Membrane metalloprotease TRABD2A restricts HIV-1 progeny production in resting CD4+ T cells by degrading viral Gag polyprotein.

Resting CD4+ T cells are highly resistant to the production of human immunodeficiency virus type 1 (HIV-1). However, the mechanism by which resting CD4+ T cells restrict such production in the late viral replication phase of infection has remained unclear. In this study, we found that the cell membrane metalloprotease TRAB domain-containing protein 2A (TRABD2A) inhibited this production in resting CD4+ T cells by degrading the virion structural precursor polyprotein Gag at the plasma membrane. Depletion or inhibition of metalloprotease activity by TRABD2A profoundly enhanced HIV-1 production in resting CD4+ T cells. TRABD2A expression was much higher in resting CD4+ T cells than in activated CD4+ T cells and was considerably reduced by T cell activation. Moreover, reexpressing TRABD2A reinforced the resistance of activated CD4+ T cells to the production of HIV-1 progeny. Collectively, these results elucidate the molecular mechanism employed by resting CD4+ T cells to potently restrict the assembly and production of HIV-1 progeny.

Direct Laser Writing Photonic Crystal Hydrogels with a Supramolecular Sacrificial Scaffold.

Photonic crystal hydrogels (PCHs), with smart stimulus-responsive abilities, have been widely exploited as colorimetric sensors for years. However, the current fabrication technologies are mostly applicable to produce PCHs with simple geometries at the sub-millimeter scale, limiting the introduction of structural design into PCH sensors as well as the accompanied advanced applications. This paper reports the microfabrication of three-dimensional (3D) PCHs with the help of supramolecular agarose PCH as a sacrificial scaffold by two-photon lithography (TPL). The supramolecular PCHs, formulated with SiO2 colloidal nanoparticles and agarose aqueous solutions, show bright structural color and are degradable upon short-time dimethyl sulfoxide treatment. Leveraging the supramolecular PCH as a sacrificial scaffold, PCHs with precise 3D geometries can be fabricated in an economical and efficient way. This work demonstrates the application of such a strategy in the creation of structural-designed PCH mechanical microsensors that have not been explored before.

MRI Assessment of Diastolic Dysfunction in People Living With the Human Immunodeficiency Virus: Correlation With Markers of Disease Activity.

BACKGROUND: Despite the advent of combination antiretroviral therapy, people living with human immunodeficiency virus (PLWH) are at an increased risk for cardiac disease. PURPOSE: To explore the presence and extent of diastolic atrial and left ventricular dysfunction in PLWH using cardiac MRI in correlation with clinical markers of disease activity. STUDY TYPE: Prospective. POPULATION: A total of 163 participants comprising 101 HIV-infected individuals (age: 52 years [42-62 years]; 92% male) and 62 age- and sex-matched healthy volunteers (age: 51 years [30-72 years]; 85% male). FIELD STRENGTH/SEQUENCE: 3.0 T, cardiac MRI including balanced steady-state free precession (SSFP) for the short-axis, two-, three-, and four-chamber views were performed. ASSESSMENT: Assessment of cardiac function and strain analysis were accomplished by CVI42 software. Blood samples for CD4+ T cells and cardiac risk factors were also collected before MRI. STATISTICAL TESTS: Independent t tests, Mann-Whitney U test, Pearson's correlation analysis, and multivariate linear analyses (significance level: P < 0.05). RESULTS: PLWH had a significantly larger left atrial volume maximum index (LAVImax: 32.6 ± 8.7 vs. 28.7 ± 8.1 mL/m2), minimum (LAVImin: 14.8 ± 5.5 vs. 11.5 ± 5.4 mL/m2,), and prior to atrial contraction (LAVIpre-a: 23.4 ± 6.7 vs. 19.7 ± 7.2 mL/m2) as compared to healthy volunteers. The LA reservoir (LAtEF: 55.0 ± 10.2 vs. 61.4 ± 10.4; Sls: 29.0 ± 8.1 vs. 33.8 ± 11.8), conduit (LApEF: 28.4 ± 8.2 vs. 32.3 ± 11.3, P = 0.01; Sle: 16.3 ± 6.5 vs. 18.9 ± 8.2), and booster pump function (LAaEF: 37.4 ± 12.4 vs. 42.7 ± 13.1, P = 0.01, Sla: 12.7 ± 5.1 vs. 14.9 ± 5.7) were all significant impaired in PLWH. Global circumferential left ventricular diastolic strain rate (LVGCS-d) was significantly lower in the HIV patients. Multivariate analysis results showed that Nadir CD4+ T cells had a significant adverse association with LVGCS-d (ß = 0.51). CONCLUSION: LA structure abnormalities and LV diastolic dysfunction were manifested in PLWH, with Nadir CD4+ T cell counts potentially serving as a risk factor for early cardiac diastolic dysfunction. LEVEL OF EVIDENCE: 2 TECHNICAL EFFICACY: Stage 3.

Network Pharmacology-Based Identification of Key Pharmacological Mechanism of Shen-qi-di-huang Decoction Acting on Uremia.

This study employs network pharmacology to uncover the pharmacological mechanisms underlying Shen-qi-di-huang decoction's efficacy in treating uremia. We identified a total of 927 differentially expressed genes (DEGs) through differential expression analysis and the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database and analysis platform, of which 607 were downregulated and 320 were upregulated. We also obtained the effective biological components and related target gene information of Chinese herbal medicines such as Renshen, Huangqi, shudihuang, Shanyao, Fuling, Mudanpi, and Shanzhuyu in Shen-qi-di-huang decoction and constructed a regulatory relationship network between molecular components and target genes in Shen-qi-di-huang decoction. We then constructed a protein-protein interaction (PPI) network of 15 targeted genes (RXRA, ND6, CYP1B1, SLPI, CDKN1A, RB1, HIF1A, MYC, HSPB1, IFNGR1, NQO1, IRF1, RASA1, PSMG1 and MAP2K4) using the STRING database and visualized the PPI network using the software Cytoscape. In addition, we revealed the key molecular functions of uremia through Gene Ontology (GO) enrichment analysis, mainly including neuron apoptotic process, cellular response to oxidative stress, regulation of neuron apoptotic process, neuron projection cytoplasm, RNA polymerase II transcription regulator complex, plasma membrane bounded cell projection cytoplasm, NADH and NADPH dehydrogenase (quinone) activity, protein kinase inhibitor and ubiquitin protein ligase binding, etc. Finally, we identified important biological pathways in uremia through Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, which mainly concentrated in Kaposi sarcoma-associated, small cell lung cancer, Gastric cancer, Hepatitis B and C, Hepatocellular carcinoma, Thyroid cancer, Bladder cancer, MAPK signaling pathway, ErbB signaling pathway, Th17 cell differentiation, HIF-1 signaling pathway, Thyroid hormone signaling pathway and Cell cycle, etc. Using integrated bioinformatical analysis, we elucidated key pharmacological mechanisms based on targeted genes, which was enable early identification of patients with uremia and would contribute to early clinical diagnosis and treatment of patients.

Inhibitory receptor CD47 binding to plasma TSP1 suppresses NK-cell IFN-γ production via activating the JAK/STAT3 pathway during HIV infection.

BACKGROUND: Natural killer (NK) cells play an important first-line role against tumour and viral infections and are regulated by inhibitory receptor expression. Among these inhibitory receptors, the expression, function, and mechanism of cluster of differentiation 47 (CD47) on NK cells during human immunodeficiency virus (HIV) infection remain unclear. METHODS: Fresh peripheral blood mononuclear cells (PBMCs) were collected from people living with HIV (PLWH) and HIV negative controls (NC) subjects. Soluble ligand expression levels of CD47 were measured using ELISA. HIV viral proteins or Toll-like receptor 7/8 (TLR7/8) agonist was used to investigate the mechanisms underlying the upregulation of CD47 expression. The effect of CD47 on NK cell activation, proliferation, and function were evaluated by flow cytometry. RNA-seq was used to identify downstream pathways for CD47 and its ligand interactions. A small molecule inhibitor was used to restore the inhibition of NK cell function by CD47 signalling. RESULTS: CD47 expression was highly upregulated on the NK cells from PLWH, which could be due to activation of the Toll-like receptor 7/8 (TLR7/8) pathway. Compared with NC subjects, PLWH subjects exhibited elevated levels of CD47 ligands, thrombospondin-1 (TSP1), and counter ligand signal regulatory protein-α (SIRPα). The TSP1-CD47 axis drives the suppression of interferon gamma (IFN-γ) production and the activation of the Janus kinase signal transducer and activator of transcription (JAK-STAT) pathway in NK cells. After treatment with a STAT3 inhibitor, the NK cells from PLWH showed significantly improved IFN-γ production. CONCLUSIONS: The current data indicate that the binding of the inhibitory receptor CD47 to plasma TSP1 suppresses NK cell IFN-γ production by activating the JAK/STAT3 pathway during HIV infection. Our results suggest that CD47 and its related signalling pathways could be targets for improving NK cell function in people living with HIV.

First complete-genome documentation of HIV-1 intersubtype superinfection with transmissions of diverse recombinants over time to five recipients.

Human immunodeficiency virus type 1 (HIV-1) recombinants in the world are believed to be generated through recombination between distinct HIV-1 strains among coinfection or superinfection cases. However, direct evidence to support transmission of HIV-1 recombinants from a coinfected/superinfected donor to putative recipient is lacking. Here, we report on the origin and evolutionary relationship between a set of recombinants from a CRF01_AE/CRF07_BC superinfected putative donor and diverse CRF01_AE/CRF07_BC recombinants from five putative recipients. Interviews on sociodemographic characteristics and sexual behaviors for these six HIV-1-infected men who have sex with men showed that they had similar ways of partner seeking: online dating sites and social circles. Phylogenetic and recombination analyses demonstrated that the near-full-length genome sequences from six patients formed a monophyletic cluster different from known HIV-1 genotypes in maximum likelihood phylogenetic trees, were all composed of CRF01_AE and CRF07_BC fragments with two common breakpoints on env, and shared 4-7 breakpoints with each other. Moreover, 3' half-genomes of recombinant strains from five recipients had identical/similar recombinant structures with strains at longitudinal samples from the superinfected donor. Recombinants from the donor were paraphyletic, whereas five recipients were monophyletic or polyphyletic in the maximum clade credibility tree. Bayesian analyses confirmed that the estimated time to the most recent common ancestor (tMRCA) of CRF01_AE and CRF07_BC strains of the donor was 2009.2 and 2010.7, respectively, and all were earlier than the emergence of recombinants from five recipients. Our results demonstrated that the closely related unique recombinant forms of HIV-1 might be the descendent of a series of recombinants generated gradually in a superinfected patient. This finding highlights the importance of early initiation of antiretroviral therapy as well as tracing and testing of partners in patients with multiple HIV-1 infection.

Cardiac MRI-Based Assessment of Myocardial Injury in Asymptomatic People Living With Human Immunodeficiency Virus: Correlation With nadir CD4 Count.

BACKGROUND: There are known cardiac manifestations of HIV, but the findings in asymptomatic subjects are still not fully explored. PURPOSE: To evaluate for the presence of subclinical myocardial injury in asymptomatic people living with human immunodeficiency virus (PLWH) by cardiac MRI and to explore the possible association between subclinical myocardial injury and HIV-related clinical characteristics. STUDY TYPE: Cross-sectional. POPULATION: A total of 80 asymptomatic PLWH (age: 53 years [47-56 years]; 90% male) and 50 age- and sex-matched healthy participants. FIELD STRENGTH/SEQUENCE: A 3-T, cine sequence, T1, T2, and T2* mapping. ASSESSMENT: Function analysis was derived from short axis, two-, three-, and four-chamber cine images by feature tracking. Regions of interest were manually selected in the midventricular septum T1, T2, and T2* mapping sequences. PLWH were evaluated for T1 increment (△T1 mapping = native T1 - cutoff values) and HIV-related clinical characteristics, particularly the nadir CD4 count. And PLWH were stratified into two groups according to the cutoff value of native T1: elevated native T1 and normal. STATISTICAL TESTS: T test, Wilcoxon rank-sum test, Chi-square test, Spearman rank correlation, and logistic regression. P <0.05 indicated statistical significance. RESULTS: Asymptomatic PLWH revealed significantly higher native myocardial T1 values (1241 ± 29 msec vs. 1189 ± 21 msec), T2 values (40.7 ± 1.5 msec vs. 37.9 ± 1.4 msec), and lower LVGRS (30.2% ± 6.2% vs. 35.8% ± 6.4%), LVGCS (-18.0% ± 2.5% vs. -19.5% ± 2.0%), and LVGLS (-16.0% ± 3.8% vs. -17.9% ± 2.6%) but showed no difference in T2* values (17.3 msec [16.3-19.1 msec] vs. 18.3 msec [16.5-19.3 msec], P = 0.201). A negative correlation between the native T1 increment in PLWH with subclinical myocardial injury and the nadir CD4 count (u = -0.316). Nadir CD4 count <500 cells/mm3 was associated with higher odds of elevated native T1 myocardial values (odds ratio, 6.12 [95% CI, 1.07-34.91]) in PLWH. DATA CONCLUSION: Subclinical myocardial inflammation and dysfunction were present in asymptomatic PLWH, and a lower nadir CD4 count may be a risk factor for subclinical myocardial injury. EVIDENCE LEVEL: 1. TECHNICAL EFFICACY: Stage 2.

Molecular Network Analysis Discloses the Limited Contribution to HIV Transmission for Patients with Late HIV Diagnosis in Northeast China.

In the "treat all" era, the high rate of late HIV diagnosis (LHD) worldwide remains an impediment to ending the HIV epidemic. In this study, we analyzed LHD in newly diagnosed people living with HIV (PLWH) and its impact on HIV transmission in Northeast China. Sociodemographic information, baseline clinical data, and plasma samples obtained from all newly diagnosed PLWH in Shenyang, the largest city in Northeast China, between 2016 and 2019 were evaluated. Multivariate logistic regression analysis was performed to identify risk factors associated with LHD. A molecular network based on the HIV pol gene was constructed to assess the risk of HIV transmission with LHD. A total of 2882 PLWH, including 882 (30.6%) patients with LHD and 1390 (48.2%) patients with non-LHD, were enrolled. The risk factors for LHD were older age (≥ 30 years: p < .01) and diagnosis in the general population through physical examination (p < .0001). Moreover, the molecular network analysis revealed that the clustering rate (p < .0001), the fraction of individuals with ≥ 4 links (p = .0847), and the fraction of individuals linked to recent HIV infection (p < .0001) for LHD were significantly or marginally significantly lower than those recorded for non-LHD. Our study indicates the major risk factors associated with LHD in Shenyang and their limited contribution to HIV transmission, revealing that the peak of HIV transmission of LHD at diagnosis may have been missed. Early detection, diagnosis, and timely intervention for LHD may prevent HIV transmission.

A longitudinal analysis of immune escapes from HLA-B*13-restricted T-cell responses at early stage of CRF01_AE subtype HIV-1 infection and implications for vaccine design.

BACKGROUND: Identifying immunogens which can elicit effective T cell responses against human immunodeficiency virus type 1 (HIV-1) is important for developing a T-cell based vaccine. It has been reported that human leukocyte antigen (HLA)-B*13-restricted T-cell responses contributed to HIV control in subtype B' and C infected individuals. However, the kinetics of B*13-restricted T-cell responses, viral evolution within epitopes, and the impact on disease progression in CRF01_AE subtype HIV-1-infected men who have sex with men (MSM) are not known. RESULTS: Interferon-γ ELISPOT assays and deep sequencing of viral RNAs were done in 14 early HLA-B*13-positive CRF01_AE subtype HIV-1-infected MSM. We found that responses to RQEILDLWV (Nef106-114, RV9), GQMREPRGSDI (Gag226-236, GI11), GQDQWTYQI (Pol487-498, GI9), and VQNAQGQMV (Gag135-143, VV9) were dominant. A higher relative magnitude of Gag-specific T-cell responses, contributed to viral control, whereas Nef-specific T-cell responses were associated with rapid disease progression. GI11 (Gag) was conserved and strong GI11 (Gag)-specific T-cell responses showed cross-reactivity with a dominant variant, M228I, found in 3/12 patients; GI11 (Gag)-specific T-cell responses were positively associated with CD4 T-cell counts (R = 0.716, P = 0.046). Interestingly, the GI9 (Pol) epitope was also conserved, but GI9 (Pol)-specific T-cell responses did not influence disease progression (P > 0.05), while a D490G variant identified in one patient did not affect CD4 T-cell counts. All the other epitopes studied [VV9 (Gag), RQYDQILIEI (Pol113-122, RI10), HQSLSPRTL (Gag144-152, HL9), and RQANFLGRL (Gag429-437, RL9)] developed escape mutations within 1 year of infection, which may have contributed to overall disease progression. Intriguingly, we found early RV9 (Nef)-specific T-cell responses were associated with rapid disease progression, likely due to escape mutations. CONCLUSIONS: Our study strongly suggested the inclusion of GI11 (Gag) and exclusion of RV9 (Nef) for T-cell-based vaccine design for B*13-positive CRF01_AE subtype HIV-1-infected MSM and high-risk individuals.

Outcomes of persistent low-level viremia among HIV patients on antiretroviral therapy: A prospective cohort study.

INTRODUCTION: We aimed to investigate the relationship between low-level viremia (LLV) and virological failure (VF), death, and non-AIDS events (NAEs). METHODS: A prospective cohort study of people living with HIV (PLHIV) on antiretroviral therapy (ART) was conducted from 2011-2018 at an HIV clinic in Shenyang, China. The incidence of VF and the mortality and NAEs due to LLV were assessed. Cox proportional hazards regression was performed to investigate risk factors for VF, mortality, and NAEs. RESULTS: In total, 1288 patients, contributing 3915 person-years of follow-up (median follow-up, 2.5 years [interquartile range: 2-4 years]), were enrolled. Thirty-one patients (2.4%) experienced VF, 5 (0.4%) died, and 38 (3.0%) experienced NAEs. The risk of VF was significantly increased among patients with a viral load (VL) of 200-499 copies/mL (adjusted hazard ratio [aHR]: 14.92, 95% confidence interval [CI]: 5.92-37.60) or 500-999 copies/mL (aHR: 13.68, 95% CI: 3.61-51.87), but not among patients with a VL of 50-199 copies/mL (aHR: 3.10, 95% CI: 0.86-11.09). The risk of NAEs was significantly increased among patients with LLV (aHR: 7.33, 95% CI: 3.73-14.42). Compared to no LLV, a VL of 50-199 copies/mL (aHR: 4.11, 95% CI: 1.73-9.74), 200-499 copies/mL (aHR: 18.31, 95% CI: 6.66-50.33), and 500-999 copies/mL (aHR: 21.34, 95% CI: 5.69-80.01) showed higher risk of NAEs. CONCLUSION: Low-level viremia was associated with VF and NAEs. Patients with LLV, especially those with a VL ≥200 copies/mL, may need more frequent VL testing and NAE screening.

Rapid HIV Progression Is Associated with Extensive Ongoing Somatic Hypermutation.

The Ab response to HIV is of great interest, particularly in the context of a protective vaccine and broadly neutralizing Abs, but research is typically geared toward elite controllers because of their ability to successfully control the virus. In this study, we studied the evolution of the Ab repertoire over the first year of HIV infection in people classified as rapid progressors (RP) compared with typical progressors. HIV RPs are an important yet understudied group of HIV patients classified by a rapid decline in CD4 counts and accelerated development of AIDS. We found that the global IgG somatic hypermutation load negatively correlated with disease progression, possibly because of exaggerated isotype switching of unmutated sequences in patients with low CD4 counts. We measured Ab sequence evolution over time using longitudinal samples taken during the early stages of infection and 1 year postinfection. Within clonal lineages spanning both timepoints, visit 2-derived sequences harbored considerably more mutations than their visit 1 relatives. Despite extensive ongoing somatic hypermutation, the initially strong signs of Ag selection pressure observed in visit 1-derived sequences decayed by visit 2. These data suggest that excessive immune activation in RPs leads to a hyperactive B cell response that fails to confer protection.

Evaluation of the learning curve for conformal sphincter preservation operation in the treatment of ultralow rectal cancer.

BACKGROUND: To investigate the learning curve of conformal sphincter preservation operation (CSPO) in the treatment of ultralow rectal cancer and to further explore the influencing factors of operation time. METHODS: From August 2011 to April 2020, 108 consecutive patients with ultralow rectal cancer underwent CSPO by the same surgeon in the Department of Colorectal Surgery of Changhai Hospital. The moving average and cumulative sum control chart (CUSUM) curve were used to analyze the learning curve. The preoperative clinical baseline data, postoperative pathological data, postoperative complications, and survival data were compared before and after the completion of learning curve. The influencing factors of CSPO operation time were analyzed by univariate and multivariate analysis. RESULTS: According to the results of moving average and CUSUM method, CSPO learning curve was divided into learning period (1-45 cases) and learning completion period (46-108 cases). There was no significant difference in preoperative clinical baseline data, postoperative pathological data, postoperative complications, and survival data between the two stages. Compared with the learning period, the operation time (P < 0.05), blood loss (P < 0.05), postoperative flatus and defecation time (P < 0.05), liquid diet time (P < 0.05), and postoperative hospital stay (P < 0.05) in the learning completion period were significantly reduced, and the difference was statistically significant. Univariate and multivariate analysis showed that distance of tumor from anal verge (≥ 4cm vs. < 4cm, P = 0.039) and T stage (T3 vs. T1-2, P = 0.022) was independent risk factors for prolonging the operation time of CSPO. CONCLUSIONS: For surgeons with laparoscopic surgery experience, about 45 cases of CSPO are needed to cross the learning curve. At the initial stage of CSPO, beginners are recommended to select patients with ultralow rectal cancer whose distance of tumor from anal verge is less than 4 cm and tumor stage is less than T3 for practice, which can enable beginners to reduce the operation time, accumulate experience, build self-confidence, and shorten the learning curve on the premise of safety.

[Co-amorphous technology to improve dissolution and physical stability of silybin].

The present study explored the effect of co-amorphous technology in improving the dissolution rate and stability of silybin based on the puerarin-silybin co-amorphous system prepared by the spray-drying method. Solid-state characterization was carried out by powder X-ray diffraction(PXRD), polarizing microscopy(PLM), Fourier transform infrared spectroscopy(FT-IR), differential scanning calorimetry(DSC), etc. Saturated powder dissolution, intrinsic dissolution rate, moisture absorption, and stability were further investigated. The results showed that puerarin and silybin formed a co-amorphous system at a single glass transition temperature which was higher than that of any crude drug. The intrinsic dissolution rate and supersaturated powder dissolution of silybin in the co-amorphous system were higher than those of the crude drug and amorphous system. The co-amorphous system kept stable for as long as three months under the condition of 40 ℃, 75% relative humidity, which was longer than that of the single amorphous silybin. Therefore, the co-amorphous technology could significantly improve the dissolution and stability of silybin.

The inhibitory receptor Tim-3 fails to suppress IFN-γ production via the NFAT pathway in NK-cell, unlike that in CD4+ T cells.

BACKGROUND: T cell immunoglobulin and mucin domain-containing-3 (Tim-3) is a negative regulator expressed on T cells, and is also expressed on natural killer (NK) cells. The function of Tim-3 chiefly restricts IFNγ-production in T cells, however, the impact of Tim-3 on NK cell function has not been clearly elucidated. RESULTS: In this study, we demonstrated down-regulation of Tim-3 expression on NK cells while Tim-3 is upregulated on CD4+ T cells during HIV infection. Functional assays indicated that Tim-3 mediates suppression of CD107a degranulation in NK cells and CD4+ T cells, while it fails to inhibit the production of IFN-γ by NK cells. Analyses of downstream pathways using an antibody to block Tim-3 function demonstrated that Tim-3 can inhibit ERK and NFκB p65 signaling; however, it failed to suppress the NFAT pathway. Further, we found that the NFAT activity in NK cells was much higher than that in CD4+ T cells, indicating that NFAT pathway is important for promotion of IFN-γ production by NK cells. CONCLUSIONS: Thus, our data show that the expression of Tim-3 on NK cells is insufficient to inhibit IFN-γ production. Collectively, our findings demonstrate a potential mechanism of Tim-3 regulation of NK cells and a target for HIV infection immunotherapy.

Facile Surface Functionalization Strategy for Two-Photon Lithography Microstructures.

Two-photon lithography (TPL) is a powerful tool to construct small-scale objects with complex and precise 3D architectures. While the limited selection of chemical functionalities on the printed structures has restricted the application of this method in fabricating functional objects and devices, this study presents a facile, efficient, and extensively applicable method to functionalize the surfaces of the objects printed by TPL. TPL-printed objects, regardless of their compositions, can be efficiently functionalized by combining trichlorovinylsilane treatment and thiol-ene chemistry. Various functionalities can be introduced on the printed objects, without affecting their micro-nano topographies. Hence, microstructures with diverse functions can be generated using non-functional photoresists. Compared to existed strategies, this method is fast, highly efficient, and non photoresist-dependent. In addition, this method can be applied to various materials, such as metals, metal oxides, and plastics that can be potentially utilized in TPL or other 3D printing technologies. The applications of this method on the biofunctionalization of microrobots and cell scaffolds are also demonstrated.

Integrated analysis of lncRNA, miRNA and mRNA profiles reveals potential lncRNA functions during early HIV infection.

BACKGROUND: Long noncoding RNAs (lncRNAs) can regulate gene expression in a cis-regulatory fashion or as "microRNA sponges". However, the expression and functions of lncRNAs during early human immunodeficiency virus (HIV) infection (EHI) remain unclear. METHODS: 3 HAART-naive EHI patients and 3 healthy controls (HCs) were recruited in this study to perform RNA sequencing and microRNA (miRNA) sequencing. The expression profiles of lncRNAs, mRNAs and miRNAs were obtained, and the potential roles of lncRNAs were analysed based on discovering lncRNA cis-regulatory target mRNAs and constructing lncRNA-miRNA-mRNA competing endogenous RNA (ceRNA) networks. Then, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed on 175 lncRNA-associated differentially expressed (DE) mRNAs to investigate the potential functions of DE lncRNAs in ceRNA networks. RESULTS: A total of 242 lncRNAs, 1240 mRNAs and 21 mature known miRNAs were determined as differentially expressed genes in HAART-naive EHI patients compared to HCs. Among DE lncRNAs, 44 lncRNAs were predicted to overlap with 41 target mRNAs, and 107 lncRNAs might regulate their nearby DE mRNAs. Two DE lncRNAs might regulate their cis-regulatory target mRNAs BTLA and ZAP70, respectively, which were associated with immune activation. In addition, the ceRNA networks comprised 160 DE lncRNAs, 21 DE miRNAs and 175 DE mRNAs. Seventeen DE lncRNAs were predicted to regulate HIF1A and TCF7L2, which are involved in the process of HIV-1 replication. Twenty DE lncRNAs might share miRNA response elements (MREs) with FOS, FOSB and JUN, which are associated with both immune activation and HIV-1 replication. CONCLUSIONS: This study revealed that lncRNAs might play a critical role in HIV-1 replication and immune activation during EHI. These novel findings are helpful for understanding of the pathogenesis of HIV infection and provide new insights into antiviral therapy.

The accumulation of plasma acylcarnitines are associated with poor immune recovery in HIV-infected individuals.

BACKGROUND: Antiretroviral therapy (ART) can reduce opportunistic infections and mortality rates among individuals infected with human immunodeficiency virus (HIV); however, some HIV-infected individuals exhibit poor immune recovery after ART. Hence, we explored the association between metabolome profiles and immune recovery in HIV-infected individuals following ART. METHODS: An untargeted metabolomics approach was used to analyze plasma samples from 18 HIV-negative individuals and 20 HIV-infected individuals, including 10 immunological non-responders (INR, CD4+ T cell rise < 100 cells/µl) and 10 immunological responders (IR, CD4+ T cell rise > 300 cells/µl) after 2 years of ART. These individuals were followed for the next 6 years and viral loads and CD4+ T cell count were measured regularly. Orthogonal projection on latent structures discriminant analysis (OPLS-DA), ANOVA, correlation, receiver operating characteristic (ROC), and survival analyses were used for selection of discriminant metabolites. RESULTS: Eighteen lipid metabolites were identified which could distinguish among control, INR, and IR groups. Among them, myristoylcarnitine (MC), palmitoylcarnitine (PC), stearoylcarnitine (SC), and oleoylcarnitine (OC) were significantly elevated in INR plasma samples compared with those from the IR and control groups and were negatively associated with CD4+ T cell count. Additionally, ROC analysis using a combination of MC, PC, SC, and OC had high sensitivity and specificity for differentiating INR from IR (AUC = 0.94). Finally, survival analysis for the combination of MC, PC, SC, and OC demonstrated that it could predict CD4+ T cell count in patients undergoing long-term ART. CONCLUSIONS: High levels of lipid metabolites, MC, PC, SC, and OC are associated with poor immune recovery in patients receiving ART and these data provide potential new insights into immune recovery mechanisms.

Transmitted drug resistance to Tenofovir/Emtricitabine among persons with newly diagnosed HIV infection in Shenyang city, Northeast China from 2016 to 2018.

BACKGROUND: To assess transmitted drug resistance (TDR) to tenofovir (TDF)/emtricitabine (FTC), using as pre-exposure prophylaxis, among newly diagnosed human immunodeficiency virus-1 (HIV-1)-infected residents in Shenyang city, northeast China. METHODS: Demographic and epidemiological information of all newly diagnosed HIV-1 infected residents in Shenyang city from 2016 to 2018 were anonymously collected from the local HIV epidemic database. HIV-1 pol sequences were amplified from RNA in cryopreserved plasma samples and sequenced directly. Viral subtypes were inferred with phylogenetic analysis and drug resistance mutations (DRMs) were determined according to the Stanford HIVdb algorithm. Recent HIV infection was determined with HIV Limiting Antigen avidity electro immunoassay. RESULTS: A total of 2176 sequences (92.4%, 2176/2354) were obtained; 70.9% (1536/2167) were CRF01_AE, followed by CRF07_BC (18.0%, 391/2167), subtype B (4.7%, 102/2167), other subtypes (2.6%, 56/2167), and unique recombinant forms (3.8%, 82/2167). The prevalence of TDR was 4.9% (107/2167), among which, only 0.6% (13/2167) was resistance to TDF/FTC. Most of these subjects had CRF01_AE strains (76.9%, 10/13), were unmarried (76.9%, 10/13), infected through homosexual contact (92.3%, 12/13), and over 30 years old (median age: 33). The TDF/FTC DRMs included K65R (8/13), M184I/V (5/13), and Y115F (2/13). Recent HIV infection accounted for only 23.1% (3/13). Most cases were sporadic in the phylogenetic tree, except two CRF01_AE sequences with K65R (Bootstrap value: 99%). CONCLUSIONS: The prevalence of TDR to TDF/FTC is low among newly diagnosed HIV-infected cases in Shenyang, suggesting that TDR may have little impact on the protective effect of the ongoing CROPrEP project in Shenyang city.

Internet-Based HIV Self-Testing Among Men Who Have Sex With Men Through Pre-exposure Prophylaxis: 3-Month Prospective Cohort Analysis From China.

BACKGROUND: Routine HIV testing accompanied with pre-exposure prophylaxis (PrEP) requires innovative support in a real-world setting. OBJECTIVE: This study aimed to determine the usage of HIV self-testing (HIVST) kits and their secondary distribution to partners among men who have sex with men (MSM) in China, who use PrEP, in an observational study between 2018 and 2019. METHODS: In 4 major cities in China, we prospectively followed-up MSM from the China Real-world oral PrEP demonstration study, which provides daily or on-demand PrEP for 12 months, to assess the usage and secondary distribution of HIVST on quarterly follow-ups. Half of the PrEP users were randomized to receive 2 HIVSTs per month in addition to quarterly facility-based HIV testing. We evaluated the feasibility of providing HIVST to PrEP users. RESULTS: We recruited 939 MSM and randomized 471 to receive HIVST, among whom 235 (49.9%) were daily and 236 (50.1%) were on-demand PrEP users. At baseline, the median age was 29 years, 390 (82.0%) men had at least college-level education, and 119 (25.3%) had never undergone facility-based HIV testing before. Three months after PrEP initiation, 341 (74.5%) men had used the HIVST provided to them and found it very easy to use. Among them, 180 of 341 (52.8%) men had distributed the HIVST kits it to other MSM, and 132 (51.6%) among the 256 men who returned HIVST results reported that used it with their sexual partners at the onset of intercourse. Participants on daily PrEP were more likely to use HIVST (adjusted hazard ratio=1.3, 95% CI 1.0-1.6) and distribute HIVST kits (adjusted hazard ratio=1.3, 95% CI 1.1-1.7) than those using on-demand PrEP. CONCLUSIONS: MSM who used PrEP had a high rate of usage and secondary distribution of HIVST kits, especially among those on daily PrEP, which suggested high feasibility and necessity for HIVST after PrEP initiation. Assuming that fourth-generation HIVST kits are available, HIVST may be able to replace facility-based HIV testing to a certain extent. TRIAL REGISTRATION: Chinese Clinical Trial Registry ChiCTR1800020374; https://www.chictr.org.cn/showprojen.aspx?proj=32481. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): RR2-10.1136/bmjopen-2019-036231.

Determination of the Peptide AWRK6 in Rat Plasma by Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) and Its Application to Pharmacokinetics.

AWRK6 was a synthesized peptide developed based on the natural occurring peptide dybowskin-2CDYa, which was discovered in frog skin in our previous study. Here, a quantitative determination method for AWRK6 analysis in rat plasma by using liquid chromatography-tandem mass spectrometry (LC-MS/MS) was established and validated following U.S. FDA guidelines. A combination of plasma precipitation and liquid-liquid extraction was applied for the extraction. For pharmacokinetics study, the rats were administrated with AWRK6 via intraperitoneal and intravenous injection. The prepared plasma samples were separated on an ODS column and analyzed by tandem MS using precursor-to-product ion pairs of m/z: 533.4→84.2 for AWRK6 and m/z: 401.9→101.1 for internal standard Polymyxin B sulfate in multiple reaction monitoring mode. AWRK6 concentrations in rat plasma peaked at about 1.2 h after intraperitoneal injections at 2.35, 4.7 and 9.4 mg/kg bodyweight. The terminal half-life was around 2.8 h. The absolute bioavailability of AWRK6 was 50% after 3 doses via injection, and the apparent volume of distribution was 4.884 ± 1.736 L. The obtained determination method and pharmacokinetics profiles of AWRK6 provides a basis for further development, and forms a benchmark reference for peptide quantification.