Serine Protease Research Articles

Distinct granzyme k expression in immune cells: a single-cell rna-seq meta-analysis.

Granzymes are essential serine proteases in cytotoxic T cells and natural killer (NK) cells, with GZMK's expression being less understood. This study aims to uncover GZMK expression profiles across various immune cell types using single-cell RNA sequencing meta-analysis. This study aims to uncover GZMK expression profiles across various immune cell types using single-cell RNA sequencing meta-analysis. We conducted a meta-analysis using cellxgene, an interactive data exploration platform developed by the Chan Zuckerberg Initiative. We focused on mature T cells, NK cells, B cells, and NKT cells. We also checked transcription factor binding sites at the granzyme gene promoter regions using JASPAR. Comparative analysis was also done using mouse single-cell RNA sequencing data. GZMK was the most lowly expressed in NK cells and mature NKT cells in most tissues except for colon and lymph nodes. In mature T cells, GZMK is similarly or more highly expressed than other granzymes. HBCA data revealed weak expression of GZMK in NK cells but strong expression in effector memory CD8-positive, alpha-beta T cells. Combined data shows no significant difference in GZMK expression between cell types. Subtype analysis shows that GZMK expression was higher in CD16-negative, CD56-bright NK cells when compared to CD16-positive, CD56-dim NK cells. We also identified unique transcription factor binding sites for GZMK. While this pattern in mouse data with low Gzmk expression in NK cells and higher T cells was repeated. GZMK expression is distinctively regulated among immune cells and tissues, with unique promoter regions and transcription factor binding sites contributing to this differential expression. These insights into GZMK's role in immune function and regulation offer potential therapeutic targets.

Peptidic Boronic Acid Plasmodium falciparum SUB1 Inhibitors with Improved Selectivity over Human Proteasome.

Plasmodium falciparum subtilisin-like serine protease 1 (PfSUB1) is essential for egress of invasive merozoite forms of the parasite, rendering PfSUB1 an attractive antimalarial target. Here, we report studies aimed to improve drug-like properties of peptidic boronic acid PfSUB1 inhibitors including increased lipophilicity and selectivity over human proteasome (H20S). Structure-activity relationship investigations revealed that lipophilic P3 amino acid side chains as well as N-capping groups were well tolerated in retaining PfSUB1 inhibitory potency. At the P1 position, replacing the methyl group with a carboxyethyl substituent led to boralactone PfSUB1 inhibitors with remarkably improved selectivity over H20S. Combining lipophilic end-capping groups with the boralactone reduced the selectivity over H20S. However, compound 4c still showed >60-fold selectivity versus H20S and low nanomolar PfSUB1 inhibitory potency. Importantly, this compound inhibited the growth of a genetically modified P. falciparum line expressing reduced levels of PfSUB1 13-fold more efficiently compared to a wild-type parasite line.

A comparative review of the application value of FAPI PET/CT and 18F-FDG PET/CT in lung cancer.

Fibroblast activation protein inhibitor (FAPI) positron emission tomography/computed tomography (PET/CT) is a multimodal imaging technique that combines PET and CT, utilizing FAP inhibitors as radiotracers. Fibroblast activation protein, a serine protease highly expressed in many epithelial tumor-associated fibroblasts, plays a crucial role in tumor stroma formation and remodeling. Through the detection of FAP expression, FAPI PET/CT facilitates the diagnosis and staging of both benign and malignant pulmonary tumors. In contrast to traditional fluorine-18-fluorodeoxyglucose (18F-FDG) PET/CT focusing on glucose metabolism, FAPI PET/CT offers benefits such as enhanced specificity, reduced background noise, accelerated imaging speed, and decreased radiation exposure. This review provides an overview of the progress in applying FAPI PET/CT and 18F-FDG PET/CT in pulmonary malignancies and discusses current challenges and future prospects.

Biochemical characteristics of carbohydrase and serine protease enzymes of beet moth, Scrobipalpa ocellatella (Lepidoptera; Gelechiidae)

Biochemical characteristics of carbohydrase and serine protease enzymes of beet moth, Scrobipalpa ocellatella (Lepidoptera; Gelechiidae)

Receptors Involved in COVID-19-Related Anosmia: An Update on the Pathophysiology and the Mechanistic Aspects

Olfactory perception is an important physiological function for human well-being and health. Loss of olfaction, or anosmia, caused by viral infections such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has received considerable attention, especially in persistent cases that take a long time to recover. This review discusses the integration of different components of the olfactory epithelium to serve as a structural and functional unit and explores how they are affected during viral infections, leading to the development of olfactory dysfunction. The review mainly focused on the role of receptors mediating the disruption of olfactory signal transduction pathways such as angiotensin converting enzyme 2 (ACE2), transmembrane protease serine type 2 (TMPRSS2), neuropilin 1 (NRP1), basigin (CD147), olfactory, transient receptor potential vanilloid 1 (TRPV1), purinergic, and interferon gamma receptors. Furthermore, the compromised function of the epithelial sodium channel (ENaC) induced by SARS-CoV-2 infection and its contribution to olfactory dysfunction are also discussed. Collectively, this review provides fundamental information about the many types of receptors that may modulate olfaction and participate in olfactory dysfunction. It will help to understand the underlying pathophysiology of virus-induced anosmia, which may help in finding and designing effective therapies targeting molecules involved in viral invasion and olfaction. To the best of our knowledge, this is the only review that covered all the receptors potentially involved in, or mediating, the disruption of olfactory signal transduction pathways during COVID-19 infection. This wide and complex spectrum of receptors that mediates the pathophysiology of olfactory dysfunction reflects the many ways in which anosmia can be therapeutically managed.

The expanded immunoregulatory protease network in mosquitoes is governed by gene co-expression.

Serine protease cascades regulate key innate immune responses. In mosquitoes, these cascades involve clip-domain serine proteases and their non-catalytic homologs (CLIPs), forming a complex network whose make-up and structural organization is not fully understood. This study assessed the impact of 85 CLIPs on humoral immunity in Anopheles gambiae. By coupling RNAi with assays measuring antimicrobial activity and melanization, we identified 27 CLIPs as immunoregulators that together form two distinct subnetworks. CLIPs regulating antimicrobial activity were found to control infection resistance, as knockdowns reduced bacterial load and improved survival. Furthermore, our analysis of CLIP gene expression unveiled a novel immunoregulatory mechanism reliant on protease baseline co-expression rather than infection-induced upregulation. These findings underscore that despite its complexity mosquito immune regulation may be targeted for malaria interventions.

Proteolytic Regulation in the Biosynthesis of Natural Product Via a ClpP Protease System.

Protein degradation is a tightly regulated biological process that maintains bacterial proteostasis. ClpPs are a highly conserved family of serine proteases that associate with the AAA + ATPase (an ATPase associated with diverse cellular activities) to degrade protein substrates. Identification and biochemical characterization of protein substrates for the AAA + ATPase-dependent ClpP degradation systems are considered essential for gaining an understanding of the molecular operation of the complex ClpP degradation machinery. Consequently, expanding the repertoire of protein substrates that can be degraded in vitro and within bacterial cells is necessary. Here, we report that AAA + ATPase-ClpP proteolytic complexes promote degradation of the secondary metabolite surfactin synthetases SrfAA, SrfAB, and SrfAC in Bacillus subtilis. On the basis of in vitro and in-cell studies coupled with activity-based protein profiling of nonribosomal peptide synthetases, we showed that SrfAC is targeted to the ClpC-ClpP proteolytic complex, whereas SrfAA is hydrolyzed not only by the ClpC-ClpP proteolytic complex but also by different ClpP proteolytic complexes. Furthermore, SrfAB does not appear to be a substrate for the ClpC-ClpP proteolytic complex, thereby implying that other ClpP proteolytic complexes are involved in the degradation of this surfactin synthetase. Natural product biosynthesis is regulated by the AAA + ATPase-ClpP degradation system, indicating that protein degradation plays a role in the regulatory stages of biosynthesis. However, few studies have examined the regulation of protein degradation levels. Furthermore, SrfAA, SrfAB, and SrfAC were identified as protein substrates for AAA + ATPase-ClpP degradation systems, thereby contributing to a better understanding of the complex ClpP degradation machinery.

Association of VEGFA polymorphisms with the risk of oesophageal cancer in Punjab, India: A case-control study

Background & objectives Vascular endothelial growth factor (VEGF) is one of the most important angiogenic factors which stimulates tumour progression induction of endothelial cell migration and division, inhibition of the apoptosis of endothelial cells, induction of serine protease activity and enhancement of vascular permeability. This study aimed to investigate the correlation of VEGF+405G/C,-7C/T and+936C/T polymorphisms with oesophageal cancer risk. Methods DNA samples of 464 subjects (231 sporadic oesophageal cancer affected individuals and 233 controls) were genotyped forVEGF+936C/T,+405G/C and-7C/T polymorphisms. VEGF+936C/T and +405G/C polymorphisms were genotyped by PCR-RFLP method whereas VEGF-7C/T polymorphism was genotyped using Amplification refractory mutation system-polymerase chain reaction (ARMS-PCR). Results CT genotype of VEGF-7C/T polymorphism was significantly associated with reduced risk of oesophageal cancer. VEGF-7C/T polymorphism was significantly associated with reduced risk of oesophageal cancer underdominant, co-dominant, over dominant and log-additive genetic models in total patients and in the female group. C+936G+405T-7 haplotype was significantly associated with decreased risk (P=0.01)of oesophageal cancer in total patients and also in the male group (P=0.02). Interpretation & conclusions In future, replication of the findings of the present study in a larger sample from different ethnic groups, along with functional analysis, may be insightful for the role of VEGFA polymorphisms in the pathogenesis of oesophageal cancer. Identification of the correlation of VEGF variants with specific therapy in oesophageal cancer may help in better selection of patients and monitoring treatment response in VEGF-therapy.

Pharmacophore-Assisted Covalent Docking Identifies a Potential Covalent Inhibitor for Drug-Resistant Genotype 3 Variants of Hepatitis C Viral NS3/4A Serine Protease

The emergence of drug-resistance-inducing mutations in Hepatitis C virus (HCV) coupled with genotypic heterogeneity has made targeting NS3/4A serine protease difficult. In this work, we investigated the mutagenic variations in the binding pocket of Genotype 3 (G3) HCV NS3/4A and evaluated ligands for efficacious inhibition. We report mutations at 14 positions within the ligand-binding residues of HCV NS3/4A, including H57R and S139P within the catalytic triad. We then modelled each mutational variant for pharmacophore-based virtual screening (PBVS) followed by covalent docking towards identifying a potential covalent inhibitor, i.e., cpd-217. The binding stability of cpd-217 was then supported by molecular dynamic simulation followed by MM/GBSA binding free energy calculation. The free energy decomposition analysis indicated that the resistant mutants alter the HCV NS3/4A–ligand interaction, resulting in unbalanced energy distribution within the binding site, leading to drug resistance. Cpd-217 was identified as interacting with all NS3/4A G3 variants with significant covalent docking scores. In conclusion, cpd-217 emerges as a potential inhibitor of HCV NS3/4A G3 variants that warrants further in vitro and in vivo studies. This study provides a theoretical foundation for drug design and development targeting HCV G3 NS3/4A.

Biotechnological potential of Brazilian snake venom in the production of biologically active drugs: Literature review

Despite being feared, Brazilian venomous snakes can be reservoirs of important resources of medicinal interest, since the venoms of these animals have an invaluable potential for biologically active substances found in nature. Pharmacological and biochemical studies carried out in recent decades have shown the diversity of proteins with enzymatic activity, toxins, peptides, bioactive amines, among other compounds in snake venoms. The aim of this literature review was to describe the pharmaceutical biotechnological potential of Brazilian snake venom and to demonstrate the importance of these toxins and their pharmacological derivatives for public health. Since 1894, when they were studied by Vital Brazil, molecules derived from Brazilian snake venom have shown significant biological activity. These molecules have been proven to be efficient antimicrobial, antifungal and antiprotozoal agents, mainly due to their activity as serine proteases, hyaluronidases, L-amino acid oxidases (LAAO) and acetylcholinesterases, among other well-known substances. Brazilian snake venoms have thus been the target of a series of studies, which have borne excellent fruit, for example in the development of various drugs, such as Captopril®, which is derived from a peptide isolated from the venom of Bothrops jararaca, and fibrin sealant, derived from molecules in the venom of Crotalus durissus terrificus associated with cryoprecipitate from the blood plasma of buffaloes. With the analysis of biotechnologies applied to various molecules with the capacity for pharmacological, biologically active action, it is clear that this field is vast for new research and due to the great diversity of compounds in snake venoms. Certainly, as research progresses, new biologically active drugs derived from snake venom should become available for use and benefit of the population. Thus, studies related to the chemical composition of snake venoms should be encouraged, as well as those related to their pharmaceutical biotechnological potential.

Snake Venom : A Bioactive Cocktail and its Therapautic use

From ancient time whenever we face any problem, we came to nature for the solution. Our nature is self renewable, she has solution for every problem, but we have to find the right path to respond more quickly. This review is regarding chemical nature of snake venom and about different life saving drugs extracted from snake venom. Venoms were originated from non poisonous proteins, and with subsequent generations under evolution these proteins become mutated and modified multiple times to develop deadly poison. With every modification venom becomes enriched with new diverse group of compounds. Main constituents of venom is hydrolyses, PLA2, phosphodiesterases, serine proteases and metalloproteases. Snakes use their venom to paralyzed their prey and to digest food, but these components can be used to treat various life threatening disease. Cobra, krait and rattle snakes are most poisionous snakes and their venom contains different enzymes, are classified as neurotoxic, haemotoxic and cytotoxic in nature and these have different impact upon exposure to animal. Neurotoxic venom destroy nervous system, haemotoxic venom leads to the destruction of red blood cells and also interfere with blood clotting factors, whereas cytotoxic venom can cause necrosis and apoptosis. With the discovary of modern techniques, isolation of different proteins and peptides and analysis of their nature are become easier. Most widely used drugs extracted from snake venom are Tirofiban, Eptifibatide, Captopril, enalapril, cobratide and batroxobin ete. The source of tirofiban, captopril, enalapril batroxobin is the venom of different viper species. On the other hand, eptifibatide is extracted from rattle snake and cobratide is from Chinese cobra .

Characterization of Inflammation/Immune-, Acute Phase-, Extracellular Matrix-, Adhesion-, and Serine Protease-Related Proteins in the Amniotic Fluid of Women With Early Preterm Prelabor Rupture of Membranes.

To determine whether altered concentrations of various inflammation/immune-, acute phase-, extracellular matrix-, adhesion-, and serine protease-related proteins in the amniotic fluid (AF) are independently associated with microbial invasion of the amniotic cavity and/or intra-amniotic inflammation (MIAC/IAI), imminent spontaneous preterm delivery (SPTD; ≤7 days), and major neonatal morbidity/mortality (NMM) in women with early preterm prelabor rupture of membranes (PPROM). This was a retrospective cohort study involving 111 singleton pregnant women with PPROM (24-31 weeks) undergoing amniocentesis to diagnose MIAC/IAI. The following proteins were measured in stored AF samples by enzyme-linked immunosorbent assay (ELISA): APRIL, DKK-3, Gal-3BP, IGFBP-2, IL-8, VDBP, lumican, MMP-2, MMP-8, SPARC, TGFBI, TGF-β1, E-selectin, ICAM-5, P-selectin, haptoglobin, hepcidin, SAA1, kallistatin, and uPA. Multivariate logistic regression analyses revealed that (i) elevated APRIL, IL-8, MMP-8, and TGFBI levels in the AF, reduced lumican and SPARC levels in the AF, and high percentages of samples above the lower limit of quantification for AF TGF-β1 and uPA were significantly associated with MIAC/IAI; (ii) elevated AF levels of IL-8 and MMP-8 were significantly associated with SPTD within 7 days; and (iii) elevated AF IL-6 levels were significantly associated with increased risk for major NMM, when adjusted for baseline covariates. ECM (lumican, SPRAC, TGFBI, and TGF-β1)- and serine protease (uPA)-associated proteins in the AF are involved in the regulation of the host response to infection/inflammation in the amniotic cavity, whereas AF inflammation (IL-8, MMP-8, and IL-6)-associated mediators are implicated in the development of preterm parturition and major NMM in early PPROM.

Use of protease substrate specificity screening in the rational design of selective protease inhibitors with unnatural amino acids: Application to HGFA, matriptase, and hepsin.

Inhibition of the proteolytic processing of hepatocyte growth factor (HGF) and macrophage stimulating protein (MSP) is an attractive approach for the drug discovery of novel anticancer therapeutics which prevent tumor progression and metastasis. Here, we utilized an improved and expanded version of positional scanning of substrate combinatorial libraries (PS-SCL) technique called HyCoSuL to optimize peptidomimetic inhibitors of the HGF/MSP activating serine proteases, HGFA, matriptase, and hepsin. These inhibitors have an electrophilic ketone serine trapping warhead and thus form a reversible covalent bond to the protease. We demonstrate that by varying the P2, P3, and P4 positions of the inhibitor with unnatural amino acids based on the protease substrate preferences learned from HyCoSuL, we can predictably modify the potency and selectivity of the inhibitor. We identified the tetrapeptide JH-1144 (8) as a single digit nM inhibitor of HGFA, matriptase and hepsin with excellent selectivity over Factor Xa and thrombin. These unnatural peptides have increased metabolic stability relative to natural peptides of similar structure. The tripeptide inhibitor PK-1-89 (2) has excellent pharmacokinetics in mice with good compound exposure out to 24 h. In addition, we obtained an X-ray structure of the inhibitor MM1132 (15) bound to matriptase revealing an interesting binding conformation useful for future inhibitor design.

Pan-cancer analysis of SERPINE1 with a concentration on immune therapeutic and prognostic in gastric cancer.

The serine protease inhibitor clade E member 1 (SERPINE1) is a key modulator of the plasminogen/plasminase system and has been demonstrated to promote tumor progression and metastasis in various tumours. However, although much literature has explored the cancer-promoting mechanism of SERPINE1, the pan-cancer analyses of its predictive value and immune response remain unexplored. The differential expression, and survival analysis of SERPINE1 expression in multiple cancers were analysed using The Cancer Genome Atlasand Genotype-Tissue Expression database. Kaplan-Meier (K-M) plotter and survival data analysis were used to analyze the prognostic value of SERPINE1 expression, including overall survival (OS), disease-specific survival, disease-free interval and progression-free interval and investigated the relationship of SERPINE1 expression with microsatellite instability. We further analysed the correlation between the expression of SERPINE1 and immune infiltration. The Kyoto Encyclopaedia of Genes and Genomes pathway was used for enrichment analysis, and the Gene Set Enrichment Analysis (GSEA) database was used to perform pathway analysis. Finally, invitro experiments demonstrated that knockdown or overexpression of SERPINE1 could alter the proliferation and migration of gastric cancer (GC) cells. The results indicated that SERPINE1 expression levels different significantly between cancer and normal tissues, meanwhile, it was highly expressed in various cancers. By analysing online data, it has been observed that the gene SERPINE1 exhibits heightened expression levels across a variety of human cancers, significantly impacting patient survival rates. Notably, the presence of SERPINE1 was strongly associated with decrease OS and disease-free survival in individuals diagnosed with GC. Furthermore, an observed link indicates that higher levels of SERPINE expression are associated with increased infiltration of immune cells in GC. Finally, invitro experiments showed that knockdown or overexpression of SERPINE1 inhibited the growth, and migration, of GC cells. SERPINE1expression potentially represents a novel prognostic biomarker due to its significant association with immune cell infiltration in GC. This study shows that SERPINE1 is an oncogene that participates in regulating the immune infiltration and affecting the prognosis of patients in multiple cancers, especially in GC. These findings underscore the importance of further investigating the role of SERPINE1 in cancer progression and offer a promising direction for the development of new therapeutic strategies.

S. aureus Eap is a polyvalent inhibitor of neutrophil serine proteases

Staphylococcus aureus expresses three high-affinity neutrophil serine protease (NSP) inhibitors known as the extracellular adherence protein domain (EAPs) proteins. Whereas EapH1 and EapH2 are comprised of a single EAP domain, the modular extracellular adherence protein (Eap) from S. aureus strain Mu50 consists of four EAP domains. We recently reported that EapH2 can simultaneously bind and inhibit cathepsin-G (CG) and neutrophil elastase (NE), which are the two most abundant NSPs. This unusual property of EapH2 arises from independent CG and NE-binding sites that lie on opposing faces of its EAP domain. Here we used X-ray crystallography and enzyme assays to show that all four individual domains of Eap (i.e. Eap1, Eap2, Eap3, and Eap4) exhibit an EapH2-like ability to form ternary complexes with CG and NE that inhibit both enzymes simultaneously. We found that Eap1, Eap2, and Eap3 have similar functional profiles insofar as NSP inhibition is concerned, but that Eap4 displays an unexpected ability to inhibit two NE enzymes simultaneously. Using X-ray crystallography, we determined that this second NE-binding site in Eap4 arises through the same region of its EAP domain that also comprises its CG-binding site. Interestingly, small angle X-ray scattering data showed that stable tail-to-tail dimers of the NE/Eap4/NE ternary complex exist in solution. This arrangement is compatible with NSP-binding at all available sites in a two-domain fragment of Eap. Together, our work implies that Eap is a polyvalent inhibitor of NSPs. It also raises the possibility that higher-order structures of NSP-bound Eap may have unique functional properties.

Gene editing of pigs to control influenza A virus infections

Proteolytic activation of the hemagglutinin (HA) glycoprotein by host cellular proteases is pivotal for influenza A virus (IAV) infectivity. Highly pathogenic avian influenza viruses possess the multibasic cleavage site of the HA which is cleaved by ubiquitous proteases, such as furin; in contrast, the monobasic HA motif is recognized and activated by trypsin-like proteases, such as the transmembrane serine protease 2 (TMPRSS2). Here, we aimed to determine the effects of TMPRSS2 on the replication of pandemic H1N1 and H3N2 subtype IAVs in the natural host, the pig. The use of the CRISPR/Cas 9 system led to the establishment of homozygous gene edited (GE) TMPRSS2 knockout (KO) pigs. Delayed IAV replication was demonstrated in primary respiratory cells of KO pigs in vitro. IAV infection in vivo resulted in significant reduction of virus shedding in the upper respiratory tract, and lower virus titers and pathological lesions in the lower respiratory tract of TMPRSS2 KO pigs as compared to wild-type pigs. Our findings support the commercial use of GE pigs to mitigate influenza A virus infection in pigs, as an alternative approach to prevent zoonotic influenza A transmissions from pigs to human.

SERPINA3 is a marker of cartilage differentiation and is essential for the expression of extracellular matrix genes during early chondrogenesis

Serine proteinase inhibitors (serpins) are a family of structurally similar proteins which regulate many diverse biological processes from blood coagulation to extracellular matrix (ECM) remodelling. Chondrogenesis involves the condensation and differentiation of mesenchymal stem cells (MSCs) into chondrocytes which occurs during early development. Here, and for the first time, we demonstrate that one serpin, SERPINA3 (gene name SERPINA3, protein also known as alpha-1 antichymotrypsin), plays a critical role in chondrogenic differentiation. We observed that SERPINA3 expression was markedly induced at early time points during in vitro chondrogenesis. We examined the expression of SERPINA3 in human cartilage development, identifying significant enrichment of SERPINA3 in developing foetal cartilage compared to total limb, which correlated with well-described markers of cartilage differentiation. When SERPINA3 was silenced using siRNA, cartilage pellets were smaller and contained lower proteoglycan as determined by dimethyl methylene blue assay (DMMB) and safranin-O staining. Consistent with this, RNA sequencing revealed significant downregulation of genes associated with cartilage ECM formation perturbing chondrogenesis. Conversely, SERPINA3 silencing had a negligible effect on the gene expression profile during osteogenesis suggesting the role of SERPINA3 is specific to chondrocyte differentiation. The global effect on cartilage formation led us to investigate the effect of SERPINA3 silencing on the master transcriptional regulator of chondrogenesis, SOX9. Indeed, we observed that SOX9 protein levels were markedly reduced at early time points suggesting a role for SERPINA3 in regulating SOX9 expression and activity. In summary, our data support a non-redundant role for SERPINA3 in enabling chondrogenesis via regulation of SOX9 levels.

Analysis of the dynamics of gene expression in patients with acute COVID-19 and in recovery period

Massive infections of people with the previously unidentified SARS-CoV-2 virus have been a shock to the global healthcare system. While many studies have focused on the clinical manifestations of the disease and its treatment methods, understanding the molecular and genetic aspects of infection has proved critical to understanding the pathogenetic mechanisms of host-pathogen interaction. The scientists focused on issues of gene expression and their regulation in response to SARS-CoV-2 infection. In particular, we conducted a study of gene expression within the framework of the project «COVID-19: Scientific and technological justification of the response system to the spread of new respiratory infections, including coronavirus infection», the results of which are presented in the article [1].Hence, variations in the gene expression profiles of TLR 3 (Toll-Like receptor), TLR 7, TLR 4, ACE 2 (Angiotensing converting enzyme), TMPRSS 13 (Transmembrane serine protease), INF-Ɣ (Interferon gamma), and IL 4 (Interleukin) were discerned across all investigated cohorts, including those assessed six months post-recovery. There is a complex of pathways of immune system to defense with the infection including the involvement of toll-like receptor signaling pathways which contact of viral particles. Moreover, the surge in ACE2 expression demonstrates the multifunctional role as gates for Sars-Cov-2 to enter cells and activation of innate immunity responses. Although, the upregulation of INF-Ɣ and IL-4 as proinflammatory cytokines were contributed to the initiation and the progression of the cytokine storm.

Single-cell RNA-sequencing reveals Eriocheir sinensis hemocyte subpopulations and their molecular responses to Spiroplasma eriocheiris infection

The hemocytes is the central part of the Eriocheir sinensis innate immune system, however, their subclasses have not been completely defined and the single-cell gene transcriptional profiles and functions are still unknown. In the present study, the single-cell mRNA sequencing (scRNA-seq) was carried out to build a single-cell gene expression map of E. sinensis hemocytes and further analysis of the interaction molecular mechanism between the crab and novel aquatic pathogens Spiroplasma eriocheiris. By scRNA-seq, we successfully identified nine cell clusters (Hem1–9) and predicted their functions according to the specific marker genes. Pseudotime inference analysis revealed that all the identified cells undergo three stages, state 1 (the initial point), state 2 and state 3 (the other two branches). After being infected by S. eriocheiris, lots of different expression genes (DEGs) were identified in each cell cluster compared with the control group (with fold change ≥2 and p-value <0.05). The bioinformatics analysis found that the Hem1 and Hem5 were the main cell clusters synthesis and release of antimicrobial peptides (AMPs) for help the host against the pathogen infection. The Hem5 also could activate the proPO system to participate in the process of host eliminate the invading S. eriocheiris. Hem7, Hem8, and Hem9 were mainly responsible for killing the invading pathogen through phagocytosis. In particular, there were also many immune-related genes identified, including β-glucan binding protein (BGBP), Clip-domain serine proteases (CLIPs), phenoloxidase 1 (PO1), anti-lipopolysaccharide factors (ALFs), syndecan, and lysozyme C, which not only played critical roles in the crab immune system but also could serve as the marker genes for identifying cell subgroup. In conclusion, the results obtained in the present study could provide pivotal insights into the crab hemocytes cell-type-specific molecular mechanisms response S. eriocheiris infection at a single-cell level.

In vitro and in vivo analyses of eFAP: a novel FAP-targeting small molecule for radionuclide theranostics and other oncological interventions

BackgroundFibroblast activation protein (FAP), a transmembrane serine protease overexpressed by cancer-associated fibroblasts in the tumor stroma, is an interesting biomarker for targeted radionuclide theranostics. FAP-targeting radiotracers have demonstrated to be superior to [18F]FDG PET/CT in various solid cancers. However, these radiotracers have suboptimal tumor retention for targeted radionuclide therapy (TRT). We aimed to develop a novel FAP-targeting pharmacophore with improved pharmacokinetics by introducing a substitution at the 8-position of (4-quinolinoyl)-glycyl-2-cyanopyrrolidine, which allows for conjugation of a chelator, dye, or other payloads.ResultsHere we showed the synthesis of DOTA-conjugated eFAP-6 and sulfo-Cyanine5-conjugated eFAP-7. After chemical characterization, the uptake and specificity of both tracers were determined on FAP-expressing cells. In vitro, [111In]In-eFAP-6 demonstrated a superior affinity and a more rapid, although slightly lower, peak uptake than gold standard [111In]In-FAPI-46. Confocal microscopy demonstrated a quick FAP-mediated internalization of eFAP-7. Studies with HT1080-huFAP xenografted mice confirmed a more rapid uptake of [177Lu]Lu-eFAP-6 vs. [177Lu]Lu-FAPI-46. However, tumor retention at 24 h post injection of [177Lu]Lu-eFAP-6 was lower than that of [177Lu]Lu-FAPI-46, hereby currently limiting its use for TRT.ConclusionThe superior affinity and faster tumor accumulation of eFAP-6 over FAPI-46 makes it a suitable compound for radionuclide imaging. After further optimization, the eFAP series has great potential for various oncological interventions, including fluorescent-guided surgery and effective targeted radionuclide theranostics.