellular vesicles (EVs) containing pathogen-derived factors 12,13 , protecting these factors from hydrolases and allowing them to accumulate in the circulation 14 . Hepatitis

This study investigates age-related macular degeneration (AMD) genetic risk factors through identification of a functional singlenucleotide polymorphism (SNP) and its disease association. We chose ERCC6 because of its roles in the aging process, DNA repair, and ocular degeneration from the gene disruption. Bioinformatics indicated a putative binding-element alteration on the sequence containing C؊6530>G SNP in the 5 flanking region of ERCC6 from Sp1 on the C allele to SP1, GATA-1, and OCT-1 on the G allele. Electrophoretic mobility shift assays displayed distinctive C and G allele-binding patterns to nuclear proteins. Luciferase expression was higher in the vector construct containing the G allele than that containing the C allele. A cohort of 460 advanced AMD cases and 269 age-matched controls was examined along with pathologically diagnosed 57 AMD and 18 age-matched non-AMD archived cases. ERCC6 C؊6530>G was associated with AMD susceptibility, both independently and through interaction with an SNP (rs380390) in the complement factor H (CFH) intron reported to be highly associated with AMD. A disease odds ratio of 23 was conferred by homozygozity for risk alleles at both ERCC6 and CFH compared with homozygozity for nonrisk alleles. Enhanced ERCC6 expression was observed in lymphocytes from healthy donors bearing ERCC6 C؊6530>G alleles. Intense immunostaining of ERCC6 was also found in AMD eyes from ERCC6 C؊6530>G carriers. The strong AMD predisposition conferred by the ERCC6 and CFH SNPs may result from biological epistasis, because ERCC6 functions in universal transcription as a component of RNA pol I transcription complex.Cockayne syndrome ͉ single nucleotide polymorphism ͉ gene regulation ͉ interaction ͉ DNA repair A ge-related macular degeneration (AMD) is a leading cause of visual impairment and blindness in Western countries among the elderly (1). AMD is a significant health problem in the United States, with a current estimate of Ϸ1.75 million persons with advanced AMD in the general population and Ϸ7.3 million people with early stages of AMD defined by large retinal drusen. By the year 2020, it is estimated that Ϸ2.95 million people will suffer advanced AMD, and an additional 6.4 million white individuals will have the early stages of AMD in at least one eye (2).The etiology of AMD remains elusive. To date, age and cigarette smoking have been identified as AMD risk factors (3-8). Various studies have indicated a significant genetic contribution to AMD risk (9). For example, it has been reported that advanced AMD is more prevalent in Caucasian populations as compared with other races that possess higher levels of uveal pigmentation and skin melanin (10). Furthermore, the population-attributed AMD risk related to genetic factors was 23% (11, 12). First-degree relatives of patients with late AMD developed the disease at an increased rate at a relatively young age (12, 13). The higher occurrence of AMD among monozygotic twins and first-degree relatives of AMD patients as compared with spouses and unrelated ...

E-cigarettes are portrayed as safer relative to conventional tobacco. However, burgeoning evidence suggests that E-cigarettes may adversely affect host defenses. However, the precise mechanisms by which E-cigarette vapor alters innate immune cell function have not been fully elucidated. We determined the effects of E-cigarette exposure on the function and responses to infectious challenge of the most abundant innate immune cell, the neutrophil, using isolated human neutrophils and a mouse model of gram-negative infection. Our results revealed that human neutrophils exposed to E-cigarette vapor had 4.2-fold reductions in chemotaxis toward the bacterial cell-well component f-Met-Leu-Phe ( P < 0.001). F-actin polarization and membrane fluidity were also adversely affected by E-cigarette vapor exposure. E-cigarette-exposed human neutrophils exhibited a 48% reduction in production of reactive oxygen species (ROS; P < 0.001). Given the central role of ROS in neutrophil extracellular trap (NET) production, NET production was quantified, and E-cigarette vapor exposure was found to reduce NETosis by 3.5-fold ( P < 0.01); formulations with and without nicotine containing propylene glycol exhibiting significant suppressive effects. However, noncanonical NETosis was unaffected. In addition, exposure to E-cigarette vapor lowered the rate of phagocytosis of bacterial bioparticles by 47% ( P < 0.05). In our physiological mouse model of chronic E-cigarette exposure and sepsis, E-cigarette vapor inhalation led to reduced neutrophil migration in infected spaces and a higher burden of Pseudomonas. These findings provide evidence that E-cigarette use adversely impacts the innate immune system and may place E-cigarette users at higher risk for dysregulated inflammatory responses and invasive bacterial infections.

Age-related macular degeneration (AMD) is the leading cause of blindness in the United States. Ccl2 knock-out (KO) mice sporadically develop the cardinal features of AMD in their senescent stage. Humans bearing a loss of function variant or single nucleotide polymorphism (SNP) in CX3CR1 are at increased risk of developing AMD. We recently developed Ccl2(-/-)/Cx3cr1(-/-) mice, which consistently develop retinal degeneration with many AMD features. Since there is strong evidence for an immunological role in AMD pathogenesis, we examined ocular immune protein expression levels in Ccl2(-/-)/Cx3cr1(-/-), Ccl2(-/-), Cx3cr1(-/-), and age-matched wild-type (WT) mice. Immunohistochemistry revealed increased complement C3d in Bruch's membrane, retinal pigment epithelium (RPE), choroidal capillaries, and particularly drusen of the Ccl2(-/-)/Cx3cr1(-/-) mice relative to the WT controls. No change was detected in single KO mice. Real-time RT-PCR revealed a 2.5-fold increase in C3 expression in the Ccl2(-/-)/Cx3cr1(-/-). While the retinas of four month old WT and Ccl2(-/-) showed minimal immunoreactivity for markers of macrophages and microglia, infiltrates of these mononuclear phagocytic cells were detected in the Ccl2(-/-)/Cx3cr1(-/-)retinal lesions and a few foci in the Cx3cr1(-/-) retina. The Ccl2(-/-)/Cx3cr1(-/-) had reduced toll-like receptor 4 (TLR4) expression in the RPE. Following LPS injection, the Ccl2(-/-)/Cx3cr1(-/-) had significantly reduced endotoxin-induced uveitis scores and showed a diminished increase in Tlr4 mRNA expression. No changes in TLR4 expression were detected in either single KO. Autoantibodies against the retina and photoreceptors were also detected in the Ccl2(-/-)/Cx3cr1(-/-) serum. Real-time RT-PCR revealed significant increases in Ccl5 transcript in the Ccl2(-/-)/Cx3cr1(-/-) relative to the WT. These results suggest that innate immunity and possibly adaptive immunity play an important role in Ccl2(-/-)/Cx3cr1(-/-) retinal degeneration. Moreover, since human AMD patients show similar immunopathological profiles, these results support the Ccl2(-/-)/Cx3cr1(-/-) as a suitable model for human AMD.

Age-related macular degeneration (AMD) is a leading cause of blindness in the United States and developed countries. Although the etiology and pathogenesis of AMD remain unknown, a complex interaction of genetic and environmental factors is thought to exist. The incidence and progression of all of the features of AMD are known to increase significantly with age. The tendency for familial aggregation and the findings of gene variation association studies implicate a significant genetic component in the development of AMD. This review summarizes in detail the AMD-related genes identified by studies on genetically engineered and spontaneously gene-mutated (naturally mutated) animals, AMD chromosomal loci identified by linkage studies, AMD-related genes identified through studies of monogenic degenerative retinal diseases, and AMD-related gene variation identified by association studies.