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PubMed1978

Neutrophil chemotaxis: the kinetics of cellular locomotion in vitro.

The Boyden chamber system has been used to study the motile response of rabbit peritoneal neutrophils to partially purified casein cytotaxin. The results of these in vitro experiments indicate that neutrophils respond in two distinct ways to cytotaxin. Firstly, they increase their innate random mobility. Secondly, they respond to the information contained in a cytotaxin concentration gradient by exhibiting directional movement with a velocity vector directed towards the higher cytotaxin concentration. Since the two responses are kinetically distinct, one may infer that they are mechanistically distinct.

Animals↗

PubMed2026

Establishment of a STAT6 Reporter Assay for Screening Environmental Toxicants Affecting Allergic Airway Inflammation.

Air pollution-associated allergic airway inflammation is an increasing public health concern. Interleukin‑4 (IL‑4) and interleukin‑13 (IL‑13), which activate the Signal Transducer and Activator of Transcription 6 (STAT6) pathway, a central mediator of allergic airway inflammation, may modulate the respiratory toxicities of pollutants. The present study describes the generation and validation of a stable STAT6 luciferase reporter assay in human airway epithelial cells for evaluating environmental toxicants that modulate STAT6 signaling. Human bronchial epithelial 16HBE14o- cells were transduced with a STAT6-responsive luciferase reporter using a lentiviral vector, followed by optimization of puromycin selection and multiplicity of infection, and monoclonal isolation by limiting dilution. A stable clone with strong and reproducible induction across serial passages was selected. Reporter responsiveness was validated by IL-4/IL-13 stimulation, and STAT6 dependence was confirmed using selective STAT6, STAT5, and STAT3 inhibitors. Assay performance was quantified by Z'-factor analysis, which indicated reproducible signal separation. Furthermore, the assay was applied to individual air-pollution constituents, and benzo[b]fluoranthene and particulate matter significantly increased STAT6 reporter activity. This method provides a scalable approach for measuring STAT6 activity in airway epithelial cells and for prioritizing environmental toxicants that modulate allergic airway signaling.

STAT6 Transcription Factor↗

PubMed2026

Factor IX Padua AAV gene therapy in adolescents with hemophilia B: a phase 1 trial.

Adeno-associated virus (AAV)-mediated factor IX Padua gene therapy has demonstrated good safety and efficacy in adult patients with hemophilia B, but its safety and efficacy in adolescent patients with hemophilia B remain unknown. Here we report a multicenter, single-arm, phase 1 study involving 11 adolescent participants (aged 12-18 years) with severe or moderately severe hemophilia B (factor IX coagulant activity (FIX:C) ≤2 IU dl-1) in China. All of them received the AAV gene therapy BBM-H901 at a dose of 5 × 1012 vector genomes per kilogram of body weight and were then enrolled in a 52-week follow-up. The primary endpoint was safety. No dose-limiting toxicity was observed throughout the study. The most common adverse events included elevation of white blood cell count, elevation of neutrophil count and rash associated with corticosteroid use. One serious adverse event and two grade 3 adverse events occurred. Abnormal liver function was observed in one participant, with elevated alanine aminotransferase (197.0 U l-1) and aspartate aminotransferase (75.0 U l-1) levels, which returned to normal 4 weeks after immunosuppression therapy. The key secondary endpoints included the mean (s.d.) FIX:C at week 52 following infusion, which was 41.8 (30.1) IU dl-1 (one-stage SynthASil method), and the mean annualized bleeding rate, which decreased from 13.9 to 0.5. In this study, we show that the gene therapy drug BBM-H901 for hemophilia B is safe in adolescent patients, providing initial insights into its efficacy. ClinicalTrials.gov identifier: NCT05709288 .

Journal Article↗

PubMed2026

Modified Plasmids and Inverted Terminal Repeats Enhance Adeno-Associated Virus Production and Performance.

Recombinant adeno-associated virus (rAAV) is a preferred vector in gene therapy, although high production costs inhibit widespread adoption. The most common approach for rAAV production involves transfection of HEK293 cells with three plasmids: pTransgene, pRep/Cap and pHelper. Producing sufficient amounts of these plasmids accounts for up to 40% of total batch costs. Initially, this work aimed to increase plasmid yields by replacing the backbones. While this approach increased pHelper yields, pRep/Cap and pTransgene yields were unaffected. A possible reason was identified: pTransgene contains inverted terminal repeat (ITR) sequences that are essential for rAAV production. ITRs have strong secondary structures (including hairpin loops termed B and C arms) that likely interfere with plasmid production. Therefore, targeted deletions were performed within the ITRs. Partial deletions in both the B and C arms of the ITR were most beneficial, as both plasmid yield and transgene expression increased. Importantly, partial deletions did not reduce rAAV yield, as had been previously observed when the B and C arms were fully deleted. In summary, we report a 140% increase in pHelper plasmid production, while the most successful ITR variant increased pTransgene plasmid yields by 57% and transgene expression by 28%, without reducing rAAV yields or transduction efficiency.

Humans↗

PubMed2026

Cerebrospinal delivery of a bidirectional AAV9 vector improves optic nerve and retinal pathology in a sheep model of Tay-Sachs disease.

Tay-Sachs disease (TSD) is a fatal neurodegenerative lysosomal storage disease. The Jacob sheep is the only large-animal model of TSD, yet ocular pathology and the therapeutic potential of gene therapy remain poorly defined. Sheep cohorts included normal controls (n = 3); untreated TSD-affected (n = 4); intravenous AAV9-Bic_HexA/HexB-treated (n = 3); and intracerebroventricular, cisterna magna, and lumbar intrathecal AAV9- Bic_HexA/HexB-treated sheep (cerebrospinal fluid [CSF] therapy; n = 7). Retinal histopathology and immunohistochemistry, retinal whole-mount analyses for retinal ganglion cell (RGC) morphology and density, optic nerve evaluation with p-phenylenediamine (PPD )semi-thin sections, qPCR assessment for vector genomes, and RNAscope probes for transgene expression were performed. Untreated TSD sheep exhibited RGCs with abundant microvesicular cytoplasmic expansion and optic nerve spheroids, with storage material variably staining with periodic acid-Schiff. Marked astrocytosis, microgliosis, and GM2 accumulation within RGCs were present. Optic nerve axon counts and RGC density were significantly reduced, and optic nerve damage scores increased, in untreated and IV-treated sheep but were rescued with short-term CSF therapy. GM2 volume and signal intensity per RGC were significantly reduced following short-term CSF therapy. Minimal but detectable retinal vector genomes and transgene expression were observed. These findings demonstrate retinal and optic nerve pathology in Jacob sheep with TSD and AAV9 therapy.

Animals↗

PubMed2024

Intrahepatic Exhausted Antiviral Immunity in an Immunocompetent Mouse Model of Chronic Hepatitis B.

BACKGROUND & AIMS: Targeting exhausted immune systems would be a promising therapeutic strategy to achieve a functional cure for HBV infection in patients with chronic hepatitis B (CHB). However, animal models recapitulating the immunokinetics of CHB are very limited. We aimed to develop an immunocompetent mouse model of CHB for intrahepatic immune profiling. METHODS: CHB mice were created by intrahepatic delivery of the Sleeping Beauty transposon vector tandemly expressing the hepatitis B virus (HBV) genome and fumarylacetoacetate hydrolase (FAH) cDNA into C57BL/6J congenic FAH knockout mice via hydrodynamic tail vein injection. We profiled the viral and intrahepatic immune kinetics in CHB mice with or without treatment with recombinant IFNα or the hepatotropic Toll-like receptor 7 agonist SA-5 using single-cell RNA-seq. RESULTS: CHB mice exhibited sustained HBV viremia and persistent hepatitis. They showed intrahepatic expansion of exhausted CD8+ T (Tex) cells, the frequency of which was positively associated with viral load. Recruited macrophages increased in number but impaired inflammatory responses in the liver. The cytotoxicity of mature natural killer (NK) cells also increased in CHB mice. IFNα and SA-5 treatment both resulted in viral suppression with mild hepatic flares in CHB mice. Although both treatments activated NK cells, SA-5 had the capacity to revitalize the impaired function of Tex cells and liver-recruited macrophages. CONCLUSIONS: Our novel CHB mouse model recapitulated the intrahepatic exhausted antiviral immunity in patients with CHB, which might be able to be reinvigorated by a hepatotropic TLR7 agonist.

Animals↗

PubMed2025

A suite of enhancer AAVs and transgenic mouse lines for genetic access to cortical cell types.

The mammalian cortex is comprised of cells classified into types according to shared properties. Defining the contribution of each cell type to the processes guided by the cortex is essential for understanding its function in health and disease. We use transcriptomic and epigenomic cortical cell-type taxonomies from mouse and human to define marker genes and putative enhancers and create a large toolkit of transgenic lines and enhancer adeno-associated viruses (AAVs) for selective targeting of cortical cell populations. We report creation and evaluation of fifteen transgenic driver lines, two reporter lines, and >1,000 different enhancer AAV vectors covering most subclasses of cortical cells. The tools reported here have been made publicly available, and along with the scaled process of tool creation, evaluation, and modification, they will enable diverse experimental strategies toward understanding mammalian cortex and brain function.

Animals↗

PubMed2026

A randomized trial of viral vector and adjuvanted protein HBV therapeutic vaccine in people with chronic hepatitis B on nucleos(t)ide analogs.

BACKGROUND: This study assessed the safety, efficacy, and immunogenicity of a therapeutic immunization strategy aimed at reaching a functional cure for chronic hepatitis B (CHB), relying on a heterologous prime-boost with viral vectors ChAd155-hIi-HBV and MVA-HBV, combined with sequential or concomitant administration of adjuvanted recombinant HBV proteins (HBc-HBs/AS01B). METHODS: This single-blind, randomized, controlled, first-in-human, phase 1/2 trial enrolled adults aged 18-65 years with HBeAg-negative CHB, virally suppressed on nucleos(t)ide analogs (NAs), with HBsAg >50 IU/mL. Participants received NAs and the following regimens of 4 doses (8-week intervals): sequential administration of ChAd155-hIi-HBV, MVA-HBV, and 2 HBc-HBs/AS01B doses; co-administration of ChAd155-hIi-HBV+HBc-HBs/AS01B, followed by 3 co-administered MVA-HBV+HBc-HBs/AS01B doses; 4 HBc-HBs/AS01B doses; 2 placebo doses followed by ChAd155-hIi-HBV and MVA-HBV administered alone or with HBc-HBs/AS01B; or 4 placebo doses. Safety, efficacy (≥1-log decrease in quantitative (q)HBsAg or HBsAg loss 24 weeks post-dose 4 [day (D)337]), antibody, and T-cell responses were evaluated. RESULTS: In all, 134 participants were vaccinated. Grade 3 solicited adverse events (AEs) (median duration: 2-3 days) were more frequent after co-administration (systemic: 59.3%; administration-site: 33.3%) than sequential administration (systemic: 10.3%; administration-site: 12.8%) of high-dose viral vectors and proteins. No vaccine-related or fatal serious AEs were reported. After 4 doses, no participant had HBsAg loss or ≥1-log decrease in qHBsAg (D337 vs. D1). Co-administration induced the strongest anti-HBs response (73.7% achieved anti-HBs ≥10 mIU/mL 2 weeks post-dose 4 vs. 40.0% after sequential administration). Both sequential and co-administration induced HBc-specific CD4+ and CD8+ T-cell responses, with a prime-boost effect of the viral vectors. CONCLUSIONS: Heterologous prime-boost with ChAd155-hIi-HBV and MVA-HBV, combined with sequential or co-administration of HBc-HBs/AS01B, had an acceptable safety profile, were moderately immunogenic, but no participants showed the expected efficacy outcome.

Humans↗

PubMed1978

Construction and properties of recombinant plasmids containing the rII genes of bacteriophage T4.

The EcoRI digestion products of phage T4 DNA have been examined using a phage DNA transformation assay. A 2.6 X 10(6) Dalton fragment was found to contain the rII genes. This fragment was purified and then treated with HindIII endonuclease. The cleavage products were ligated to the vector plasmid pBR313 and viable recombinant plasmids recovered. A genetic assay was employed to demonstrate that the recombinants contained T4 DNA and to localize on the phage genetic map the EcoRI and HindIII sites cleaved during the construction of the plasmids. Preliminary characterization suggests that a fragment covering the beginning of the rIIA gene possibly contains a promotor which is active in uninfected cells.

Cell Transformation, Viral↗

PubMed2026

Optimized AAV5-RPGR ORF15 Gene Therapy Rescues Photoreceptor Structure and Function in X-Linked Retinitis Pigmentosa Mouse Model.

PURPOSE: To develop and evaluate an rAAV5-based gene therapy vector expressing an optimized human RPGR ORF15 transgene (rAAV5-RPGR) for the treatment of X-linked retinitis pigmentosa caused by RPGR mutations, addressing the challenges of cloning the unstable wild-type ORF15 sequence. DESIGN: This was a prospective experimental study. SUBJECTS: This was an animal study. METHODS: An optimized RPGR ORF15 sequence was designed to eliminate problematic secondary structures and cryptic splice sites. In vitro expression was validated in HEK 293T and photoreceptor-like 661 W cells. A complete Rpgr knockout mouse model (Rpgr-knockout [KO]) was generated and characterized phenotypically. Therapeutic efficacy was assessed in Rpgr-KO mice via subretinal injection of rAAV5-RPGR at low (1 × 10⁹ vg/eye), medium (3 × 10⁹ vg/eye), or high (1 × 10¹⁰ vg/eye) doses. Structural and functional outcomes were evaluated at 12- and 14-month postinjection. Short-term safety was assessed in rabbits 1 month after subretinal injection. MAIN OUTCOME MEASURES: Level of RPGR protein expression and Protein isoform profile (elimination of truncated isoforms), Cellular localization of transgene expression and Dose-dependence of expression, outer nuclear layer thickness, and electroretinography parameters. RESULTS: (1) The optimized vector increased RPGR protein expression 3.3-fold in vitro compared to wild-type and eliminated truncated isoforms. (2) Subretinal delivery of rAAV5-RPGR in mice demonstrated dose-dependent transgene expression localized correctly to photoreceptor inner segments. (3) In Rpgr-KO mice, high-dose treatment significantly preserved outer nuclear layer thickness at the injection site (42% greater than controls at 14 months, P < .01) and central retina (P < .05), reduced aberrant rhodopsin mislocalization (P < .01), and partially restored retinal function. ERG showed significantly improved scotopic a-wave (≥100 vs <90 µV in controls at 10 cd·s/m²) and photopic b-wave amplitudes (49-66 vs 31-46 µV at 30 cd·s/m²) in treated mice. (4) No vector-related toxicity was observed in rabbits. CONCLUSIONS: rAAV5-RPGR mediated efficiently, targeted expression of optimized RPGR-ORF15, significantly preserved photoreceptor structure and function in a severe X-linked retinitis pigmentosa mouse model, and demonstrated a favorable safety profile. This study provides preclinical proof-of-concept for RPGR-targeted gene replacement therapy.

Animals↗

PubMed1979

Interaction of inflammatory cells and oral microorganisms. VIII. Detection of leukotoxic activity of a plaque-derived gram-negative microorganism.

In the present study we identified a gram-negative anaerobic rod referred to as Y4 which was cytotoxic for human polymorphonuclear leukocytes. Y4 was isolated from dental plaque of a patient with juvenile periodontitis and presented most of the taxonomic characteristics of Actinobacillus species. Under experimental conditions, viable Y4 were cytotoxic for human peripheral blood polymorphonuclear leukocytes in serum-free cultures. Cytotoxicity was dependent on bacterial concentrations and was enhanced in the presence of a fresh or heat-inactivated (56 degrees C, 30 min) autologous serum. Leukotoxicity was independent of phagocytosis. Y4 leukotoxic effect was abolished when bacteria were heat treated (56 degrees C, 30 min) or when incubations were carried out at 4 degrees C instead of at 37 degrees C. The leukotoxicity was monitored by electron microscopy and biochemically by measuring lactate dehydrogenase indicator of cell viability. No cytotoxic effects of Y4 on human mononuclear cells, chicken fibroblasts, or mouse macrophages were detected under the conditions studied. Polymorphonuclear leukocytes may play an important role in the host defense against bacteria in periodontal disease. The cytotoxic effect of Y4 for polymorphonuclear leukocytes presented in this study is the first report of a direct offensive microbial vector in a plaque-derived microorganism and may prove to be relevant in the pathogenesis of juvenile periodontitis.

Dental Plaque↗

PubMed2016

In Vivo Selection Yields AAV-B1 Capsid for Central Nervous System and Muscle Gene Therapy.

Adeno-associated viral (AAV) vectors have shown promise as a platform for gene therapy of neurological disorders. Achieving global gene delivery to the central nervous system (CNS) is key for development of effective therapies for many of these diseases. Here we report the isolation of a novel CNS tropic AAV capsid, AAV-B1, after a single round of in vivo selection from an AAV capsid library. Systemic injection of AAV-B1 vector in adult mice and cat resulted in widespread gene transfer throughout the CNS with transduction of multiple neuronal subpopulations. In addition, AAV-B1 transduces muscle, β-cells, pulmonary alveoli, and retinal vasculature at high efficiency. This vector is more efficient than AAV9 for gene delivery to mouse brain, spinal cord, muscle, pancreas, and lung. Together with reduced sensitivity to neutralization by antibodies in pooled human sera, the broad transduction profile of AAV-B1 represents an important improvement over AAV9 for CNS gene therapy.

Animals↗

PubMed1978

Molecular analysis of cloned human 18S ribosomal DNA segments.

A fraction of DNA from the human fetal lung fibroblast line IMR-90, 30-fold enriched for ribosomal DNA, was cloned in the lambda phage vector Charon 16A. Of 978 clones assayed by hybridization to a mixture of 125I-labeled 18S and 28S ribosomal RNA, 11 recombinants containing a 3.8-megadalton segment of human 18S ribosomal DNA were identified. Restriction endonuclease analysis of these clones demonstrated variation only in orientation of the human gene segment within the phage vector. Restriction sites that we had previously detected from analysis of restriction products of unfractionated human DNA by using the Southern transfer method were also present in the cloned DNA segment. Recombinant DNA technology thus provides a valid and efficient means to define structural conservation or variation within families of human genes.

Cell Line↗

PubMed2016

Immunodominant Dengue Virus-Specific CD8+ T Cell Responses Are Associated with a Memory PD-1+ Phenotype.

UNLABELLED: Dengue disease is a large public health problem that mainly afflicts tropical and subtropical regions. Understanding of the correlates of protection against dengue virus (DENV) is poor and hinders the development of a successful human vaccine. The present study aims to define DENV-specific CD8(+)T cell responses in general and those of HLA alleles associated with dominant responses in particular. In human blood donors in Nicaragua, we observed a striking dominance of HLA B-restricted responses in general and of the allele B*35:01 in particular. Comparing these patterns to those in the general population of Sri Lanka, we found a strong correlation between restriction of the HLA allele and the breadth and magnitude of CD8(+)T cell responses, suggesting that HLA genes profoundly influence the nature of responses. The majority of gamma interferon (IFN-γ) responses were associated with effector memory phenotypes, which were also detected in non-B*35:01-expressing T cells. However, only the B*35:01 DENV-specific T cells were associated with marked expression of the programmed death 1 protein (PD-1). These cells did not coexpress other inhibitory receptors and were able to proliferate in response to DENV-specific stimulation. Thus, the expression of particular HLA class I alleles is a defining characteristic influencing the magnitude and breadth of CD8 responses, and a distinct, highly differentiated phenotype is specifically associated with dominant CD8(+)T cells. These results are of relevance for both vaccine design and the identification of robust correlates of protection in natural immunity. IMPORTANCE: Dengue is an increasingly significant public health problem as its mosquito vectors spread over greater areas; no vaccines against the virus have yet been approved. An important step toward vaccine development is defining protective immune responses; toward that end, we here characterize the phenotype of the immunodominant T cell responses. These DENV-reactive T cells express high levels of the receptor programmed death 1 protein (PD-1), while those from disease-susceptible alleles do not. Not only does this represent a possible correlate of immunodominance, but it raises the hypothesis that PD-1 might be a regulator that prevents excessive damage while preserving antiviral function. Further, as this study employs distinct populations (Nicaraguan and Sri Lankan donors), we also confirmed that this pattern holds despite geographic and ethnic differences. This finding indicates that HLA type is the major determinant in shaping T cell responses.

Alleles↗

PubMed2020

Battling Neurodegenerative Diseases with Adeno-Associated Virus-Based Approaches.

Neurodegenerative diseases (NDDs) are most commonly found in adults and remain essentially incurable. Gene therapy using AAV vectors is a rapidly-growing field of experimental medicine that holds promise for the treatment of NDDs. To date, the delivery of a therapeutic gene into target cells via AAV represents a major obstacle in the field. Ideally, transgenes should be delivered into the target cells specifically and efficiently, while promiscuous or off-target gene delivery should be minimized to avoid toxicity. In the pursuit of an ideal vehicle for NDD gene therapy, a broad variety of vector systems have been explored. Here we specifically outline the advantages of adeno-associated virus (AAV)-based vector systems for NDD therapy application. In contrast to many reviews on NDDs that can be found in the literature, this review is rather focused on AAV vector selection and their preclinical testing in experimental and preclinical NDD models. Preclinical and in vitro data reveal the strong potential of AAV for NDD-related diagnostics and therapeutic strategies.

Animals↗

PubMed2026

Distribution of transgene in the rodent choroid plexus after intracerebroventricular injection of adeno-associated virus.

Using adeno-associated virus to transfer genetic information to the choroid plexus has emerged as a promising route for long-term gene therapy in the brain for a variety of conditions. Overexpression of proteins has proved effective in small animal models but few attempts have been made to translate this technology to clinic, suppress the activity of a protein of interest, or further expand the limited capsid serotypes known to have choroid plexus tropism. We utilise transfer of green fluorescent protein to show choroid plexus epithelium tropism for novel AAV6 derived capsid ShH10Y445F in mouse, rat and porcine tissue explant cultures. In vivo tropism is shown in the mouse following stereotactic intracerebroventricular injection. We examined the distribution of viral transduction across the choroid plexus in all four ventricles following a single unilateral intracerebroventricular injection using both green fluorescent protein as a transgene, but also the CRISPR/Cas9 system to deliver permanent knockdown of apical water channel aquaporin-1. Quantitative immunofluorescence and SURVEYOR assay were used to statistically assess the magnitude and extent of choroid plexus knockdown across the ventricular system. We conclude that serotype ShH10Y445F targets choroid plexus epithelium in mouse, rat and pig; and when carrying the CRISPR/Cas9 system can reduce target protein expression in these cells. Transduced choroid plexus epithelial cells distribute unevenly with a bias toward the lateral ventricle on the injected side and a preferential infection of choroid plexus in the lateral over the third and fourth ventricles. Overcoming irregular distribution represents a challenge for clinical translation of this technology where clinical efficacy may require manipulation of the entire choroid plexus.

Animals↗

PubMed1979

Identification of the C. coli dnaK (groPC756) gene product.

The E. coli dnaK (groPC756) gene product is essential for bacteriophage lambda DNA replication. Bacterial DNA segments carrying this gene have been cloned onto a bacteriophage lambda vector. The product of the dnaK gene has been identified on SDS polyacrylamide gels after infection of UV-irradiated E. coli cells. The dnaK gene codes for a polypeptide with an apparent molecular weight of 93,000-Mr. Transducing phages carrying amber mutations in the dnaK gene fail to induce the synthesis of the 93,000-Mr polypeptide chain upon infection of sup+ bacteria, but do so upon infection of supF bacteria. E coli carrying the dnaK756 mutation are, in addition, temperature sensitive for growth at 43 degrees C. It is shown that the dnaK756 mutation results in an overproduction of the dnaK gene product at that temperature.

Bacterial Proteins↗

PubMed2025

Identification of potential biomarkers and mechanisms for keloid disorder based on comprehensive bioinformatics analysis and machine learning algorithms.

BACKGROUND: Keloid disorder (KD) encompasses a spectrum of fibroproliferative dermal conditions, the pathogenesis remains complex and incompletely understood. This study sought to identify biomarkers and potential therapeutic targets for KD through an integrative bioinformatics approach and machine learning analysis of RNA sequencing data. METHODS: RNA sequencing was performed on skin tissue samples from 13 patients with KD and 14 healthy controls. Using weighted gene co-expression network analysis and differential expression analysis revealed differentially expressed key module genes, and the CytoHubba plugin identified candidate genes. Subsequently analyzed using least absolute shrinkage and selection operator (LASSO) and support vector machine recursive feature elimination (SVM-RFE) methods to pinpoint feature genes associated with KD. Following this, biomarkers were determined through expression level validation, enrichment analysis, and immune infiltration analysis. RESULTS: A total of 420 differentially expressed key module genes were identified, and the top 10 genes with DMNC values were selected as candidate genes. Five feature genes were selected through LASSO and SVM-RFE, with NID2, MFAP2, COL8A1, and P4HA3 showing significant expression differences between KD and control samples, along with consistent expression patterns across datasets, identified as potential biomarkers. These four biomarkers were proved to possess high diagnostic potential, and they were found to exhibit significant positive correlations with one another. Functional enrichment analysis indicated that the primary KEGG pathways associated with these biomarkers included "steroid hormone biosynthesis" and "cytokine-cytokine receptor interaction." Moreover, immune infiltration analysis revealed that the four biomarkers were negatively correlated with type 17 T helper cells and positively correlated with 15 immune cell types, including activated B cells and central memory CD4 T cells. CONCLUSION: In conclusion, NID2, MFAP2, COL8A1, and P4HA3 were identified as key biomarkers for KD, offering new avenues for more targeted and effective diagnostic and therapeutic strategies for managing this condition.

Humans↗