Detecting Sources of Immune Activation and Viral Rebound in HIV Infection.

Anti-retroviral therapy (ART) generally suppresses HIV replication to undetectable levels in peripheral blood, but immune activation associated with increased morbidity and mortality is sustained during ART, and infection rebounds when treatment is interrupted. To identify drivers of immune activation and potential sources of viral rebound, we modified RNAscope in situ hybridization to visualize HIV-producing cells as a standard against which to compare the following assays of potential sources of immune activation and virus rebound following treatment interruption: (i) envelope detection by induced transcription-based sequencing (EDITS) assay; (ii) HIV-Flow; (iii) Flow-FISH assays that can scan tissues and cell suspensions to detect rare cells expressing env mRNA, gag mRNA/Gag protein and p24; and (iv) an ultrasensitive immunoassay that detects p24 in cell/tissue lysates at subfemtomolar levels. We show that the sensitivities of these assays are sufficient to detect one rare HIV-producing/env mRNA+/p24+ cell in one million uninfected cells. These high-throughput technologies provide contemporary tools to detect and characterize rare cells producing virus and viral antigens as potential sources of immune activation and viral rebound. IMPORTANCE Anti-retroviral therapy (ART) has greatly improved the quality and length of life for people living with HIV, but immune activation does not normalize during ART, and persistent immune activation has been linked to increased morbidity and mortality. We report a comparison of assays of two potential sources of immune activation during ART: rare cells producing HIV and the virus' major viral protein, p24, benchmarked on a cell model of active and latent infections and a method to visualize HIV-producing cells. We show that assays of HIV envelope mRNA (EDITS assay), gag mRNA, and p24 (Flow-FISH, HIV-Flow. and ultrasensitive p24 immunoassay) detect HIV-producing cells and p24 at sensitivities of one infected cell in a million uninfected cells, thereby providing validated tools to explore sources of immune activation during ART in the lymphoid and other tissue reservoirs.

Nucleotide-Tackified Organohydrogel Electrolyte for Environmentally Self-Adaptive Flexible Supercapacitor with Robust Electrolyte/Electrode Interface.

Hydrogel electrolytes have attracted enormous attention in flexible and safe supercapacitors. However, the interfacial contact problem between hydrogel electrolyte and electrodes, and the environmental instability are the key factors restricting the development of hydrogel-based supercapacitors. Here, a nucleotide-tackified adhesive organohydrogel electrolyte is successfully constructed and exhibits freezing resistance and water-holding ability based on the water/glycerol binary solvent system. Adenosine monophosphate enables the organohydrogels to possess outstanding adhesion and mechanical robustness. The robust adhesion can ensure close contact between the organohydrogel electrolyte and electrodes for constructing an all-in-one supercapacitor with low interfacial contact resistance. Impressively, the integrated organohydrogel-based supercapacitors display an areal specific capacitance of 163.6 mF cm-2 . Besides, the supercapacitors feature prominent environmental stability with capacitance retention of 90.6% after 5000 charging/discharging cycles at -20 °C. Furthermore, based on the strong interfacial adhesion, the supercapacitors present excellent electrochemical stability without delamination/displacement between electrolyte and electrodes even under severe deformations such as bending and twisting. It is anticipated that this work will provide an encouraging way for developing flexible energy storage devices with electrochemical stability and environmental adaptability.

Hierarchical colloidosomes self-assembled from block copolymer micelles via emulsion interfacial confinement.

Hierarchical self-assembly of polymeric building blocks into high-level colloidosomes is desirable to not only design novel nanostructures but also fabricate the complex artificial materials across many length scales with multifunctionality. Although great progress has been made in the designing the hierarchical colloidosomes, the fabrication of polymeric colloidosomes self-assembled from block copolymer (BCP) colloidal nanoparticles still remains challenge. Here, we report the fabrication of the hierarchical polymeric colloidosomes with typical hollow internal structures self-assembled from the polystyrene-block-poly (2-vinyl pyridine) (PS-b-P2VP) BCP spherical micelles through the emulsion interfacial confinement, which is constructed through the water-in-1-butanol emulsion system. Moreover, the hierarchical colloidosomes can disassemble into the original uniform spherical micelles under the acid aqueous solution, indicating that the colloidosomes possess good pH stimuli-responsibility. Finally, the stability of the colloidosomes can be greatly improved by cross-linking the P2VP corona of original spherical micelles, offering the effective templates for construction of the multifunctional materials. This finding provides a simple yet effective method for the fabrication of the hierarchical colloidosomes from the BCP building blocks.

Lead removal from water using organic acrylic amine fiber (AAF) and inorganic-organic P-AAF, fixed bed filtration and surface-induced precipitation.

Granular porous sorbents were normally used for heavy metals removal from water. To search for the new commercial sorbent and treatment strategy, an organic acrylic amine fiber (AAF) and phosphorus loading inorganic-organic AAF (P-AAF) were prepared and used for lead (Pb) removal from water. A new strategy of inorganic-organic coupling technology was proposed for Pb removal, based on the hypothesis of surface-induced precipitation mechanism. The AAF showed a Pb adsorption capacity of 417 mg/g from the Langmuir fitting, while the column filtration technology was further applied to measure the adsorption edge and applications. Effects of different initial Pb concentrations, hydraulic retention time, and co-existing P were considered in the filtration experiments. The presence of 0.8 mg/L P in water significantly improved the Pb breakthrough point from 15,000 to 41,000 bed volumes of water spiked with 85 µg/L Pb, while the P-AAF fixed bed showed better removal of Pb than AAF SEM/EDX and XRD spectra were employed for determining the surface functional groups and the formation of surface-induced precipitation of pyromorphite (Pb5(PO4)3OH) on AAF. This study verified the application of AAF sorbent for Pb removal and the enhanced effect of coating P on AAF, thus improved our fundamental understanding and application of the surface chemistry process of Pb with P.

Anti-inflammatory effects of doxepin hydrochloride against LPS-induced C6-glioma cell inflammatory reaction by PI3K-mediated Akt signaling.

Recent studies have shown that tricyclic antidepressants (TCAs) may have anti-inflammatory and anticonvulsant effects in addition to its antidepressant effects. So far, the nonantidepressant effects of TCAs and their molecular pharmacological mechanisms remain completely unclear. Chronic inflammation in the brain parenchyma may be related to the pathogenesis and progression of various neurodegenerative diseases. As a common antidepressant and anti-insomnia drug, doxepin also may be a potential anti-inflammatory and anticonvulsant drug, so the study on the anti-inflammatory protective effect of doxepin and its molecular mechanism has become a very important issue in pharmacology and clinical medicine. Further elucidating the anti-inflammatory and neuroprotective effects of doxepin and its molecular mechanism may provide the important theoretical and clinical basis for the prevention and treatment of neurodegenerative disease. This study was designed to understand the glio-protective mechanism of doxepin against the inflammatory damage induced by lipopolysaccharide (LPS) exposure in C6-glioma cells. We found the treatment of C6-glioma cells with LPS results in deleterious effects, including the augmentation of inflammatory cytokine levels (tumor necrosis factor-α, interleukin-1ß), and suppresses the Akt phosphorylation. Furthermore, our outcomes demonstrated that doxepin was able to suppress these effects induced by LPS, through activation of the phosphatidylinositol-3-kinase-mediated protein kinase B (Akt) pathway. To sum up, these results highlight the potential role of doxepin against neuroinflammatory-related disease in the brain.

Robust and anti-fatigue hydrophobic association hydrogels assisted by titanium dioxide for photocatalytic activity.

Currently, robust and functional hydrogels have attracted extensive attention due to their potential applications in wastewater treatment, farmland water conservation and other fields. Herein, a series of hydrophobic association hydrogels assisted by titanium dioxide (TiO2) was fabricated via one-pot in situ photo-induced polymerization. TiO2 nanoparticles could act as both photo-initiators and physical crosslinking points. The TiO2-assisted hydrophobic association hydrogels exhibited a high tensile strength of 306 kPa, superior compression strength of 2.17 MPa and excellent fatigue resistance. Simultaneously, the incorporation of TiO2 endowed the hydrogel with photocatalytic capacity for dye wastewater treatment based on the inherent nature of TiO2. The results indicated that the hydrogels contributed to the degradation of various ionic dyes including methylene blue, rhodamine B and bromophenol blue, and the removal of methylene blue achieved a rate of 96.63%. Significantly, the hydrogel could be repeatedly utilized and the removal rate showed no evident decrease after five cycles, indicating that the hydrogels could be powerful candidates as photocatalysts for dye wastewater treatment.

Multipurpose and Durable Adhesive Hydrogel Assisted by Adenine and Uracil from Ribonucleic Acid.

Nucleobase pairs of adenine and uracil (A-U) from ribonucleic acid are of particular interest for various promising material properties. Herein, a novel polyacrylamide hydrogel with adhesive properties that are assisted by adenine and uracil has been designed and investigated. The incorporation of adenine and uracil enables the formation of a polyacrylamide hydrogel with remarkable adhesive behaviour with various materials including polytetrafluoroethylene (PTFE), plastics, rubber, glasses, metal, ceramics and wood. Moreover, the adhesive hydrogel can easily and directly adhere to humid biological tissues without any extra process, including heart, liver, spleen, lung, kidney, bone and muscle of mouse. More impressively, even after repeated peeling tests (10×), the AU-mediated polyacrylamide hydrogel still exhibits excellent durable adhesion for various materials. From mechanical contact tests, the adhesion energy of the A-U adhesive hydrogel is 47.9 J m-2 , which is nearly nine times that of polyacrylamide hydrogel (5.3 J m-2 ). The 90° peeling strength for aluminium, titanium, silica rubbers, glasses, PTFE and hogskin is 518, 645, 445, 396, 349, and 119 N m-1 , respectively. The multipurpose and durable adhesive behaviour of hydrogels assisted by adenine and uracil indicated the promise of nucleobase pairs from ribonucleic acid for the future development of adhesive materials.

Vaccine-induced CD8+ T cells control AIDS virus replication.

Developing a vaccine for human immunodeficiency virus (HIV) may be aided by a complete understanding of those rare cases in which some HIV-infected individuals control replication of the virus. Most of these elite controllers express the histocompatibility alleles HLA-B*57 or HLA-B*27 (ref. 3). These alleles remain by far the most robust associations with low concentrations of plasma virus, yet the mechanism of control in these individuals is not entirely clear. Here we vaccinate Indian rhesus macaques that express Mamu-B*08, an animal model for HLA-B*27-mediated elite control, with three Mamu-B*08-restricted CD8(+) T-cell epitopes, and demonstrate that these vaccinated animals control replication of the highly pathogenic clonal simian immunodeficiency virus (SIV) mac239 virus. High frequencies of CD8(+) T cells against these Vif and Nef epitopes in the blood, lymph nodes and colon were associated with viral control. Moreover, the frequency of the CD8(+) T-cell response against the Nef RL10 epitope (Nef amino acids 137-146) correlated significantly with reduced acute phase viraemia. Finally, two of the eight vaccinees lost control of viral replication in the chronic phase, concomitant with escape in all three targeted epitopes, further implicating these three CD8(+) T-cell responses in the control of viral replication. Our findings indicate that narrowly targeted vaccine-induced virus-specific CD8(+) T-cell responses can control replication of the AIDS virus.

Rapidly recoverable, anti-fatigue, super-tough double-network hydrogels reinforced by macromolecular microspheres.

In this study, a novel strategy was designed to prepare rapidly recoverable, anti-fatigue, super-tough double-network hydrogels by introducing macromolecular microspheres (MMs) as cross-linking centers for hydrophobic associations. MMs were prepared via emulsion polymerization using butyl acrylate (BA) as a main component and dicyclopentyl acrylate (DCPA) as a cross-linker. Then, a double-network (DN) hydrogel was prepared using gelatin as the first network and a copolymer of acrylamide and hexadecyl methacrylate stabilized by MMs as the second network. As a result, the DN hydrogels that were toughened by MMs exhibited an excellent fracture strength of 1.48 MPa and a fracture strain of 2100%. Moreover, the hydrogels exhibited rapid recoverability and fatigue resistance. Therefore, the strategy would open up a novel avenue for the toughening of DN hydrogels for biomedical applications.

Super-tough, ultra-stretchable and strongly compressive hydrogels with core-shell latex particles inducing efficient aggregation of hydrophobic chains.

Toughness, strechability and compressibility for hydrogels were ordinarily balanced for their use as mechanically responsive materials. For example, macromolecular microsphere composite hydrogels with chemical crosslinking exhibited excellent compression strength and strechability, but poor tensile stress. Here, a novel strategy for the preparation of a super-tough, ultra-stretchable and strongly compressive hydrogel was proposed by introducing core-shell latex particles (LPs) as crosslinking centers for inducing efficient aggregation of hydrophobic chains. The core-shell LPs always maintained a spherical shape due to the presence of a hard core even by an external force and the soft shell could interact with hydrophobic chains due to hydrophobic interactions. As a result, the hydrogels reinforced by core-shell LPs exhibited not only a high tensile strength of 1.8 MPa and dramatic elongation of over 20 times, but also an excellent compressive performance of 13.5 MPa at a strain of 90%. The Mullins effect was verified for the validity of core-shell LP-reinforced hydrogels by inducing aggregation of hydrophobic chains. The novel strategy strives to provide a better avenue for designing and developing a new generation of hydrophobic association tough hydrogels with excellent mechanical properties.

Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody Gene Transfer Protects Nonhuman Primates from Mucosal Simian-Human Immunodeficiency Virus Infection.

Broadly neutralizing antibodies (bnAbs) can prevent lentiviral infection in nonhuman primates and may slow the spread of human immunodeficiency virus type 1 (HIV-1). Although protection by passive transfer of human bnAbs has been demonstrated in monkeys, durable expression is essential for its broader use in humans. Gene-based expression of bnAbs provides a potential solution to this problem, although immune responses to the viral vector or to the antibody may limit its durability and efficacy. Here, we delivered an adeno-associated viral vector encoding a simianized form of a CD4bs bnAb, VRC07, and evaluated its immunogenicity and protective efficacy. The expressed antibody circulated in macaques for 16 weeks at levels up to 66 g/ml, although immune suppression with cyclosporine (CsA) was needed to sustain expression. Gene-delivered simian VRC07 protected against simian-human immunodeficiency virus (SHIV) infection in monkeys 5.5 weeks after treatment. Gene transfer of an anti-HIV antibody can therefore protect against infection by viruses that cause AIDS in primates when the host immune responses are controlled.

Live simian immunodeficiency virus vaccine correlate of protection: immune complex-inhibitory Fc receptor interactions that reduce target cell availability.

Principles to guide design of an effective vaccine against HIV are greatly needed, particularly to protect women in the pandemic's epicenter in Africa. We have been seeking these principles by identifying correlates of the robust protection associated with SIVmac239Δnef vaccination in the SIV-rhesus macaque animal model of HIV-1 transmission to women. We identified one correlate of SIVmac239Δnef protection against vaginal challenge as a resident mucosal system for SIV-gp41 trimer Ab production and neonatal FcR-mediated concentration of these Abs on the path of virus entry to inhibit establishment of infected founder populations at the portal of entry. In this study, we identify blocking CD4(+) T cell recruitment to thereby inhibit local expansion of infected founder populations as a second correlate of protection. Virus-specific immune complex interactions with the inhibitory FcγRIIb receptor in the epithelium lining the cervix initiate expression of genes that block recruitment of target cells to fuel local expansion. Immune complex-FcγRIIb receptor interactions at mucosal frontlines to dampen the innate immune response to vaginal challenge could be a potentially general mechanism for the mucosal immune system to sense and modulate the response to a previously encountered pathogen. Designing vaccines to provide protection without eliciting these transmission-promoting innate responses could contribute to developing an effective HIV-1 vaccine.

NK cell responses to simian immunodeficiency virus vaginal exposure in naive and vaccinated rhesus macaques.

NK cell responses to HIV/SIV infection have been well studied in acute and chronic infected patients/monkeys, but little is known about NK cells during viral transmission, particularly in mucosal tissues. In this article, we report a systematic study of NK cell responses to high-dose vaginal exposure to SIVmac251 in the rhesus macaque female reproductive tract (FRT). Small numbers of NK cells were recruited into the FRT mucosa following vaginal inoculation. The influx of mucosal NK cells preceded local virus replication and peaked at 1 wk and, thus, was in an appropriate time frame to control an expanding population of infected cells at the portal of entry. However, NK cells were greatly outnumbered by recruited target cells that fuel local virus expansion and were spatially dissociated from SIV RNA+ cells at the major site of expansion of infected founder populations in the transition zone and adjoining endocervix. The number of NK cells in the FRT mucosa decreased rapidly in the second week, while the number of SIV RNA+ cells in the FRT reached its peak. Mucosal NK cells produced IFN-γ and MIP-1α/CCL3 but lacked several markers of activation and cytotoxicity, and this was correlated with inoculum-induced upregulation of the inhibitory ligand HLA-E and downregulation of the activating receptor CD122/IL-2Rß. Examination of SIVΔnef-vaccinated monkeys suggested that recruitment of NK cells to the genital mucosa was not involved in vaccine-induced protection from vaginal challenge. In summary, our results suggest that NK cells play, at most, a limited role in defenses in the FRT against vaginal challenge.

Live simian immunodeficiency virus vaccine correlate of protection: local antibody production and concentration on the path of virus entry.

We sought design principles for a vaccine to prevent HIV transmission to women by identifying correlates of protection conferred by a highly effective live attenuated SIV vaccine in the rhesus macaque animal model. We show that SIVmac239Δnef vaccination recruits plasma cells and induces ectopic lymphoid follicle formation beneath the mucosal epithelium in the rhesus macaque female reproductive tract. The plasma cells and ectopic follicles produce IgG Abs reactive with viral envelope glycoprotein gp41 trimers, and these Abs are concentrated on the path of virus entry by the neonatal FcR in cervical reserve epithelium and in vaginal epithelium. This local Ab production and delivery system correlated spatially and temporally with the maturation of local protection against high-dose pathogenic SIV vaginal challenge. Thus, designing vaccines to elicit production and concentration of Abs at mucosal frontlines could aid in the development of an effective vaccine to protect women against HIV-1.

Glycerol monolaurate prevents mucosal SIV transmission.

Although there has been great progress in treating human immunodeficiency virus 1 (HIV-1) infection, preventing transmission has thus far proven an elusive goal. Indeed, recent trials of a candidate vaccine and microbicide have been disappointing, both for want of efficacy and concerns about increased rates of transmission. Nonetheless, studies of vaginal transmission in the simian immunodeficiency virus (SIV)-rhesus macaque (Macacca mulatta) model point to opportunities at the earliest stages of infection in which a vaccine or microbicide might be protective, by limiting the expansion of infected founder populations at the portal of entry. Here we show in this SIV-macaque model, that an outside-in endocervical mucosal signalling system, involving MIP-3alpha (also known as CCL20), plasmacytoid dendritic cells and CCR5(+ )cell-attracting chemokines produced by these cells, in combination with the innate immune and inflammatory responses to infection in both cervix and vagina, recruits CD4(+) T cells to fuel this obligate expansion. We then show that glycerol monolaurate-a widely used antimicrobial compound with inhibitory activity against the production of MIP-3alpha and other proinflammatory cytokines-can inhibit mucosal signalling and the innate and inflammatory response to HIV-1 and SIV in vitro, and in vivo it can protect rhesus macaques from acute infection despite repeated intra-vaginal exposure to high doses of SIV. This new approach, plausibly linked to interfering with innate host responses that recruit the target cells necessary to establish systemic infection, opens a promising new avenue for the development of effective interventions to block HIV-1 mucosal transmission.

Predatory Bdellovibrio-and-like organism mixtures on efficient MBR in-situ membrane fouling diminishment and mechanisms.

Membrane fouling is a major hindrance that restricts the application of membrane bioreactors (MBRs). Bdellovibrio-and-like organisms (BALOs), as obligatory parasitic bacteria, prey upon various bacteria. In this study, the BALO mixtures were screened and found more effective in membrane fouling mitigation compared to the single BALO species and extended the membrane filtration period by as long as 33.3%. The higher BALO diversity reduced the potential foulants generation in the activated sludge by decreasing the sludge viscosity as high as 13.8 ± 0.6% than the pure culture of BALO. Meanwhile, the mixed BALOs demonstrated superior biofilm predation capabilities, with the content of soluble microbial products and extracellular polymeric substances on the biofilm decreasing by 26.1 ± 0.5% and 38.3 ± 0.2% as the most compared to the single BALO species involved system. Additionally, the BALO mixtures expanded the single strains' host lysis spectrum of both the activated sludge and biofilm. The abundance of membrane-fouling-related bacteria such as Flavobacterium, Rhodobacter, and Labilithrix and pioneer bacteria such as Sphingorhabdus and Pseudomonas was significantly reduced. In summary, this study disclosed the significantly better membrane fouling mitigation effects of the BALOs with higher diversity, suggesting that the expansion of the host range is crucial for the further application of BALOs to enhance the anti-fouling performance of the MBR system.

The immunosuppressive role of IL-32 in lymphatic tissue during HIV-1 infection.

One pathological hallmark of HIV-1 infection is chronic activation of the immune system, driven, in part, by increased expression of proinflammatory cytokines. The host attempts to counterbalance this prolonged immune activation through compensatory mediators of immune suppression. We recently identified a gene encoding the proinflammatory cytokine IL-32 in microarray studies of HIV-1 infection in lymphatic tissue (LT) and show in this study that increased expression of IL-32 in both gut and LT of HIV-1-infected individuals may have a heretofore unappreciated role as a mediator of immune suppression. We show that: 1) IL-32 expression is increased in CD4(+) T cells, B cells, macrophages, dendritic cells, and epithelial cells in vivo; 2) IL-32 induces the expression of immunosuppressive molecules IDO and Ig-like transcript 4 in immune cells in vitro; and 3) in vivo, IL-32-associated IDO/Ig-like transcript 4 expression in LT macrophages and gut epithelial cells decreases immune activation but also may impair host defenses, supporting productive viral replication, thereby accounting for the correlation between IL-32 levels and HIV-1 replication in LT. Thus, during HIV-1 infection, we propose that IL-32 moderates chronic immune activation to avert associated immunopathology but at the same time dampens the antiviral immune response and thus paradoxically supports HIV-1 replication and viral persistence.

Tough, recyclable and biocompatible carrageenan-modified polyvinyl alcohol ionic hydrogel with physical cross-linked for multimodal sensing.

Biocompatibility hydrogel conductors are considered as sustainable bio-electronic materials for the application of wearable sensors and implantable devices. However, they mostly face the limitations of mismatched mechanical properties with skin tissue and the difficulty of recycling. In this regard, here, a biocompatible, tough, reusable sensor based on physical crosslinked polyvinyl alcohol (PVA) ionic hydrogel modified with ι-carrageenan (ι-CG) helical network was reported. Through simulating the ion transport and network structure of biological systems, the ionic hydrogels with skin-like mechanical features exhibit large tensile strain of 640 %, robust fracture strength of 800 kPa, soft modulus and high fatigue resistance. Meanwhile, the ionic hydrogel-based sensors possess a high response to strain/pressure over a wide range and could be utilized for multimodal sensing of human activity signals. Benefit from biosafety and temperature reversibility of ι-CG and PVA endow hydrogels with not only biocompatibility, but also meaningfully recyclability. The as-prepared hydrogels could be freely reconstructed into new flexible electronics and safely integrated with the human skin. It could be anticipated that the physically cross-linked ionic hydrogel conductor could expand the options for next-generation bio-based sensors.

New insights into exogenous N-acyl-homoserine lactone manipulation in biological nitrogen removal system against ZnO nanoparticle shock.

The effects and mechanisms of three N-acyl-homoserine lactones (AHLs) (C4-HSL, C6-HSL, and C10-HSL) on responses of biological nitrogen removal (BNR) systems to zinc oxide nanoparticle (NP) shock were investigated. All three AHLs improved the NP-impaired ammonia oxidation rates by up to 50.0 % but inhibited the denitrification process via regulating nitrogen metabolism-related enzyme activities. C4-HSL accelerated the catalase activity by 13.2 %, while C6-HSL and C10-HSL promoted the superoxide dismutase activity by 26.6 % and 18.4 %, respectively, to reduce reactive oxygen species levels. Besides, the enhancements of tryptophan protein and humic acid levels in tightly-bound extracellular polymeric substance by AHLs were vital for NP toxicity attenuation. The metabonomic analysis demonstrated that all three AHLs up-regulated the levels of lipid- and antioxidation-related metabolites to advance the system's resistance to NP shock. The "dual character" of AHLs emphasized the concernment of legitimately employing AHLs to alleviate NP stress for BNR systems.

Improved export coefficient model for identification of watershed environmental risk areas.

As a complex system under the joint action of man and nature, land use/cover directly or indirectly affects the environmental quality of the freshwater ecosystem. Studying the response of water environment quality to land use/cover change was significant to accurately simulate lake water quality and effectively enhance the management level. As an empirical model, the classical export coefficient model has been widely used and developed in agricultural non-point source pollution research because of its simple structure and convenient application. However, it assumes that the export coefficient of a particular type of land use/cover was constant, ignoring the influence of surface runoff and interception on the output intensity of pollutants in pollutant migration. This study improved the classical export coefficient model by adding factors such as precipitation, surface cover, and topography, evaluated the contribution of land use/cover to total nitrogen load into the lake in Dianchi Lake Basin, and applied the pollution assessment results to the identification of watershed environmental risk areas. The results showed that the improved export coefficient model could better simulate the relationship between land use/cover and total nitrogen load into Dianchi Lake from the basin. At the same time, spatial characteristics of the total nitrogen load contribution of the terrestrial could be represented. The high-risk areas in the basin were mainly cultivated land and construction areas with low vegetation coverage around lakes or downstream. The contribution per unit area to the TN load into the lake from areas with a high risk was 14.28 t/km2, which was 3.47 times that of medium-high-risk areas and 52.28 times that of the medium-risk area. Land use control measures in high-risk areas in the basin should be further strengthened, especially in the lakeside zone.