Multi-omics study reveals different pathogenesis of the generation of skin lesions in SLE and IDLE patients.

Lupus erythematosus (LE) is a heterogeneous, antibody-mediated autoimmune disease. Isolate discoid LE (IDLE) and systematic LE (SLE) are traditionally regarded as the two ends of the spectrum, ranging from skin-limited damage to life-threatening multi-organ involvement. Both belong to LE, but IDLE and SLE differ in appearance of skin lesions, autoantibody panels, pathological changes, treatments, and immunopathogenesis. Is discoid lupus truly a form of LE or is it a completely separate entity? This question has not been fully elucidated. We compared the clinical data of IDLE and SLE from our center, applied multi-omics technology, such as immune repertoire sequencing, high-resolution HLA alleles sequencing and multi-spectrum pathological system to explore cellular and molecular phenotypes in skin and peripheral blood from LE patients. Based on the data from 136 LE patients from 8 hospitals in China, we observed higher damage scores and fewer LE specific autoantibodies in IDLE than SLE patients, more uCDR3 sharing between PBMCs and skin lesion from SLE than IDLE patients, elevated diversity of V-J recombination in IDLE skin lesion and SLE PBMCs, increased SHM frequency and class switch ratio in IDLE skin lesion, decreased SHM frequency but increased class switch ratio in SLE PBMCs, HLA-DRB1*03:01:01:01, HLA-B*58:01:01:01, HLA-C*03:02:02:01, and HLA-DQB1*02:01:01:01 positively associated with SLE patients, and expanded Tfh-like cells with ectopic germinal center structures in IDLE skin lesions. These findings suggest a significant difference in the immunopathogenesis of skin lesions between SLE and IDLE patients. SLE is a B cell-predominate systemic immune disorder, while IDLE appears limited to the skin. Our findings provide novel insights into the pathogenesis of IDLE and other types of LE, which may direct more accurate diagnosis and novel therapeutic strategies.

Systemic lupus erythematosus patients contain B-cell receptor repertoires sensitive to immunosuppressive drugs.

Immune repertoire (IR) during treatment may be a surrogate biomarker for disease response. Changes of the IR in systemic lupus erythematosus patients in response to immunosuppressive drugs were identified in ten SLE patients. Patients provided peripheral blood mononuclear cells at two time points for sequencing. They were divided into sensitive and nonsensitive groups by their clinical responses to immunosuppressive drugs. After treatment, the BCR expression significantly decreased in patients from the sensitive group while there was no change in patients from the nonsensitive group. IgM comprised a dominant portion of the BCR repertoire and increased slightly in all patients in the sensitive group but decreased in the nonsensitive group. IgA also exhibited opposing changes between the two groups. Shorter CDR3 of TRB and TRG chains occurred in the sensitive group. CDR3 length of IGK decreased significantly in the sensitive group. CDR3 of TCR δ/γ changed distinctly between time points in the sensitive group. Six immune-related genes showed differential expression levels in sensitive and nonsensitive groups. Our study shows that it is BCR repertoire sensitivity to immunosuppressive drugs in SLE patients and sheds light on personalized therapy for SLE.

Application of magnetic nanoparticles in nucleic acid detection.

Nucleic acid is the main material for storing, copying, and transmitting genetic information. Gene sequencing is of great significance in DNA damage research, gene therapy, mutation analysis, bacterial infection, drug development, and clinical diagnosis. Gene detection has a wide range of applications, such as environmental, biomedical, pharmaceutical, agriculture and forensic medicine to name a few. Compared with Sanger sequencing, high-throughput sequencing technology has the advantages of larger output, high resolution, and low cost which greatly promotes the application of sequencing technology in life science research. Magnetic nanoparticles, as an important part of nanomaterials, have been widely used in various applications because of their good dispersion, high surface area, low cost, easy separation in buffer systems and signal detection. Based on the above, the application of magnetic nanoparticles in nucleic acid detection was reviewed.

Topical application of a BCL-2 inhibitor ameliorates imiquimod-induced psoriasiform dermatitis by eliminating senescent cells.

BACKGROUND: Psoriasis is an inflammatory skin disease with unclear pathogenesis and unmet therapeutic needs. OBJECTIVE: To investigate the role of senescent CD4+ T cells in psoriatic lesion formation and explore the application of senolytics in treating psoriasis. METHODS: We explored the expression levels of p16INK4a and p21, classical markers of cellular senescence, in CD4+ T cells from human psoriatic lesions and imiquimod (IMQ)-induced psoriatic lesions. We prepared a senolytic gel using B-cell lymphoma 2 (BCL-2) inhibitor ABT-737 and evaluated its therapeutic efficacy in treating psoriasis. RESULTS: Using multispectrum immunohistochemistry (mIHC) staining, we detected increased expression levels of p16INK4a and p21 in CD4+ T cells from psoriatic lesions. After topical application of ABT-737 gel, significant alleviation of IMQ-induced psoriatic lesions was observed, with milder pathological alterations. Mechanistically, ABT-737 gel significantly decreased the percentage of senescent cells, expression of T cell receptor (TCR) α and ß chains, and expression of Tet methylcytosine dioxygenase 2 (Tet2) in IMQ-induced psoriatic lesions, as determined by mIHC, high-throughput sequencing of the TCR repertoire, and RT-qPCR, respectively. Furthermore, the severity of psoriatic lesions in CD4creTet2f/f mice was milder than that in Tet2f/f mice in the IMQ-induced psoriasis model. CONCLUSION: We revealed the roles of senescent CD4+ T cells in developing psoriasis and highlighted the therapeutic potential of topical ABT-737 gel in treating psoriasis through the elimination of senescent cells, modulation of the TCR αß repertoire, and regulation of the TET2-Th17 cell pathway.

Deep immunoglobulin repertoire sequencing depicts a comprehensive atlas of spike-specific antibody lineages shared among COVID-19 convalescents.

Neutralizing antibodies are a key component in protective humoral immunity against SARS-CoV-2. Currently, available technologies cannot track epitope-specific antibodies in global antibody repertoires. Thus, the comprehensive repertoire of spike-specific neutralizing antibodies elicited by SARS-CoV-2 infection is not fully understood. We therefore combined high-throughput immunoglobulin heavy chain (IgH) repertoire sequencing, and structural and bioinformatics analysis to establish an antibodyomics pipeline, which enables tracking spike-specific antibody lineages that target certain neutralizing epitopes. We mapped the neutralizing epitopes on the spike and determined the epitope-preferential antibody lineages. This analysis also revealed numerous overlaps between immunodominant neutralizing antibody-binding sites and mutation hotspots on spikes as observed so far in SARS-CoV-2 variants. By clustering 2677 spike-specific antibodies with 360 million IgH sequences that we sequenced, a total of 329 shared spike-specific antibody clonotypes were identified from 33 COVID-19 convalescents and 24 SARS-CoV-2-naïve individuals. Epitope mapping showed that the shared antibody responses target not only neutralizing epitopes on RBD and NTD but also non-neutralizing epitopes on S2. The immunodominance of neutralizing antibody response is determined by the occurrence of specific precursors in human naïve B-cell repertoires. We identified that only 28 out of the 329 shared spike-specific antibody clonotypes persisted for at least 12 months. Among them, long-lived IGHV3-53 antibodies are likely to evolve cross-reactivity to Omicron variants through accumulating somatic hypermutations. Altogether, we created a comprehensive atlas of spike-targeting antibody lineages in COVID-19 convalescents and antibody precursors in human naïve B cell repertoires, providing a valuable reference for future vaccine design and evaluation.

Epidemiological survey of bovine gammaherpesvirus 4 (BoHV-4) infection in cattle and buffalo from Pakistan.

This study was conducted to evaluate the prevalence of bovine gammaherpesvirus 4 (BoHV-4) among healthy cattle and buffaloes as well as those associated with different diseases (respiratory tract infection, mastitis and reproductive tract infection) in District Chakwal, Pakistan. Blood, swab and milk samples of cattle and buffaloes were randomly collected from different areas of Chakwal. DNA was isolated from the samples and subjected to nested PCR using thymidine kinase gene primers. Out of 300 samples (200 blood, 50 swab and 50 milk samples) from both species (cattle and buffalo), an overall prevalence of BoHV-4 of 3.33% was obtained. Samples from cattle showed a higher species-specific prevalence (4.16%) than samples from buffalo (2.78%). One sample out of 50 swab samples and 1 out of 50 milk samples were also positive for BoHV-4. DNA sequencing of a positive PCR product from cattle confirmed that the sequence was from the thymidine kinase gene of BoHV-4. Phylogenetic analysis also revealed close similarities with other BOHV-4 thymidine kinase sequences. To detect BoHV-4 antibodies, an indirect ELISA was also performed. Two hundred blood samples were also collected from the same animals in nonanticoagulant-containing tubes for the isolation of serum and were subjected to indirect ELISA. Sixteen samples (8%) were positive for BoHV-4 antibodies. This study will be useful in further diagnoses of BoHV-4 in Pakistan and in devising measures to control the spread of BoHV-4.

Naked-Eye Detection of Food-Borne Pathogens Using Multiplex Hyperbranched Rolling Circle Amplification and Magnetic Particles.

Food safety is a significant public health issue in both developed and developing countries. Previous detection methods struggle to meet the current demands. We have proposed a new way to detect pathogens, allowing detection to be visualized by the naked eye. Using our newly developed assay, when target genes are present in the reaction, corresponding padlock probes form closed-loop molecules. Each reaction tube contains a pair of universal primers for identifying target genes. The ring padlock probes and corresponding universal primers start hyperbranched rolling circle amplification (HRCA) under the action of the polymerase, so as to gain branched chain amplification products, which are irreversibly entangled with magnetic particles to form aggregated magnetic particle clusters, and the detection results are visible to naked eyes. On the contrary, by using linear probes, the clustering of magnetic particles will not be produced. This method was applied to the detection of five food-borne pathogens enterohemorrhagic Escherichia coli (EHEC), enterotoxigenic Escherichia coli (ETEC), enteropathogenic Escherichia coli (EPEC), enteroinvasive Escherichia coli (EIEC) and Escherichia coli (E. coli), with detection limits of 1 × 103, 1 × 104, 1 × 103, 1 × 104 and 1 × 102 CFU/mL, respectively. This method can realize multiplex automatic detection of nucleic acid and shows great development potential in the field of molecular diagnosis.

Immune repertoire analysis of normal Chinese donors at different ages.

OBJECTIVES: This study investigated the characteristics of the immune repertoire in normal Chinese individuals of different ages. MATERIALS AND METHODS: In this study, all seven receptor chains from both B and T cells in peripheral blood of 16 normal Chinese individuals from two age groups were analyzed using high-throughput sequencing and dimer-avoided multiplex PCR amplification. Normal in this study is defined as no chronic, infectious or autoimmune disease within 6 months prior to blood draw. RESULTS: We found that compared with the younger group, the clonal expression of T-cell receptor repertoire increased in the older group, while diversity decreased. In addition, we found that the T-cell receptor repertoire was more significantly affected by age than the B-cell receptor repertoire, including significant differences in the use of the unique TCR-alpha and TCR-beta V-J gene combinations, in the two groups of normal participants. We further analyzed the degree of complementarity determining region 3 sequence sharing between the two groups, and found shared TCR-alpha, TCR-gamma, immunoglobulin-kappa and immunoglobulin-lambda chain complementarity determining region 3 sequences in all subjects. CONCLUSION: Taken together, our study gives us a better understanding of the immune repertoire of different normal Chinese people, and these results can be applied to the treatment of age-related diseases. Immune repertoire analysis also allows us to observe participant's wellness, aiding in early-stage diagnosis.

The methods and advances of adaptive immune receptors repertoire sequencing.

The adaptive immune response is a powerful tool, capable of recognizing, binding to, and neutralizing a vast number of internal and external threats via T or B lymphatic receptors with widespread sets of antigen specificities. The emergence of high-throughput sequencing technology and bioinformatics provides opportunities for research in the fields of life sciences and medicine. The analysis and annotation for immune repertoire data can reveal biologically meaningful information, including immune prediction, target antigens, and effective evaluation. Continuous improvements of the immunological repertoire sequencing methods and analysis tools will help to minimize the experimental and calculation errors and realize the immunological information to meet the clinical requirements. That said, the clinical application of adaptive immune repertoire sequencing requires appropriate experimental methods and standard analytical tools. At the population cell level, we can acquire the overview of cell groups, but the information about a single cell is not obtained accurately. The information that is ignored may be crucial for understanding the heterogeneity of each cell, gene expression and drug response. The combination of high-throughput sequencing and single-cell technology allows us to obtain single-cell information with low-cost and high-throughput. In this review, we summarized the current methods and progress in this area.

Recent advances in biosensor for detection of lung cancer biomarkers.

Lung cancer is primary cancer threatening human life worldwide with the highest mortality rate. The early detection of lung cancer plays a critical role in the early diagnosis and subsequent treatment. However, the conventional methodologies limit the applications due to the low sensitivity, being expensive, and invasive procedure. Tumor markers as biochemical parameters can reflect cancer occurrence and progression, which show sensitivity, convenience, and low cost in developing biosensors, and act as good candidates for fabricating biosensors of detecting lung cancer. This review describes various biosensors (2013-2019) for detection of lung cancer biomarkers. Firstly, the various reported tumor markers of lung cancer are briefly described. Then, the advancements of designing biosensors for sensitive, stable, and selective identification of lung cancer biomarkers are systematically provided, with a specific focus on the main clinical biomarkers such as neuron-specific enolase (NSE), cytokeratin 19 fragment (CYFRA 21-1). Finally, the recent challenges and further opportunities for developing effective biosensors for early diagnosis of lung cancer are discussed.

Immunosensors Based on Nanomaterials for Detection of Tumor Markers.

Nanomaterials have been widely used to immobilize biomolecules, amplify the signals and concentrate the analytes for detection with good properties including large surface area, good adsorption capacity and high surface activity. In recent years, nanomaterials such as carbon nanomaterials, noble metal nanomaterials, polymers, are widely applied to research and develop immunosensors with high sensitivity and selectivity, which monitor the antigen-antibody reaction for the detection of tumor markers. This review provides an introduction of immunosensors and focuses on the design of electrochemical (EC) immunosensors, electrochemical luminscence (ECL) immunosensors and photoelectrochemical (PEC) immunosensors based on nanomaterials in nearly three years.