Extracellular Membrane Proteins Research Articles

Development of a novel prostate Cancer-Stroma Sphere (CSS) model for In Vitro tumor microenvironment studies

Tumor employs non-cancerous cells to gain beneficial features that promote growth and survival of cancer cells. Despite intensive research in the area of tumor microenvironment, there is still a lack of reliable and reproducible in vitro model for tumor and tumor-microenvironment cell interaction studies. Herein we report the successful development of a heterogeneous cancer-stroma sphere (CSS) model composed of prostate adenocarcinoma PC3 cells and immortalized mesenchymal stem cells (MSC). The CSS model demonstrated a structured spatial layout of the cells, with stromal cells concentrated at the center of the spheres and tumor cells located on the periphery. Significant increase in the levels of VEGFA, IL-10, and IL1a has been detected in the conditioned media of CSS as compared to PC3 spheres. Single cell RNA sequencing data revealed that VEGFA was secreted by MSC cells within heterogeneous spheroids. Enhanced expression of extracellular membrane (ECM) proteins was also shown for CSS-derived MSCs. Furthermore, we demonstrated that the multicellular architecture altered cancer cell response to chemotherapeutic agents: the inhibition of sphere formation by topotecan was 74.92 ± 4.56 % for PC3 spheres and 45.95 ± 7.84 % for CSS spheres (p < 0.01), docetaxel showed 37,51± 20,88 % and 15,67± 14,08 % inhibition, respectively (p < 0.05). Thus, CSS present an effective in vitro model for examining the extracellular matrix composition and cell-to-cell interactions within the tumor, as well as for evaluating the antitumor activity of drugs.

A co-assembly platform engaging macrophage scavenger receptor A for lysosome-targeting protein degradation

Targeted degradation of proteins has emerged as a powerful method for modulating protein homeostasis. Identification of suitable degraders is essential for achieving effective protein degradation. Here, we present a non-covalent degrader construction strategy, based on a modular supramolecular co-assembly system consisting of two self-assembling peptide ligands that bind cell membrane receptors and the protein of interest simultaneously, resulting in targeted protein degradation. The developed lysosome-targeting co-assemblies (LYTACAs) can induce lysosomal degradation of extracellular protein IL-17A and membrane protein PD-L1 in several scavenger receptor A-expressing cell lines. The IL-17A-degrading co-assembly has been applied in an imiquimod-induced psoriasis mouse model, where it decreases IL-17A levels in the skin lesion and alleviates psoriasis-like inflammation. Extending to asialoglycoprotein receptor-related protein degradation, LYTACAs have demonstrated the versatility and potential in streamlining degraders for extracellular and membrane proteins.

Identification of common fungal extracellular membrane (CFEM) proteins in Fusarium sacchari that inhibit plant immunity and contribute to virulence

ABSTRACT Fusarium sacchari is one of the primary pathogens causing pokkah boeng disease in sugarcane in China. Common fungal extracellular membrane (CFEM) domain-containing proteins only exist in fungi and play a significant role in the interaction of pathogens and plants. However, the CFEM proteins in F. sacchari were not identified and functionally resolved. Of the 20 FsCFEM proteins identified from the genome of F. sacchari , 16 FsCFEM genes were cloned, while 4 FsCFEM proteins (Fs06761, Fs08184, Fs10706, and Fs13617) were identified as effector proteins that could suppress the programmed cell death triggered by BCL2-associated X protein in Nicotiana benthamiana . Subcellular localization analysis revealed that these four effectors could enter plant cells and are mainly located in the cell membrane and nucleus. By the yeast YTK12 secretion system, the signal peptides of four CFEM proteins were functionally verified. The expression levels of these four effector proteins were significantly increased in the F. sacchari -infected sugarcane plants. Through fungal gene knockout, three genes (Fs06761, Fs08184, and Fs13617) were found to be important pathogenic factors for pathogens. Our results support that the FsCFEM effector proteins were virulence effectors and might be involved in regulating plant immunity. IMPORTANCE Common fungal extracellular membrane (CFEM) domain-containing protein has long been considered an essential effector, playing a crucial role in the interaction of pathogens and plant. Strategies aimed at understanding the pathogenicity mechanism of F. sacchari are eagerly anticipated to ultimately end the spread of pokkah boeng disease. Twenty FsCFEM proteins in the genome of F. sacchari have been identified, and four FsCFEM effector proteins have been found to suppress BCL2-associated X protein-triggered programmed cell death in N. benthamiana . These four effector proteins have the ability to enter plant cells and inhibit plant immunity. Furthermore, the expression of these four FsCFEM effector proteins significantly increases during the infection stage, with the three of them playing an essential role in achieving full virulence. These study findings provide a direction toward further exploration of the immune response in sugarcane. By applying these discoveries, we can potentially control the spread of disease through techniques such as host-induced gene silencing.

Towards a Therapeutically Pragmatic Molecular Definition of Hepatoblastoma

Hepatoblastoma (HB) is the most common pediatric liver cancer while also holding the distinction of being the least genomically altered of all malignancies. Three major oncogenic pathways have been identified as underlying the pathogenesis of this disease. Mouse models have demonstrated that deregulating any two is the minimal requirement for generating a tumor and that deregulating all three generates particularly aggressive ones. The terminal mediators of these three pathways, namely β-catenin of the Wnt/β-catenin, YAP of the Hippo pathway and NFE2L2/NRF of the NRF2 pathway are all transcription factors (TFs) that directly regulate hundreds-thousands of both unique and common downstream targets. Different pairwise combinations of these three TFs generate HBs with distinct growth patterns, histologies and gene expression patterns. Moreover, different patient-derived mutant forms of β-catenin also generate HBs with similarly distinct profiles. Based on the analyses of numerous tumors with these various combinations of TFs, and in different genetic backgrounds, it has been possible to identify a key set of 22 so-called “BYN genes” genes that are always similarly deregulated across all experimental HB types. A subset of these can be used to classify human HBs into those with long-term favorable and unfavorable outcomes and another subset is predictive of survival in hepatocellular carcinomas and over a dozen other human cancers. None of the BYN genes encode TFs; rather they mostly encode secreted or extracellular membrane proteins or enzymes. They therefore make more appealing targets than do the more refractory upstream TFs.

Forces and Flows at Cell Surfaces.

Flow is an important physiological signal and modulates a variety of cell functions. However, the molecular mechanisms that cells use to sense flow have remained surprisingly opaque. The first steps in flow sensing are likely to occur at the plasma membrane, the fluid barrier between the inside and outside of a cell. This membrane is an organized, two-dimensional molecular array that has both solid and fluid properties. The mobility of membrane proteins and lipids is constrained by complex interactions with the cytoskeletal protein network that supports the membrane. Physiologically relevant flows can only generate tiny forces on individual proteins, smaller than those from thermal noise. However, cells could overcome this problem by sensing micron-scale concentration gradients of extracellular membrane proteins. This is possible in cell plasma membranes because their particular physical state allows flow to sort membrane proteins laterally over the extracellular surface.

Accurate and Convenient Lung Cancer Diagnosis through Detection of Extracellular Vesicle Membrane Proteins via Förster Resonance Energy Transfer.

Tumor-derived extracellular vesicles (EVs) are promising to monitor early stage cancer. Unfortunately, isolating and analyzing EVs from a patient's liquid biopsy are challenging. For this, we devised an EV membrane proteins detection system (EV-MPDS) based on Förster resonance energy transfer (FRET) signals between aptamer quantum dots and AIEgen dye, which eliminated the EV extraction and purification to conveniently diagnose lung cancer. In a cohort of 80 clinical samples, this system showed enhanced accuracy (100% versus 65%) and sensitivity (100% versus 55%) in cancer diagnosis as compared to the ELISA detection method. Improved accuracy of early screening (from 96.4% to 100%) was achieved by comprehensively profiling five biomarkers using a machine learning analysis system. FRET-based tumor EV-MPDS is thus an isolation-free, low-volume (1 μL), and highly accurate approach, providing the potential to aid lung cancer diagnosis and early screening.

Clinicopathological correlation of Cathepsin K expression in salivary gland carcinomas; relation to patients` outcome

BackgroundSalivary gland carcinomas (SGCs) represent various groups of tumors that demonstrate marked diversity in their prognosis owing to different histology and clinical characteristics. One of the poor prognostic indicators is distant metastasis which is considered the major reason for death in SGC patients. Discovering new biomarkers is urgently required to aid in the detection of cancer onset and progression. Cathepsin K (CTSK), the lysosomal cysteine protease has a principal role in cancer invasion and progression through interaction with the tumor microenvironment, degradation of extracellular membrane proteins and destruction of the elastic lamina of blood vessels. In the English literature, little information was present about the role of CTSK in SGCs. The current study aimed to assess the immunohistochemical expression of CTSK in SGCs and correlate its expression to different clinicopathologic parameters.MethodsThe retrospective study applied to 45 cases of SGCs categorized as high-grade (33 cases) and low-grade SGCs (12 cases) following the criteria of WHO classification (2017) of head and neck tumors. All patients` clinicopathological and follow-up records were retrieved. The following statistical tests were used to study the variance of CTSK expression in SGCs concerning different clinicopathological parameters; Pearson`s Chi-square test, unpaired two-tailed student t-test, One-way ANOVA, and Post Hoc tests. Disease-free survival (DFS) and Overall survival (OS) were calculated and displayed with the Kaplan–Meier strategy and analyzed with the log-rank test. Univariate and multivariate survival analyses were performed with Cox regression. A P-value lesser than 0.05 was considered statistically significant.ResultsStrong CTSK expression was significantly related to high-grade SGCs (P = 0.000), large infiltrating carcinomas (P = 0.000), presence of nodal (P = 0.041) and distant metastasis (P = 0.009), advanced TNM clinical stage (P = 0.000), the incidence of recurrence (P = 0.009), and reduced DFS (P = 0.006). Distant metastasis was the independent predictor for DFS using Cox regression model.ConclusionsCTSK has a great role in cancer progression by triggering many signaling pathways. Its level in cancerous tissue is considered an effective index for predicting the severity and prognosis of cancer. Therefore, we indicate its utility as a prognostic tool and therapeutic target for cancer treatment.Trial registrationRetrospectively registered.

Measuring flow-mediated protein drift across stationary supported lipid bilayers

Fluid flow near biological membranes influences cell functions such as development, motility, and environmental sensing. Flow can laterally transport extracellular membrane proteins located at the cell-fluid interface. To determine whether this transport contributes to flow signaling in cells, quantitative knowledge of the forces acting on membrane proteins is required. Here, we demonstrate a method for measuring flow-mediated lateral transport of lipid-anchored proteins. We rupture giant unilamellar vesicles to form discrete patches of supported membrane inside rectangular microchannels and then allow proteins to bind to the upper surface of the membrane. While applying flow, we observe the formation of protein concentration gradients that span the membrane patch. By observing how these gradients dynamically respond to changes in applied shear stress, we determine the flow mobility of the lipid-anchored protein. We use simplified model membranes and proteins to demonstrate our method’s sensitivity and reproducibility. Our intention was to design a quantitative, reliable method and analysis for protein mobility that we will use to compare flow transport for a variety of proteins, lipid anchors, and membranes in model systems and on living cells.

The Conserved Cysteine-Rich Secretory Protein MaCFEM85 Interacts with MsWAK16 to Activate Plant Defenses.

Metarhizium anisopliae is an entomopathogenic fungus which may enhance plant growth and resistance when acting as an endophyte in host plants. However, little is known about the protein interactions nor their activating mechanisms. Common in fungal extracellular membrane (CFEM) proteins have been identified as plant immune regulators that suppress or activate plant resistance responses. Here, we identified a CFEM domain-containing protein, MaCFEM85, which was mainly localized in the plasma membrane. Yeast two-hybrid (Y2H), glutathione-S-transferase (GST) pull-down, and bimolecular fluorescence complementation assays demonstrated that MaCFEM85 interacted with the extracellular domain of a Medicago sativa (alfalfa) membrane protein, MsWAK16. Gene expression analyses showed that MaCFEM85 and MsWAK16 were significantly upregulated in M. anisopliae and M. sativa, respectively, from 12 to 60 h after co-inoculation. Additional yeast two-hybrid assays and amino acid site-specific mutation indicated that the CFEM domain and 52th cysteine specifically were required for the interaction of MaCFEM85 with MsWAK16. Defense function assays showed that JA was up-regulated, but Botrytis cinerea lesion size and Myzus persicae reproduction were suppressed by transient expression of MaCFEM85 and MsWAK16 in the model host plant Nicotiana benthamiana. Collectively, these results provide novel insights into the molecular mechanisms underlying interactions of M. anisopliae with host plants.

Extracellular Vesicle Membrane Protein Profiling and Targeted Mass Spectrometry Unveil CD59 and Tetraspanin 9 as Novel Plasma Biomarkers for Detection of Colorectal Cancer.

Extracellular vesicles (EVs) are valuable sources for the discovery of useful cancer biomarkers. This study explores the potential usefulness of tumor cell-derived EV membrane proteins as plasma biomarkers for early detection of colorectal cancer (CRC). EVs were isolated from the culture supernatants of four CRC cell lines by ultracentrifugation, and their protein profiles were analyzed by LC-MS/MS. Bioinformatics analysis of identified proteins revealed 518 EV membrane proteins in common among at least three CRC cell lines. We next used accurate inclusion mass screening (AIMS) in parallel with iTRAQ-based quantitative proteomic analysis to highlight candidate proteins and validated their presence in pooled plasma-generated EVs from 30 healthy controls and 30 CRC patients. From these, we chose 14 potential EV-derived targets for further quantification by targeted MS assay in a separate individual cohort comprising of 73 CRC and 80 healthy subjects. Quantitative analyses revealed significant increases in ADAM10, CD59 and TSPAN9 levels (2.19- to 5.26-fold, p < 0.0001) in plasma EVs from CRC patients, with AUC values of 0.83, 0.95 and 0.87, respectively. Higher EV CD59 levels were significantly correlated with distant metastasis (p = 0.0475), and higher EV TSPAN9 levels were significantly associated with lymph node metastasis (p = 0.0011), distant metastasis at diagnosis (p = 0.0104) and higher TNM stage (p = 0.0065). A two-marker panel consisting of CD59 and TSPAN9 outperformed the conventional marker CEA in discriminating CRC and stage I/II CRC patients from healthy controls, with AUC values of 0.98 and 0.99, respectively. Our results identify EV membrane proteins in common among CRC cell lines and altered plasma EV protein profiles in CRC patients and suggest plasma EV CD59 and TSPAN9 as a novel biomarker panel for detecting early-stage CRC.

Bifunctional Compounds as Molecular Degraders for Integrin-Facilitated Targeted Protein Degradation.

As effective ways to regulate protein levels, targeted protein degradation technologies have attracted great attention in recent years. Here, we established a novel integrin-facilitated lysosomal degradation (IFLD) strategy to degrade extracellular and cell membrane proteins using bifunctional compounds as molecular degraders. By conjugation of a target protein-binding ligand with an integrin-recognition ligand, the resulting molecular degrader proved to be highly efficient to induce the internalization and subsequent degradation of extracellular or cell membrane proteins in an integrin- and lysosome-dependent manner. As demonstrated in the development of BMS-L1-RGD, which is an efficient programmed death-ligand 1 (PD-L1) degrader validated both in vitro and in vivo, the IFLD strategy expands the toolbox for regulation of secreted and membrane-associated proteins and thus has great potential to be applied in chemical biology and drug discovery.

Emerging enzyme surface display systems for waste resource recovery.

The current century marks an inflection point for human progress, as the developed world increasingly comes to recognize that the ecological and socioeconomic impacts of resource extraction must be balanced with more sustainable modes of growth that are less reliant on non-renewable sources of energy and materials. This has opened a window of opportunity for cross-sector development of biotechnologies that harness the metabolic problem-solving power of microbial communities. In this context, recovery has emerged as an organizing principal to create value from industrial and municipal waste streams, and the search is on for new enzymes and platforms that can be used for waste resource recovery at scale. Enzyme surface display on cells or functionalized materials has emerged as a promising platform for waste valorization. Typically, surface display involves the use of substrate binding or catalytic domains of interest translationally fused with extracellular membrane proteins in a microbial chassis. Novel display systems with improved performance features include S-layer display with increased protein density, spore display with increased resistance to harsh conditions, and intracellular inclusions including DNA-free cells or nanoparticles with improved social licence for in situ applications. Combining these display systems with advances in bioprinting, electrospinning and high-throughput functional screening have potential to transform outmoded extractive paradigms into 'trans-metabolic" processes for remediation and waste resource recovery within an emerging circular bioeconomy.

Thiol Modifications in the Extracellular Space-Key Proteins in Inflammation and Viral Infection.

Posttranslational modifications (PTMs) allow to control molecular and cellular functions in response to specific signals and changes in the microenvironment of cells. They regulate structure, localization, stability, and function of proteins in a spatial and temporal manner. Among them, specific thiol modifications of cysteine (Cys) residues facilitate rapid signal transduction. In fact, Cys is unique because it contains the highly reactive thiol group that can undergo different reversible and irreversible modifications. Upon inflammation and changes in the cellular microenvironment, many extracellular soluble and membrane proteins undergo thiol modifications, particularly dithiol–disulfide exchange, S-glutathionylation, and S-nitrosylation. Among others, these thiol switches are essential for inflammatory signaling, regulation of gene expression, cytokine release, immunoglobulin function and isoform variation, and antigen presentation. Interestingly, also the redox state of bacterial and viral proteins depends on host cell-mediated redox reactions that are critical for invasion and infection. Here, we highlight mechanistic thiol switches in inflammatory pathways and infections including cholera, diphtheria, hepatitis, human immunodeficiency virus (HIV), influenza, and coronavirus disease 2019 (COVID-19).

Identification of Antigenic Properties of Acinetobacter baumannii Proteins as Novel Putative Vaccine Candidates Using Reverse Vaccinology Approach.

Multidrug-resistant Acinetobacter baumannii (A. baumannii) infections are becoming more prevalent all over the world. As a cost-effective and preventative method, vaccination seems to be required against this bacterium. In the present study, subtractive proteomics along with reverse vaccinology approaches was used to predict suitable therapeutics against A. baumannii. Using the Vaxign online tool, we studied over 35 genomes of A. baumannii strains and chose outer membrane and secreted proteins of A. baumannii 1656-2 as possible vaccine candidates. Then, investigations were performed on the immunogenicity, antigenic characteristics, physicochemical properties, B-cell and MHC class I, and MHC class II molecules epitope densities of proteins. After optimizing the codon of the proteins, the pcDNA3.1( +) expression construct was designed and the immunogenicity, allergenicity, and physicochemical properties of the vaccine construct were predicted. Hcp and OmpC proteins were predicted as extracellular and outer membrane proteins, respectively. These proteins interact with 10 other proteins to form a network of protein interactions with virulence properties. Immunoassays of Hcp and OmpC proteins showed antigenicity of 0.88 and 0.79, respectively. These proteins have 5 structural cell epitope points and 5 linear B epitope points. They are also able to bind to different HLA alleles of MCH class I/class II as selected immunogenic proteins and designed non-allergenic structures with solubility of 0.650 and immunogenicity score of 0.91. The results of this "in silico" study indicate high specificity and the development of a significant humoral and cellular immune response. It can be concluded that the Hcp and OmpC dual vaccine construct is one of the promising candidates against A. baumannii. The findings of this "in silico" study show excellent specificity and the emergence of a substantial humoral and cellular immune response. This is a computer-based study that needs to be tested in vitro and in vivo to corroborate the conclusions of the vaccine design procedures.

Sustained correction of hippocampal neurogenic and cognitive deficits after a brief treatment by Nutlin-3 in a mouse model of fragile X syndrome

BackgroundFragile X syndrome (FXS), the most prevalent inherited intellectual disability and one of the most common monogenic forms of autism, is caused by a loss of fragile X messenger ribonucleoprotein 1 (FMR1). We have previously shown that FMR1 represses the levels and activities of ubiquitin ligase MDM2 in young adult FMR1-deficient mice, and treatment by a MDM2 inhibitor Nutlin-3 rescues both hippocampal neurogenic and cognitive deficits in FMR1-deficient mice when analyzed shortly after the administration. However, it is unknown whether Nutlin-3 treatment can have long-lasting therapeutic effects.MethodsWe treated 2-month-old young adult FMR1-deficient mice with Nutlin-3 for 10 days and then assessed the persistent effect of Nutlin-3 on both cognitive functions and adult neurogenesis when mice were 6-month-old mature adults. To investigate the mechanisms underlying the persistent effects of Nutlin-3, we analyzed the proliferation and differentiation of neural stem/progenitor cells isolated from these mice and assessed the transcriptome of the hippocampal tissues of treated mice.ResultsWe found that transient treatment with Nutlin-3 of 2-month-old young adult FMR1-deficient mice prevents the emergence of neurogenic and cognitive deficits in mature adult FXS mice at 6 months of age. We further found that the long-lasting restoration of neurogenesis and cognitive function might not be mediated by changing intrinsic properties of adult neural stem cells. Transcriptomic analysis of the hippocampal tissue demonstrated that transient Nultin-3 treatment leads to significant expression changes in genes related to the extracellular matrix, secreted factors, and cell membrane proteins in the FMR1-deficient hippocampus.ConclusionsOur data indicates that transient Nutlin-3 treatment in young adults leads to long-lasting neurogenic and behavioral changes likely through modulating adult neurogenic niche that impact adult neural stem cells. Our results demonstrate that cognitive impairments in FXS may be prevented by an early intervention through Nutlin-3 treatment.

Comprehensive Glycomic Profiling of Breast Cancer Patients

Glycans are complex carbohydrates found on extracellular membrane lipids and proteins. They are known to be variably expressed by cancer cells, which can have partial or even novel glycan structures. Therefore, we hypothesize that the analysis of breast cancer patient glycomic profiles will result in elucidation of specific and generalizable signatures which will be useful for the diagnosis and treatment of breast cancer. Our study examines N‐glycan expression and its changes through breast cancer stages and compares them to healthy donor samples to create a catalogue of changes and profiles based on cancer stage and other tumor attributes. We are specifically investigating N‐glycans found in patient serum, on the tumor cells, and on adjacent normal tissue. Serum and tissue (tumor and normal) samples from breast cancer patients were analyzed using mass spectroscopy to determine the presence, type, and amount of over 300 unique glycan compositions. Bioinformatic analysis is being used to develop comprehensive glycomic profiles of patients and glycan signatures of cancer stages. We have found several sets of glycan compositions in the serum that appear to change over the progression of cancer. Most notably, glycans of low molecular mass (H4N2, H3N3, H5N2, H3N3F1, H4N3, H3N4, H3N5, H5N5) are rarely present in healthy samples, but are prevalent in all 4 stages of cancer patients. Alternatively, several glycans of varying molecular weight have low expression in healthy patients but higher expression in cancer patients (H4N4F1, H6N2, H5N4F1, H3N4F1, H5N4, H5N2, H4N5F1). Comparing serum and tumor glycan profiles to those of healthy sera and tissue controls will allow for a more comprehensive understanding of the aberrant expression of glycan structures in breast cancer. The collected information has the potential for more accurate diagnoses and personalized treatments. Prospective uses of differential glycomic profiles could allow oncologists and their patients a more nuanced understanding of the tumor and its characteristics.

Systemic Identification and Functional Characterization of Common in Fungal Extracellular Membrane Proteins in Lasiodiplodia theobromae

Plant pathogenic fungi deploy secreted proteins into apoplastic space or intracellular lumen to promote successful infections during plant-pathogen interactions. In the present study, fourteen CFEM domain-containing proteins were systemically identified in Lasiodiplodia theobromae and eight of them were functionally characterized. All eight proteins were confirmed to be secreted into extracellular space by a yeast signal peptide trapping system. The transcriptional levels of most CFEM genes, except for LtCFEM2 and LtCFEM6, were significantly elevated during infection. In addition, almost all LtCFEM genes, apart from LtCFEM2, LtCFEM3, and LtCFEM6, were transcriptionally up-regulated at 35°C in contrast to that at 25°C and 30°C. As two elicitors, LtCFEM1 induced local yellowish phenotype and LtCFEM4 triggered cell death in Nicotiana benthamiana leaves. Furthermore, these proteins displayed distinct subcellular localizations when expressed transiently in N. benthamiana. Moreover, two genes, LtCFEM7 and LtCFEM8, were found to be spliced alternatively by RT-PCR and sequencing. Therefore, our data suggest that LtCFEM proteins play important roles in multiple aspects, including pathogenicity and plant immune response, which will enhance our understanding of the sophisticated pathogenic mechanisms of plant opportunistic pathogen L. theobromae.

Seropositive Muscle-Specific Tyrosine Kinase Myasthenia Gravis Presenting as a Late-Onset Isolated Sixth Nerve Palsy: A Case Report and a Brief Review of Subtypes of Myasthenia Gravis

Autoimmune myasthenia gravis (MG) is a well-characterized post-synaptic disorder of neuromuscular transmission. Immunologically, there is complement activation with autoantibodies binding to the acetylcholine receptor (AChR), leading to cross-linking and internalization of the receptor. The diminished functional clustering leads to impaired folding of the post-synaptic membrane. The antibodies generated by the autoimmune process are directed at the various components of the post-synaptic membrane and its scaffolding, including the AChR, muscle-specific tyrosine kinase (MuSK), low-density lipoprotein receptor-related protein 4 (LRP4), and the other recently described epitopes including the extracellular membrane proteins agrin and collagen Q (ColQ). MuSK MG is phenotypically different from classic AChR-antibody-mediated MG by a more frequent presentation of bulbar weakness, less responsiveness to symptomatic therapy with acetylcholinesterase inhibitors, the absence of a thymoma, and a better therapeutic response to a cluster of differentiation (CD-20) B-cell therapy such as rituximab. The pleiotropic ocular findings of ocular MG include ptosis, fluctuating and variable involvement of cranial nerves III, IV, and VI, pseudo-internuclear ophthalmoplegia (INO), near-complete or complete ophthalmoplegia, and variable gaze palsies. To our knowledge, we present one of the very few reported cases of MuSK MG presenting as isolated sixth nerve palsy. The localization of a sixth nerve palsy with lateral rectus muscle weakness can be due to disease anywhere along its path from the abducens nucleus, coursing at the skull base through Dorello's canal, through the cavernous sinus, and along its path through the superior orbital fissure and into the orbits. A painless sixth nerve palsy should alert the clinician to MuSK-MG as we outline in this case report.

Gold nanoparticles conjugated with DNA aptamer for photoacoustic detection of human matrix metalloproteinase-9

Matrix metalloproteinase-9 (MMP-9) plays major roles in extracellular matrix (ECM) remodeling and membrane protein cleavage, suggesting a high correlation with cancer cell invasion and tumor metastasis. Here, we present a contrast agent based on a DNA aptamer that can selectively target human MMP-9 in the tumor microenvironment (TME) with high affinity and sensitivity. Surface modification of plasmonic gold nanospheres with the MMP-9 aptamer and its complementary sequences allows the nanospheres to aggregate in the presence of human MMP-9 through DNA displacement and hybridization. Aggregation of gold nanospheres enhances the optical absorption in the first near-infrared window (NIR-I) due to the plasmon coupling effect, thereby allowing us to detect the aggregated gold nanospheres within the TME via ultrasound-guided photoacoustic (US/PA) imaging. Selective and sensitive detection of human MMP-9 via US/PA imaging is demonstrated in solution of nanosensors with the pre-treatment of human MMP-9, in vitro in cell culture, and in vivo in a xenograft murine model of human breast cancer.

Selective Degradation of PD‐L1 in Cancer Cells by Enzyme‐Instructed Self‐Assembly

AbstractHere, a novel strategy to selectively degrade membrane proteins of programmed cell death‐ligand 1 (PD‐L1) in cancer cells is reported. 4T1 cells show high levels of extracellular alkaline phosphatase (ALP) and membrane proteins of PD‐L1. Therefore, peptide derivatives capable of responding to ALP and binding to PD‐L1 are designed. Enzyme‐instructed self‐assembly (EISA) by ALP and surface‐induced self‐assembly by PD‐L1 of Comp. 3 leads to the selective formation of nanomaterials around PD‐L1 on the cell membrane, which is similar to attaching a hydrophobic label to the surface of PD‐L1 to simulate its partial denaturation state. When taken up by cells, PD‐L1 could be further degraded by the proteasome pathway in the cytoplasm. This process does not occur in the normal cell line, LO2 cells, which express relatively low levels of ALP. In addition to concentration and time dependence, the selective knock‐out of PD‐L1 by the strategy is reversible by simply removing Comp. 3. In vivo studies reveal that Comp. 3 inhibits tumor growth in a tumor‐bearing mouse model. The study offers a novel platform for the precise degradation of membrane proteins using EISA, which might ultimately lead to the development of novel nanomedicines to treat diseases.