Structure and assembly of the human IL-12 signaling complex.

Interleukin (IL)-12 is a heterodimeric pro-inflammatory cytokine. Our cryoelectron microscopy structure determination of human IL-12 in complex with IL-12Rß1 and IL-12Rß2 at a resolution of 3.75 Å reveals that IL-12Rß2 primarily interacts with the IL-12p35 subunit via its N-terminal Ig-like domain, while IL-12Rß1 binds to the p40 subunit with its N-terminal fibronectin III domain. This binding mode of IL-12 with its receptors is similar to that of IL-23 but shows notable differences with other cytokines. Through structural information and biochemical assays, we identified Y62, Y189, and K192 as key residues in IL-12p35, which bind to IL-12Rß2 with high affinity and mediate IL-12 signal transduction. Furthermore, structural comparisons reveal two distinctive conformational states and structural plasticity of the heterodimeric interface in IL-12. As a result, our study advances our understanding of IL-12 signal initiation and opens up new opportunities for the engineering and therapeutic targeting of IL-12.

Structural basis for urate recognition and apigenin inhibition of human GLUT9.

Urate, the physiological form of uric acid and a potent antioxidant in serum, plays a pivotal role in scavenging reactive oxygen species. Yet excessive accumulation of urate, known as hyperuricemia, is the primary risk factor for the development of gout. The high-capacity urate transporter GLUT9 represents a promising target for gout treatment. Here, we present cryo-electron microscopy structures of human GLUT9 in complex with urate or its inhibitor apigenin at overall resolutions of 3.5 Å and 3.3 Å, respectively. In both structures, GLUT9 exhibits an inward open conformation, wherein the substrate binding pocket faces the intracellular side. These structures unveil the molecular basis for GLUT9's substrate preference of urate over glucose, and show that apigenin acts as a competitive inhibitor by occupying the substrate binding site. Our findings provide critical information for the development of specific inhibitors targeting GLUT9 as potential therapeutics for gout and hyperuricemia.

Structural mechanism of bacteriophage lambda tail's interaction with the bacterial receptor.

Bacteriophage infection, a pivotal process in microbiology, initiates with the phage's tail recognizing and binding to the bacterial cell surface, which then mediates the injection of viral DNA. Although comprehensive studies on the interaction between bacteriophage lambda and its outer membrane receptor, LamB, have provided rich information about the system's biochemical properties, the precise molecular mechanism remains undetermined. This study revealed the high-resolution cryo-electron microscopy (cryo-EM) structures of the bacteriophage lambda tail complexed with its irreversible Shigella sonnei 3070 LamB receptor and the closed central tail fiber. These structures reveal the complex processes that trigger infection and demonstrate a substantial conformational change in the phage lambda tail tip upon LamB binding. Providing detailed structures of bacteriophage lambda infection initiation, this study contributes to the expanding knowledge of lambda-bacterial interaction, which holds significance in the fields of microbiology and therapeutic development.

Architecture of the bacteriophage lambda tail.

Bacteriophage lambda has a double-stranded DNA genome and a long, flexible, non-contractile tail encoded by a contiguous block of 11 genes downstream of the head genes. The tail allows host recognition and delivery of viral DNA from the head shell to the cytoplasm of the infected cell. Here, we present a high-resolution structure of the tail complex of bacteriophage lambda determined by cryoelectron microscopy. Most component proteins of the lambda tail were determined at the atomic scale. The structure sheds light on the molecular organization of the extensively studied tail of bacteriophage lambda.

Body mass index impacts ectopic pregnancy during in vitro fertilization: an analysis of 42,362 clinical pregnancy cycles.

PURPOSE: This large, single-center, retrospective cohort study was aimed to explore the effect of female body mass index (BMI) on ectopic pregnancy (EP) following fresh and frozen-thawed embryo transfers (ET). METHODS: A total of 27,600 pregnancies after fresh ET and 14,762 pregnancies after frozen-thawed ET were included between January 2010 to June 2022. Women were divided into three groups based on BMI according to the Working Group on Obesity in China (WGOC), International Life Sciences Institute (ILSI): underweight (BMI < 18.5 kg/m2), normal weight (BMI, 18.5-23.9 kg/m2), and overweight or obesity (≥ 24 kg/m2). Compare EP rates among BMI categories in fresh and frozen-thawed ET cycles respectively. Multivariate logistic regression analyses were used to investigate the association between female BMI and EP. RESULTS: The overall EP rates in fresh, and frozen thawed transfer cycles were 2.43% (672/27,600) and 2.82% (417/14,762), respectively. In fresh ET cycles, underweight women yielded a significantly higher EP rate than those with normal and excess weight (3.29% vs. 2.29% vs. 2.54%, P = 0.029). But EP rates did not differ among the three BMI groups (2.72% vs. 2.76% vs. 2.96%, P = 0.782) in frozen-thawed ET cycles. In fresh ET cycles, after adjusting for potential confounding factors, no significant association was found between female BMI and EP occurrence (adjusted OR: 0.98, 95% CI 0.70-1.37, P = 0.894, for BMI 18.5-23.9 kg/m2; adjusted OR: 0.89, 95% CI 0.75-1.06, P = 0.205, for BMI ≥ 24 kg/m2. Reference = BMI < 18.5 kg/m2). CONCLUSION(S): Female BMI did not affect the occurrence of ectopic pregnancy in either fresh or frozen-thawed embryo transfer cycles.

Duration of late-follicular elevated progesterone and in vitro fertilization outcomes in pituitary down-regulation treatment cycles.

Background: The objective of this study was to explore whether the duration of LFEP (late-follicular elevated progesterone) affected pregnancy outcomes in IVF (in vitro fertilization) patients treated with pituitary downregulation protocols. Method: Patients with their first IVF/ICSI cycles between January 2016 and December 2016 were included. LFEP was set either at P > 1.0ng/ml or P > 1.5ng/ml. Clinical pregnancy rate was compared among three different groups (no LFEP; LFEP for 1 day; LFEP for ≥ 2 days). Then multivariate logistic regression analysis was performed to explore the influencing factors of clinical pregnancy rate. Results: This retrospective analysis involved 3,521 first IVF/ICSI cycles with fresh embryo transfers. Clinical pregnancy rate was the lowest in patients with a LFEP duration of ≥ 2 days, irrespective of whether LFEP was defined as P > 1.0 ng/ml (68.79% vs. 63.02% vs. 56.20%; P = 0.000) or as P > 1.5 ng/ml (67.24% vs. 55.95% vs. 45.51%; P = 0.000). In addition, LFEP duration was significantly associated with clinical pregnancy outcomes in unadjusted logistic regression analysis. However, in multivariate regression models after adjusting confounders, adjusted OR for LFEP duration (≥ 2 days) in the two models was 0.808 (P = 0.064; LFEP as P > 1.0 ng/ml) and 0.720 (P = 0.098; LFEP as P > 1.5 ng/ml), respectively. Conclusion: LFEP adversely affects clinical pregnancy outcomes. However, the duration of LFEP seems to have no influence on the clinical pregnancy rate in pituitary downregulation treatment cycles.

Effect of blastocyst morphology on the incidence of monozygotic twinning pregnancy after single blastocyst transfer.

Aims: To explore whether blastocyst morphology [blastocyst stage, inner cell mass (ICM), and trophectoderm (TE) grading] impacts the occurrence of monozygotic twinning (MZT) after single blastocyst transfer (SBT).Materials and methods: A single-center retrospective cohort study was conducted including all clinical pregnancies after single blastocyst transfer between January 2015 and September 2021 (n = 9229). Blastocyst morphology was assessed using Gardner grading system. MZT was defined as more than one gestational sac (GS), or two or more fetal heartbeats in a single GS via ultrasound at 5-6 gestational weeks.Results: The overall MZT rate was 2.46% (227 of 9229 cases), of which was the highest in blastocysts of grade A TE and lowest in those with grade C TE (grade A: B:C = 3.40%:2.67%:1.58%, p = .002). Higher risk of MZT pregnancy was associated with higher trophectoderm grading [A vs. C: aOR, 1.883, 95% CI 1.069-3.315, p = .028; B vs C: aOR, 1.559, 95% CI 1.066-2.279, p = .022], but not extended culture in vitro (day 5 vs. day 6), vitrification (fresh vs. frozen-thawed ET), assisted hatching (AH), blastocyst stage (stage 1-6) or ICM grading (A vs. B).Conclusions: We conclude that TE grade is an independent risk factor of MZT after single blastocyst transfer. Blastocysts with high-grade trophectoderm are more liable to obtain monozygotic multiple gestation.

Polycystic ovary syndrome increases the rate of early spontaneous miscarriage in women who have undergone single vitrified euploid blastocyst transfer.

RESEARCH QUESTION: Does polycystic ovary syndrome (PCOS) independently influence the risk of early spontaneous miscarriage in patients undergoing single euploid vitrified blastocyst transfer? DESIGN: This observational cohort study retrospectively analysed 1498 patients undergoing their first single euploid blastocyst frozen transfer cycles between October 2016 and December 2021. Patients were divided into PCOS and non-PCOS groups according to the Rotterdam criteria. Logistic regression analysis was conducted to study the independent effect of maternal PCOS status on early spontaneous miscarriage after single euploid embryo transfer after adjusting for confounding factors. RESULTS: No statistically significant differences were identified in the rates of positive pregnancy test (68.95% versus 64.86%, P = 0.196) or clinical pregnancy (59.93% versus 57.33%, P = 0.429) between the PCOS and non-PCOS groups after single euploid embryo transfer. Early spontaneous miscarriage occurred more frequently in women with PCOS compared with controls (18.67% versus 12.00%, P = 0.023). In single euploid embryo transfer cycles, PCOS significantly increased the incidence of early spontaneous miscarriage after adjusting for some potential confounders (adjusted odds ratio 1.649, 95% CI 1.032 to 2.635, P = 0.036). CONCLUSIONS: Although no significant difference was observed in clinical pregnancy rates, PCOS status increased the risk of early spontaneous miscarriage after single vitrified euploid blastocyst transfer, suggesting an additional role of endometrial dysfunction affected by endocrine disorders. Further studies are needed to investigate the specific mechanisms and effective intervention strategies.

Identification of an intraocular microbiota.

The current dogma in ophthalmology and vision research presumes the intraocular environment to be sterile. However, recent evidence of intestinal bacterial translocation into the bloodstream and many other internal organs including the eyes, found in healthy and diseased animal models, suggests that the intraocular cavity may also be inhabited by a microbial community. Here, we tested intraocular samples from over 1000 human eyes. Using quantitative PCR, negative staining transmission electron microscopy, direct culture, and high-throughput sequencing technologies, we demonstrated the presence of intraocular bacteria. The possibility that the microbiome from these low-biomass communities could be a contamination from other tissues and reagents was carefully evaluated and excluded. We also provide preliminary evidence that a disease-specific microbial signature characterized the intraocular environment of patients with age-related macular degeneration and glaucoma, suggesting that either spontaneous or pathogenic bacterial translocation may be associated with these common sight-threatening conditions. Furthermore, we revealed the presence of an intraocular microbiome in normal eyes from non-human mammals and demonstrated that this varied across species (rat, rabbit, pig, and macaque) and was established after birth. These findings represent the first-ever evidence of intraocular microbiota in humans.

Topical nanoparticles interfering with the DNA-LL37 complex to alleviate psoriatic inflammation in mice and monkeys.

Cell-free DNA (cfDNA) released from damaged or dead cells combines with LL37 and is converted into an immune response activator to exacerbate psoriasis. Here, we show that cationic nanoparticles (cNPs) efficiently compete for DNA from the DNA-LL37 immunocomplex and inhibit DNA-LL37-induced cell activation. Using phenotypical images, psoriasis area and severity index scoring, histology, and immunohistochemical analysis, we demonstrate that topical application of cNPs on psoriasiform skin of a mouse model relieves psoriatic symptoms. It is noteworthy that the results were confirmed in a cynomolgus monkey model. Moreover, topically administrated cNPs showed low in vivo toxicity because of their retention in skin. Mechanistic analyses of cytokine expression in the psoriatic site, cfDNA levels in circulation and inflamed skin, skin permeation, and biodistribution of cNPs also matched the therapeutic outcomes. Therefore, we present a previously unidentified strategy of nanomedicine to treat skin inflammatory diseases, which demonstrates great potential for clinical application.

CSMD: a computational subtraction-based microbiome discovery pipeline for species-level characterization of clinical metagenomic samples.

MOTIVATION: Microbiome analyses of clinical samples with low microbial biomass are challenging because of the very small quantities of microbial DNA relative to the human host, ubiquitous contaminating DNA in sequencing experiments and the large and rapidly growing microbial reference databases. RESULTS: We present computational subtraction-based microbiome discovery (CSMD), a bioinformatics pipeline specifically developed to generate accurate species-level microbiome profiles for clinical samples with low microbial loads. CSMD applies strategies for the maximal elimination of host sequences with minimal loss of microbial signal and effectively detects microorganisms present in the sample with minimal false positives using a stepwise convergent solution. CSMD was benchmarked in a comparative evaluation with other classic tools on previously published well-characterized datasets. It showed higher sensitivity and specificity in host sequence removal and higher specificity in microbial identification, which led to more accurate abundance estimation. All these features are integrated into a free and easy-to-use tool. Additionally, CSMD applied to cell-free plasma DNA showed that microbial diversity within these samples is substantially broader than previously believed. AVAILABILITY AND IMPLEMENTATION: CSMD is freely available at https://github.com/liuyu8721/csmd. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The DNA Methylation Inhibitor Zebularine Controls CD4+ T Cell Mediated Intraocular Inflammation.

CD4+ T cell mediated uveitis is conventionally treated with systemic immunosuppressive agents, including corticosteroids and biologics targeting key inflammatory cytokines. However, their long-term utility is limited due to various side effects. Here, we investigated whether DNA methylation inhibitor zebularine can target CD4+ T cells and control intraocular inflammation. Our results showed that zebularine restrained the expression of inflammatory cytokines IFN-γ and IL-17 in both human and murine CD4+ T cells in vitro. Importantly, it also significantly alleviated intraocular inflammation and retinal tissue damage in the murine experimental autoimmune uveitis (EAU) model in vivo, suggesting that the DNA methylation inhibitor zebularine is a candidate new therapeutic agent for uveitis.

Inhibition of Experimental Age-Related Macular Degeneration by ZQMT in Mice.

BACKGROUND: Age-related macular degeneration (AMD) is a progressive and irreversible eye disease. The anti-vascular endothelial growth factor (VEGF) therapy has revolutionized the treatment of neovascular AMD. However, the expense for such treatment is quite high. METHODS: We used a traditional Chinese medicine ZQMT as an alternative therapeutic regimen for AMD. We employed two in vivo animal models mimicking dry and wet AMD respectively to assess the therapeutic efficacy of ZQMT on treating AMD-related retinopathy. AMD-related retinopathy in Crb1rd8 mice was evaluated from week 1 to 8 by fundus photography. Laser-induced choroidal neovascularization (CNV) was evaluated by fluorescein angiography and histopathology. RESULTS: ZQMT increased CX3CR1 expression in murine CD4+ T cells either cultured in vitro or directly isolated from animals treated with ZQMT. We also performed both in vitro and in vivo studies to confirm that ZQMT has no apparent toxic effects. ZQMT alleviated AMD-related retinopathy in both Crb1rd8 and CNV models. Depletion of CCL2 and CX3CR1 in Crb1rd8 mice abolished the efficacy of ZQMT, suggesting that CCL2 and/or CX3CR1 may underlie the mechanisms of ZQMT in treating AMD-related retinopathy in mice. CONCLUSION: In summary, our study supports the protective roles of a traditional Chinese medicine ZQMT in AMD.

Structure of the human PKD1-PKD2 complex.

Mutations in two genes, PKD1 and PKD2, account for most cases of autosomal dominant polycystic kidney disease, one of the most common monogenetic disorders. Here we report the 3.6-angstrom cryo-electron microscopy structure of truncated human PKD1-PKD2 complex assembled in a 1:3 ratio. PKD1 contains a voltage-gated ion channel (VGIC) fold that interacts with PKD2 to form the domain-swapped, yet noncanonical, transient receptor potential (TRP) channel architecture. The S6 helix in PKD1 is broken in the middle, with the extracellular half, S6a, resembling pore helix 1 in a typical TRP channel. Three positively charged, cavity-facing residues on S6b may block cation permeation. In addition to the VGIC, a five-transmembrane helix domain and a cytosolic PLAT domain were resolved in PKD1. The PKD1-PKD2 complex structure establishes a framework for dissecting the function and disease mechanisms of the PKD proteins.

Cryo-EM structure of the polycystic kidney disease-like channel PKD2L1.

PKD2L1, also termed TRPP3 from the TRPP subfamily (polycystic TRP channels), is involved in the sour sensation and other pH-dependent processes. PKD2L1 is believed to be a nonselective cation channel that can be regulated by voltage, protons, and calcium. Despite its considerable importance, the molecular mechanisms underlying PKD2L1 regulations are largely unknown. Here, we determine the PKD2L1 atomic structure at 3.38 Å resolution by cryo-electron microscopy, whereby side chains of nearly all residues are assigned. Unlike its ortholog PKD2, the pore helix (PH) and transmembrane segment 6 (S6) of PKD2L1, which are involved in upper and lower-gate opening, adopt an open conformation. Structural comparisons of PKD2L1 with a PKD2-based homologous model indicate that the pore domain dilation is coupled to conformational changes of voltage-sensing domains (VSDs) via a series of π-π interactions, suggesting a potential PKD2L1 gating mechanism.

Epigenetics, microbiota, and intraocular inflammation: New paradigms of immune regulation in the eye.

Sight threatening immune responses that damage the eye characterize intraocular inflammatory diseases. These diseases including uveitis and age-related macular degeneration are worryingly common and quality of life shattering. Genetic studies in past decades significantly advanced our understanding of the etiology of these devastating diseases. Unfortunately, patient genetics alone failed to adequately explain disease origin, susceptibility, and progression. Non-genetic factors such as the epigenetic regulation of ocular diseases and the environmental factors triggering intraocular inflammation offer new insight into intraocular inflammatory disorders. Importantly, mounting evidence is signaling that dysbiosis of human microbiota leads to rapid epigenomic reprograming of host cells and results in the onset of many diseases. In this review, we discuss how epigenetic mechanisms and microbiota may cooperate to initiate and perpetuate ocular inflammation. Lastly, we propose that the discovery of intraocular microbiota presents a significant shift in thought affecting current approaches to the diagnosis, treatment, and prevention of intraocular inflammatory diseases such as uveitis and age-related macular degeneration. The geographical and genetic background difference in both disease presentation and genetic association of intraocular inflammatory diseases may be due to the variation of intraocular microbiota.